研究者総覧

北澤 理子 (キタザワ リコ)

  • 附属病院 准教授
Last Updated :2020/08/06

研究者情報

学位

  • 博士(医学)(神戸大学)

ホームページURL

J-Global ID

研究キーワード

  • 細胞生物学 破骨細胞形成 病理診断学   Cell Biology Osteoclastogenesis Diagnostic Pathology   

研究分野

  • ライフサイエンス / 実験病理学
  • ライフサイエンス / 人体病理学

経歴

  • 2014年04月 - 現在  愛媛大学附属病院病理診断科(病理部)特任教授・病理部長
  • 2013年01月 - 2014年03月  愛媛大学大学院医学系研究科分子病理学特任教授
  • 2012年04月 - 2012年12月  愛媛大学大学院医学系研究科分子病理学准教授
  • 2010年10月 - 2012年03月  神戸大学大学院医学研究科分子病理診断学准教授
  • 1996年09月 - 2010年09月  神戸大学 大学院 医学研究科 医科学 神戸大学 大学院医学研究科本務/講師
  • 1995年03月 - 1996年09月  神戸大学医学部病理学第二講座助手
  • 1994年08月 - 1995年02月  神戸大学大学院医学系研究科病理学第2講座非常勤講師
  • 1992年04月 - 1994年07月  ワシントン大学(セントルイス)内分泌内科ポスドク
  • 1988年04月 - 1992年03月  神戸大学大学院医学系研究科大学院生
  • 1987年06月 - 1988年03月  国立療養所兵庫中央病院内科医師
  • 1985年06月 - 1987年05月  神戸市立中央市民病院臨床研修医

所属学協会

  • 臨床細胞学会   日本組織細胞化学会   日本癌学会   米国骨代謝学会議(ASBMR)   日本骨代謝学会   日本内科学会   日本内分泌学会   日本病理学会   

研究活動情報

論文

  • Riko Kitazawa, Satomi Kinto-Shibahara, Ryuma Haraguchi, Yukihiro Kohara, Sohei Kitazawa
    Biochemical and biophysical research communications 515 2 268 - 274 2019年07月 [有り][無し]
     
    Receptor activator of NF-κB (RANK) expressed on osteoclasts and their precursors is a receptor for RANK ligand (RANKL). Signals transduced by RANKL-RANK interaction induce genes essential for the differentiation and function of osteoclasts. We have cloned a basic promoter region of the mouse RANK gene and have analyzed the transcription machinery by transcription factors such as PU.1 (-480), and MITF (-100). Here, we examined the regulatory mechanisms of RANK gene transcription through AP-1 binding site, agagctca (-240). RANK mRNA expression in pre-osteoclastic RAW264.7 cells was induced by Phorbol12-myristate13-acetate (PMA) and suppressed by protein kinase C (PKC) inhibitor calphostin C. In RAW264.7 cells, Fos knockdown by siRNA blocked the inducible effect of PMA on RANK expression. By EMSA, an oligonucleotide (-246/-238) showed DNA protein binding, the specificity of which was confirmed by block-shift assay with an anti-Fos antibody and by the addition of the excess of a cold consensus probe. Co-transfection with a Fos expression vector showed that Fos increased RANK promoter activity 6-fold in RAW264.7 cells, and the addition of PU.1 and MITF superinduced the activity more than twenty-fold by the addition of PU.1 and MITF. Mutagenesis of the putative AP-1 site (-240) blocked the inducible effect of Fos on promoter activity. Taken together, these results indicate that during the differentiation of bone marrow mono-nucleated cells into osteoclast precursors, RANK transcription is positively regulated by Fos/AP-1 through the binding element of its gene promoter, supporting the concept that Fos activation by continuous CSF-1 stimulation on macrophages triggers initial expression of RANK and, later, a positive feedback loop by RANKL-RANK interaction.
  • Emiri Watanabe, Masao Miyagawa, Teruyoshi Uetani, Masaki Kinoshita, Riko Kitazawa, Mie Kurata, Hayato Ishimura, Takuya Matsuda, Yuki Tanabe, Tomoyuki Kido, Teruhito Kido, Akira Kurata, Teruhito Mochizuki
    International journal of cardiology 283 171 - 177 2019年05月 [有り][無し]
     
    BACKGROUND: Direct evidence of inflammatory activity in the atria of patients with atrial fibrillation (AF) is scarce. We assessed the capability of positron-emission tomography/computed tomography (PET/CT) to diagnose AF based on fluorodeoxyglucose (FDG) uptake in the atrial wall. METHODS AND RESULTS: Among 8233 patients who underwent FDG-PET/CT as work-up for malignancies, we identified 180 consecutive patients with AF (2.2%). Of those, we selected 137 patients who had fasted >12 h before FDG injection for inclusion in the experimental group (88 men and 49 women; age: 72.7 ± 8.9 years). Controls were 62 age- and sex-matched patients without AF. For visual analysis, we used a 4-point grading system. For quantitative analysis, we used the maximum standard uptake value (SUVmax) in the left (LA) and right atrial (RA) myocardium and the target-to-background ratio (TBR) of SUVmax to blood pool activity. The sensitivity, specificity, and positive-predictive value for detecting AF visually were 54.0%, 95.2%, and 96.1%, respectively; for quantitative analysis, the respective values were 65.7%, 75.8%, and 85.7%. Multivariable analysis of 11 clinical and imaging variables showed significant associations with RA SUVmax (odds ratio [OR]: 14.353, P = 0.026) and LA volume (OR: 1.371, P = 0.0001). The RA TBR was greater in cases with persistent AF than in those with paroxysmal AF (P < 0.0001). Pathological investigation of 4 autopsy hearts confirmed infiltration of extravascular macrophages and lymphocytes in the regions with FDG uptake. CONCLUSIONS: Higher atrial FDG uptake was associated with AF. PET/CT could be a useful tool for detecting local inflammation in the atria with AF.
  • Masaki T, Tsujimoto M, Kitazawa R, Nakano E, Funasaka Y, Ichihashi M, Kitazawa S, Kakita A, Kanda F, Nishigori C
    JAAD case reports 5 3 205 - 208 2019年03月 [有り][無し]
  • Fujibuchi T, Miyawaki J, Kidani T, Imai H, Kiyomatsu H, Kitazawa R, Miura H
    BMC cancer 19 1 116  2019年02月 [有り][無し]
  • Tanigawa K, Maekawa M, Kiyoi T, Nakayama J, Kitazawa R, Kitazawa S, Semba K, Taguchi T, Akita S, Yoshida M, Ishimaru K, Watanabe Y, Higashiyama S
    Journal of cellular physiology 2019年02月 [有り][無し]
  • Manami Mizumoto, Fumihiro Ochi, Toshihiro Jogamoto, Kentaro Okamoto, Mitsumasa Fukuda, Toshifumi Yamauchi, Toyohisa Miyata, Ryo Tashiro, Mariko Eguchi, Riko Kitazawa, Eiichi Ishii
    Case reports in pediatrics 2019 5354074 - 5354074 2019年 [有り][無し]
     
    Background: Nonocclusive mesenteric ischemia (NOMI) defines acute mesenteric ischemia without occlusion of the mesenteric arteries. The most common cause of NOMI is vasoconstriction or vasospasm of a mesenteric artery. NOMI generally affects patients >50 years of age, and few cases have been reported in children. Case Presentation: A 15-year-old boy with severe neurodevelopmental disability developed sudden-onset fever, abdominal distention, and dyspnea. Laboratory and radiological findings indicated acute intestinal obstruction and prerenal failure. He developed transient cardiopulmonary arrest and hypovolemic shock. Emergent laparotomy was performed, which revealed segmentally necrotic intestine from the jejunum to the ascending colon with pulsation of peripheral intestinal arteries, leading to a diagnosis of NOMI. The necrotic intestine was resected, and stomas were created. He was discharged on postoperative day 334 with short bowel syndrome as a complication. Conclusions: NOMI should be considered a differential diagnosis for intestinal symptoms with severe general conditions in both adults and children with underlying disease. Immediate surgical exploration is essential with NOMI to save a patient's life.
  • Wei J, Gan Y, Peng D, Jiang X, Kitazawa R, Xiang Y, Dai Y, Tang Y, Yang J
    Cancer medicine 2018年11月 [有り][無し]
  • Janecki DM, Sajek M, Smialek MJ, Kotecki M, Ginter-Matuszewska B, Kuczynska B, Spik A, Kolanowski T, Kitazawa R, Kurpisz M, Jaruzelska J
    Oncotarget 9 65 32466 - 32477 2018年08月 [有り][無し]
  • Sohei Kitazawa, Ryuma Haraguchi, Riko Kitazawa
    Histochemistry and cell biology 150 1 3 - 12 2018年07月 [有り][無し]
     
    Cytosine methylation plays a major role in the regulation of sequential and tissue-specific expression of genes. De novo aberrant DNA methylation and demethylation are also crucial processes in tumorigenesis and tumor progression. The mechanisms of how and when such aberrant methylation and demethylation occur in tumor cells are still obscure, however. To evaluate subtle epigenetic alteration among minor subclonal populations, morphology-oriented epigenetic analysis is requisite, especially where heterogeneity and flexibility are as notable as in the process of cancer progression and cellular differentiation at critical stages. Therefore, establishment of reliable morphology-oriented epigenetic studies has become increasingly important in not only the experimental but also the diagnostic field. By selecting a subset of cells based on characteristic morphological features disclosed by microdissection or in situ hybridization, we discovered how methylation at certain CpG sites outside of CpG islands would play a crucial epigenetic role in the versatility and flexibility of gene expression during cancer progression. In this review, we first introduce technical aspects of two morphology-oriented epigenetic studies: (1) histoendonuclease-linked detection of methylated sites of DNA (HELMET), and (2) padlock probe and rolling circle amplification (RCA) for in situ identification of methylated cytosine in a sequence-dependent manner. We then present our observation of a novel MeCP2-mediated gene-silencing mechanism through the addition of methylation to a single-CpG-locus upstream of the TATA-box of the receptor activator of NF-κB ligand (RANKL) and of secreted frizzled-related protein 4 (SFRP4) gene promoters.
  • Inoue A, Ohnishi T, Kohno S, Ohue S, Iwata S, Matsumoto S, Nishikawa M, Ozaki S, Nakamura Y, Mizuno Y, Kitazawa R, Kunieda T
    Neurosurgical review 41 2 557 - 565 2018年04月 [有り][無し]
  • Kusakabe K, Kohno S, Inoue A, Seno T, Yonezawa S, Moritani K, Mizuno Y, Kurata M, Kitazawa R, Tauchi H, Watanabe H, Iwata S, Hirato J, Kunieda T
    Neuropathology : official journal of the Japanese Society of Neuropathology 38 2 179 - 184 2018年04月 [有り][無し]
  • Ryuma Haraguchi, Riko Kitazawa, Yuuki Imai, Sohei Kitazawa
    Histochemistry and cell biology 149 4 365 - 373 2018年04月 [有り][無し]
     
    Longitudinal bone growth progresses by continuous bone replacement of epiphyseal cartilaginous tissue, known as "growth plate", produced by columnar proliferated- and differentiated-epiphyseal chondrocytes. The endochondral ossification process at the growth plate is governed by paracrine signals secreted from terminally differentiated chondrocytes (hypertrophic chondrocytes), and hedgehog signaling is one of the best known regulatory signaling pathways in this process. Here, to investigate the developmental relationship between longitudinal endochondral bone formation and osteogenic progenitors under the influence of hedgehog signaling at the growth plate, genetic lineage tracing was carried out with the use of Gli1CreERT2 mice line to follow the fate of hedgehog-signal-responsive cells during endochondral bone formation. Gli1CreERT2 genetically labeled cells are detected in hypertrophic chondrocytes and osteo-progenitors at the chondro-osseous junction (COJ); these progeny then commit to the osteogenic lineage in periosteum, trabecular and cortical bone along the developing longitudinal axis. Furthermore, in ageing bone, where longitudinal bone growth ceases, hedgehog-signal responsiveness and its implication in osteogenic lineage commitment is significantly weakened. These results show, for the first time, evidence of the developmental contribution of endochondral progenitors under the influence of epiphyseal chondrocyte-derived secretory signals in longitudinally growing bone. This study provides a precise outline for assessing the skeletal lineage commitment of osteo-progenitors in response to growth-plate-derived regulatory signals during endochondral bone formation.
  • Riko Kitazawa, Ryuma Haraguchi, Mana Fukushima, Sohei Kitazawa
    Histochemistry and cell biology 149 4 405 - 415 2018年04月 [有り][無し]
     
    Hard tissue homeostasis is regulated by the balance between bone formation by osteoblasts and bone resorption by osteoclasts. This physiologic process allows adaptation to mechanical loading and calcium homeostasis. Under pathologic conditions, however, this process is ill-balanced resulting in either over-resorption or over-formation of hard tissue. Local over-resorption by osteoclasts is typically observed in osteolytic metastases of malignancies, autoimmune arthritis, and giant cell tumor of bone (GCTB). In tumor-related local osteolysis, tumor-derived osteoclast-activating factors induce bone resorption not by directly acting on osteoclasts but by indirectly upregulating receptor activator of NFκB ligand (RANKL) on osteoblastic cells. Similarly, synovial tissue in the autoimmune arthritis model does overexpress RANKL and contains numerous osteoclast precursors, and like a landing craft, when it comes in contact with eroded bone surfaces, osteoclast precursors are immediately polarized to become mature osteoclasts, inducing rapidly progressive bone destruction at a late stage of the disease. GCTB, on the other hand, is a common primary bone tumor, usually arising at the metaphysis of the long bone in young adults. After the discovery of RANKL, the concept of GCTB as a tumor of RANKL-expressing stromal cells was established, and comprehensive exosome studies finally disclosed the causative single-point mutation at histone H3.3 (H3F3A) in stromal cells. Thus, osteolytic lesions under various pathological conditions are ultimately attributable to the overexpression of RANKL, which opens up a common, practical and useful therapeutic target for diverse osteolytic conditions.
  • Saki Ito, Riko Kitazawa, Ryuma Haraguchi, Takeshi Kondo, Ayaka Ouchi, Yasuo Ueda, Sohei Kitazawa
    Diagnostic pathology 13 1 1 - 1 2018年01月 [有り][無し]
     
    BACKGROUND: A proper balance between the activator and the repressor form of GLI3, a zinc-finger transcription factor downstream of hedgehog signaling, is essential for proper development of various organs during development. Mutations in different domains of the GLI3 gene underlie several congenital diseases including Greig cephalopolysyndactyly syndrome (GCPS) and Pallister-Hall syndrome (PHS). CASE PRESENTATION: Here, we describe the case of an overlapped phenotype of these syndromes with agenesis of the gallbladder and the pancreas, bearing a c.2155 C > T novel likely pathogenic variant of GLI3 gene by missense point mutation causing p.P719S at the proteolytic cleavage site. CONCLUSIONS: Although agenesis of the gallbladder and the pancreas is uncommon in GLI3 morphopathy, a slight difference in the gradient or the balance between activator and repressor in this case may hinder sophisticated spatial and sequential hedgehog signaling that is essential for proper development of gallbladder and pancreas from endodermal buds.
  • Inoue A, Ohnishi T, Kohno S, Ohue S, Iwata S, Matsumoto S, Nishikawa M, Ozaki S, Mizuno Y, Kitazawa R, Kunieda T
    Neurosurgical review 41 1 197 - 206 2018年01月 [有り][無し]
  • Kusakabe K, Inoue A, Matsumoto S, Kurata M, Kitazawa R, Watanabe H, Kunieda T
    International journal of surgery case reports 48 95 - 100 2018年 [有り][無し]
  • Masahiro Nishikawa, Akihiro Inoue, Takanori Ohnishi, Shohei Kohno, Shiro Ohue, Shirabe Matsumoto, Satoshi Suehiro, Daisuke Yamashita, Saya Ozaki, Hideaki Watanabe, Hajime Yano, Hisaaki Takahashi, Riko Kitazawa, Junya Tanaka, Takeharu Kunieda
    Stem cells international 2018 5387041 - 5387041 2018年 [有り][無し]
     
    Glioblastoma multiforme (GBM) is the most aggressive malignant brain tumor and a subpopulation of glioma stem-like cells (GSCs) is likely responsible for the invariable recurrence following maximum resection and chemoradiotherapy. As most GSCs that are located in the perivascular and perinecrotic niches should be removed during tumor resection, it is very important to know where surviving GSCs are localized. Here, we investigated the existence and functions of GSCs in the tumor periphery, which is considered to constitute the invasion niche for GSCs in GBM, by analyzing expression of stem cell markers and stem cell-related molecules and measuring particular activities of cultured GSCs. In addition, the relationship between GSCs expressing particular stem cell markers and pathological features on MRI and prognosis in GBM patients was analyzed. We showed that GSCs that express high levels of CD44 are present in the tumor periphery. We also found that vascular endothelial growth factor (VEGF) is characteristically expressed at a high level in the tumor periphery. Cultured GSCs obtained from the tumor periphery were highly invasive and have enhanced migration phenotype, both of which were markedly inhibited by CD44 knockdown. Higher expression of CD44 in the tumor periphery than in the core was correlated with a highly invasive feature on MRI and was associated with early tumor progression and worse survival, whereas lower expression of CD44 in the tumor periphery corresponded to low invasion and was associated with longer survival. The low invasion type on MRI tended to show high levels of VEGF expression in the tumor periphery, thus presenting the tumor with high proliferative activity. These results imply the significance of GSCs with high levels of CD44 expression in the tumor periphery compared to the core, not only in tumor invasion but also rapid tumor progression and short survival in patients with GBM.
  • Tomoki Shinohara, Akihiro Inoue, Shohei Kohno, Yasuo Ueda, Satoshi Suehiro, Shirabe Matsumoto, Masahiro Nishikawa, Saya Ozaki, Seiji Shigekawa, Hideaki Watanabe, Riko Kitazawa, Takeharu Kunieda
    Surgical neurology international 9 226 - 226 2018年 [有り][無し]
     
    Background: Chordoid glioma of the third ventricle is a rare neuroepithelial tumor characterized by a unique histomorphology within the third ventricular region, but with radiological and histopathological features mimicking benign lesions such as meningioma. We report a case of chordoid glioma of the third ventricle and suggest a useful indicator for accurate diagnosis. Case Description: A previously healthy 46-year-old woman was admitted to our hospital with mild headache. Neuroimaging revealed a large tumor measuring approximately 18 mm in the suprasellar region, and perifocal edema in the optic tract and internal capsule on magnetic resonance imaging. Laboratory findings revealed no pituitary dysfunction including diabetes insipidus. Gross total resection of the tumor was performed by the interhemispheric translamina terminalis approach. Histological findings revealed nests of regular epithelioid cells with large nuclei and abundant eosinophilic cytoplasm within myxoid stroma. Immunohistochemical studies demonstrated diffuse cytoplasmic expression of glial fibrillary acidic protein (GFAP) and CD34, and strong nuclear staining for thyroid transcription factor 1 (TTF-1). We, therefore, histologically classified the tumor as chordoid glioma of the third ventricle. Headache improved immediately postoperatively, and follow-up neuroimaging after 12 months showed no signs of recurrence. Conclusions: Chordoid glioma of the third ventricle is a very rare tumor that is difficult to diagnose on routine neuroimaging. Accurate diagnosis requires detailed analysis of neuroimaging and immunohistochemical studies using CD34 and TTF-1 staining.
  • Nishi Y, Kitazawa R, Haraguchi R, Ouchi A, Ueda Y, Kamaoka Y, Yamamoto K, Todo Y, Miyaoka H, Kitazawa S
    Case reports in oncology 10 2 508 - 514 2017年05月 [有り][無し]
  • Ryuma Haraguchi, Riko Kitazawa, Aki Murashima, Gen Yamada, Sohei Kitazawa
    ACTA HISTOCHEMICA ET CYTOCHEMICA 50 4 127 - 133 2017年 [有り][無し]
     研究論文(学術雑誌) 
    In mammals, the mullerian duct (MD) is an embryonic tubular structure that gives rise to the female reproductive tract (FRT). The MD originates from the coelomic epithelium (CoE) and takes on a rostral to caudal shape to establish the primary structure of the FRT under the regulation of morphogenetic signals. During these developmental processes, the MD and its derivatives require proper regulation of the Wnt-signaling-pathway. Here, to investigate the developmental contribution of FRT primordia under the influence of the Wnt-signaling, genetic lineage tracing was carried out using TopCreER/Rosa-LacZ mice to follow the fate of Wnt-signal-responsive cells during reproductive tract formation. TopCreER-marked-LacZ+ cells, arising from the Wnt-signal-responsive progenitors in CoE, give rise to spatially restricted MD and the uterine luminal epithelium. Similarly, the progeny from LacZ+ mesenchymal cells surrounding the MD contribute to both the uterine smooth muscle and stroma. Furthermore, in males, the Wnt-signal-responsive MD mesenchyme develops into the epididymis. These results show, for the first time, evidence of the sequential involvement of reproductive tract progenitors under the influence of Wnt-signal throughout the developmental term. This study provides a precise outline for assessing the lineage relation between the reproductive tract and the cell fate of its primordia in a temporally regulated manner.
  • Tomoya Onishi, Yutaka Yanagihara, Tadahiko Kikugawa, Noriyoshi Miura, Terutaka Noda, Toshio Kakuda, Riko Kitazawa, Nozomu Tanji
    World journal of surgical oncology 14 1 176 - 176 2016年07月 [有り][無し]
     
    BACKGROUND: Leiomyosarcomas typically originate in smooth muscle cell. Leiomyosarcoma potentially arising from the adrenal gland is an extremely rare mesenchymal tumors associated with delayed diagnosis and poor prognosis. CASE PRESENTATION: A 34-year-old man visited our department complaining of right hypochondriac pain. Computed tomography demonstrated a solid mass measuring 5.2 cm in diameter above the right kidney, corresponding to the right adrenal gland, and a lymph node mass, which appeared to have invaded the IVC wall. Right adrenalectomy and lymphadenectomy were performed. A microscopic examination revealed primary adrenal leiomyosarcoma with lymph node metastasis. No adjuvant therapy was performed, and the patient remains recurrence-free at 10 months postoperatively. CONCLUSIONS: We experienced a very rare case of primary adrenal leiomyosarcoma. Aggressive surgical resection including vascular reconstruction may be associated with improved survival.
  • Akihiro Inoue, Takanori Ohnishi, Shohei Kohno, Yosuke Mizuno, Riko Kitazawa, Yawara Nakamura, Shiro Ohue
    WORLD JOURNAL OF SURGICAL ONCOLOGY 14 1 152  2016年05月 [有り][無し]
     研究論文(学術雑誌) 
    Background: Multicentric gliomas are well-separated tumors in different locations of the brain, without anatomical continuity between lesions. We report a rare case of multicentric gliomas that occurred in both supra- and infratentorial regions with different histopathology. Case presentation: A 27-year-old man was admitted to our hospital with mild motor weakness of the right leg. Magnetic resonance imaging (MRI) showed a large tumor occupying the left insula, extending to the left basal ganglia, so tumor resection was performed. Histological diagnosis was diffuse astrocytoma. Tumor cells showed sporadic immunoreactivity for p53 and negative immunostaining for epidermal growth factor receptor (EGFR). Postoperative course was uneventful, and adjuvant therapy was not performed. At 7 months after surgery, MRI disclosed a left cerebellar tumor displaying an irregular ring formation on enhancement with gadolinium (Gd) and marked peritumoral edema. MRI studies including T2-weighted imaging demonstrated that this paravermian tumor had no contact with the initial left insular tumor. In addition, MRI studies of the whole neuraxis, cytological examination of the cerebrospinal fluid, and neurological findings demonstrated that no dissemination had occurred through the subarachnoid space or as intracerebral metastases. Therefore, the second surgery was performed. Histological diagnosis was glioblastoma. Immunohistochemistry revealed that most tumor cells were positively stained for both p53 and EGFR but negatively stained for isocitrate dehydrogenase 1 (IDH1). Conclusions: We reported a case of multicentric gliomas occurring in both supra-and infratentorial regions with different histopathology. Immunohistochemical examinations suggest that different genetic pathways may participate in the occurrence of these tumors.
  • Ryuma Haraguchi, Riko Kitazawa, Kiyoshi Mori, Ryosuke Tachibana, Hiroshi Kiyonari, Yuuki Imai, Takaya Abe, Sohei Kitazawa
    SCIENTIFIC REPORTS 6 25198  2016年04月 [有り][無し]
     研究論文(学術雑誌) 
    sFRP4 is an extracellular Wnt antagonist that fine-tunes its signal activity by direct binding to Wnts. Bone fragility under oxidative stress by diabetes and aging is partly related to the suppression of the Wnt signal through upregulated sFRP4. Here, to explore the functions of sFRP4 as a balancer molecule in bone development and remodeling, we analyzed the sFRP4 knock-in mouse strain. X-gal and immunohistochemically stained signals in sFRP4-LacZ heterozygous mice were detectable in restricted areas, mostly in osteoblasts and osteoclasts, of the femoral diaphysis after neonatal and postnatal stages. Histological and mu CT analyses showed increased trabecular bone mass with alteration of the Wnt signal and osteogenic activity in sFRP4 mutants; this augmented the effect of the buildup of trabecular bone during the ageing period. Our results indicate that sFRP4 plays a critical role in bone development and remodeling by regulating osteoblasts and osteoclasts, and that its functional loss prevents age-related bone loss in the trabecular bone area. These findings imply that sFRP4 functions as a key potential endogenous balancer of the Wnt signaling pathway by efficiently having direct influence on both bone formation and bone absorption during skeletal bone development and maintenance through remodeling.
  • Yong Wang, Yu Gan, Zhengyu Tan, Jun Zhou, Riko Kitazawa, Xianzhen Jiang, Yuxin Tang, Jianfu Yang
    ONCOTARGETS AND THERAPY 9 409 - 420 2016年 [有り][無し]
     研究論文(学術雑誌) 
    Human testis development-related gene 1 (TDRG1) is a recently identified gene that is expressed exclusively in the testes and promotes the development of testicular germ cell tumors. In this study, the role of TDRG1 in the development of testicular seminoma, which is the most common testicular germ cell tumor, was further investigated. Based on polymerase chain reaction, Western blotting, and immunohistochemistry tests, both gene and protein expression levels of TDRG1 were significantly upregulated in testicular seminoma tissues compared with normal testicular tissues. Additionally, the levels of phosphoinositide-3 kinase (PI3K)/p110 and Akt phosphorylation were dramatically upregulated in testicular seminoma tissues. Accordingly, in our cell experiment, seminoma TCam-2 cells were subjected to different treatments: the TDRG1 knockout, TDRG1 overexpression, PI3K inhibition (LY294002 administration), or PI3K activation (insulin-like growth factor-1 administration). Cell proliferation, the proliferation index, apoptosis rate, cell adhesive capacity, and cell invasion capability were assessed. Cells with both TDRG1 knockout and PI3K inhibition exhibited decreased cell proliferation, proliferation indexes, cell adhesion capacity, and cell invasion capability and increased apoptosis rates. Most of these effects were reversed by TDRG1 overexpression or PI3K activation, indicating that both TDRG1- and PI3K-mediated signaling promote proliferation and invasion of testicular seminoma cells. The knockout of TDRG1 significantly decreased the phosphorylation levels of PI3K/p85, PI3K/p110, Akt, and mammalian target of rapamycin (mTOR; Ser(2448)). Except for PI3K/p110, TDRG1 overexpression had the opposite effects on phosphorylation levels. Phosphorylated mTOR at Ser(2481) and Thr(2446) was not affected by TDRG1 or PI3K in our tests. Thus, these results indicate that TDRG1 promotes the development and migration of seminoma cells via the regulation of the PI3K/Akt/mTOR signaling pathway; this contributes to an understanding of the precise mechanisms underlying the development and migration of seminomas and lays a theoretical foundation for the development of appropriate therapies.
  • Yu Gan, Yong Wang, Zhengyu Tan, Jun Zhou, Riko Kitazawa, Xianzhen Jiang, Yuxin Tang, Jianfu Yang
    CANCER BIOLOGY & THERAPY 17 7 741 - 750 2016年 [有り][無し]
     研究論文(学術雑誌) 
    We previously identified TDRG1 (testis developmental related gene 1), a novel gene with exclusive expression in testis, promoted the proliferation and progression of cultured human seminoma cells through PI3K/Akt/mTOR signaling. As increasing evidence reveal that aberrant activation of this signaling is involved in cisplatin resistance. Then, in this study, we further explored whether TDRG1 regulated the chemosensitivity of seminoma TCam-2 cells to cisplatin. Our researches showed TDRG1 could regulate the viability of TCam-2 cells following cisplatin treatment in vitro through control of both cell apoptosis and cell cycle. Mechanistically, we observed TDRG1 positively regulated the expression levels of the key elements in PI3K/Akt/mTOR pathway including p-PI3K, p-Akt and p-mTOR and also affected the translocation of nuclear p-Akt in TCam-2 cells during cisplatin treatment. Meanwhile, the levels of Bad, cytochrome c, caspase-9 ratio (activated/total), caspase-3 ratio (activated/total) and cleaved-PARP were negatively modulated by TDRG1, which meant the involvement of mitochondria-mediated apoptotic pathway. Furthermore, we found the effect of TDRG1 knockdown or TDRG1 overexpression could be reversed by IGF-1, a PI3K signaling activator, or LY294002, a inhibitor of this pathway, respectively. Similar effects of TDRG1 on cisplatin chemosensitivity and associated molecular mechanism were also confirmed in vivo by employing xenograft assays. In addition, the positive correlation between TDRG1 and p-PI3K, or p-Akt, was found in tumor tissues from seminoma patients. In conclusion, we uncover that TDRG1 regulates chemosensitivity of TCam-2 cells to cisplatin through PI3K/Akt/mTOR signaling and mitochondria-mediated apoptotic pathway both in vitro and in vivo.
  • Matsubara E, Yamanouchi J, Kitazawa R, Azuma T, Fujiwara H, Hato T, Yasukawa M
    Case reports in hematology 2016 8751329  2016年 [有り][無し]
  • Inoue A, Watanabe H, Kumon Y, Ohtsuka Y, Tagawa M, Seyama T, Takechi A, Mizuno Y, Kitazawa R, Ohnishi T
    No shinkei geka. Neurological surgery 43 8 713 - 719 2015年08月 [有り][無し]
  • Akihiro Inoue, Takanori Ohnishi, Shohei Kohno, Yoshihiro Ohtsuka, Yawara Nakamura, Yosuke Mizuno, Riko Kitazawa, Shiro Ohue
    WORLD JOURNAL OF SURGICAL ONCOLOGY 13 226  2015年07月 [有り][無し]
     研究論文(学術雑誌) 
    Background: We present two rare cases of pineal-region meningiomas. These tumors are the first reported cases of dura-unrelated meningiomas originating from the arachnoid membrane over the vein of Galen (AMG). Case description: In Case 1, a 37-year-old woman presented with a progressing headache. Magnetic resonance imaging (MRI) showed a large tumor in the pineal region, displacing the vein of Galen upward. Angiography disclosed occlusion of the vein of Galen, with deep venous flow draining through the veins on the right medial surface of the occipital lobe to the superior sagittal sinus. In Case 2, a 67-year-old man presented with dizziness. MRI demonstrated a large mass in the pineal region, displacing the vein of Galen inferiorly. Angiography disclosed occlusion of the vein of Galen, with deep venous flow draining through the collateral venous channel into the transverse sinus. Both tumors were totally excised (Simpson Grade III for Case 1, Grade I for Case 2) via a left occipital transtentorial approach. No dural attachment was recognized in either case, but the tumor in Case 1 was firmly adherent to the inferior portion of the AMG, while that in Case 2 was attached to the superior portion of the AMG, but remained dissectible. Conclusions: We reported two cases of pineal-region meningiomas originating from the arachnoid membrane over the vein of Galen, resulting in meningioma without dural attachment. These tumors can be totally resected by careful dissection of the tumor from the arachnoid membrane surrounding the vein of Galen.
  • Ryuma Haraguchi, Riko Kitazawa, Sohei Kitazawa
    CELL AND TISSUE RESEARCH 359 2 503 - 512 2015年02月 [有り][無し]
     研究論文(学術雑誌) 
    Endochondral bone formation is tightly regulated by the spatial and sequential expression of a series of transcription factors. To disclose the roles of TBX18, a member of the T-box transcription factor family, during endochondral bone formation, its spatial and temporal expression patterns were characterized in the limb skeletal region of the developing mouse together with those of established osteochondrogenic markers Sox9, Col2a1, and Runx2. TBX18 expression first appeared in condensed mesenchymal cells (chondro-progenitors) in embryonic-day-10.5 (E10.5) limb bud and was co-localized with Sox9 expression, whereas at E11.5 and E12.5, it became undetectable in mesenchymal cells committed to the chondrocyte lineage. From E13.5 to E18.5, TBX18 expression reappeared in chondrocytes, correlating strongly with Col2a1 expression; furthermore, low level TBX18 expression was found in the Runx2-positive perichondral osteoblastic cell lineage. At the postnatal stage, TBX18 expression was observed in epiphyseal chondrocytes and osteocytes within the lacunae of mature trabecular bone. On the assumption that such characteristic Tbx18 gene expression is epigenetically regulated during mouse limb development, we examined the methylation status of the CpG-island in the mouse Tbx18 gene by methylation-specific polymerase chain reaction. Hypermethylation of the Tbx18 gene promoter became evident at an early embryonic stage in TBX18-negative cells and then disappeared at a late embryonic stage in TBX18-positive cells. Therefore, the temporal suppression of Tbx18 gene expression by the hypermethylation of its promoter seems to trigger the differentiation of mesenchymal cells into hypertrophic chondrocytes in the early stages of endochondral ossification.
  • Yuri Kameoka, Riko Kitazawa, Kanazu Ariasu, Ryosuke Tachibana, Yosuke Mizuno, Ryuma Haraguchi, Sohei Kitazawa
    ACTA HISTOCHEMICA ET CYTOCHEMICA 48 4 115 - 124 2015年 [有り][無し]
     研究論文(学術雑誌) 
    To explore the epigenetic mechanism that reactivates CDX2 (a homeobox transcription factor that serves as a tumor-suppressor gene) in intestinal-type gastric cancer during cancer progression, we examined the methylation status of the CDX2 gene promoter and the expression pattern of methyl-CpG binding protein-2 (MeCP2). From archives of the pathology records of surgically excised advanced stomach cancer cases in the Department of Molecular Pathology, Ehime University in a past decate (n=265), 10 cases of intestinal-type tubular adenocarcinoma, well-differentiated type (wel) with minor poorly-differentiated adenocarcinoma (por) components were selected. The expression pattern of CDX2, MUC2 and MeCP2 in these 10 cases was analyzed by immunohistochemistry. The cancerous and non-cancerous areas were selectively obtained by microdissection, and the methylation status of the CDX2 promoter of each area was assessed by methylation-specific polymerase chain reaction (MSP). In all 10 cases, CDX2 expression was clearly observed in the nucleus of the non-cancerous background of the intestinal metaplasic area, where the unmethylation pattern of the CDX2 gene promoter prevailed with reduced MeCP2 expression. In this metaplastic area, CDX2 expression was co-localized with its target gene, MUC2. CDX2 expression then disappeared from the deep invasive wel area. Reflecting the reduced CDX2 expression, microdissected samples from all the wel areas showed hypermethylation of the CDX2 gene promoter by MSP, with prominent MeCP2 expression. Interestingly, while hypermethylation of the CDX2 gene promoter was maintained in the por area in 8 of the 10 cases, CDX2 expression was restored in por areas where MeCP2 expression was markedly and selectively reduced. The other two cases, however, showed a constant MeCP2 expression level comparable to the surrounding deep invasive wel area with negative CDX2 expression. Therefore, gene silencing by hypermethylation may be overcome by the reduction of methyl-CpG binding proteins, resulting in apparent but non-functional reactivation of CDX2 as a mere molecular mark for gene silencing memory.
  • Masaru Kinomura, Noriaki Shimada, Mana Nishikawa, Kazuyoshi Omori, Tomoyasu Jo, Yasunori Ueda, Kenji Notohara, Riko Kitazawa, Sohei Kitazawa, Masaki Fukushima, Kenichiro Asano
    INTERNAL MEDICINE 54 23 3029 - 3033 2015年 [有り][無し]
     研究論文(学術雑誌) 
    A 68-year-old man was hospitalized and examined for renal impairment. A laboratory analysis showed hypercalcemia. Although the serum parathyroid hormone and serum 1-25(OH)(2) vitamin D3 levels were not elevated, the serum parathyroid hormone-related peptide (PTHrP) level was increased. Immunoelectrophoresis of the urine and bone marrow aspiration indicated multiple myeloma (MM). He was diagnosed with the coexistence of cast nephropathy and light chain deposition disease by a renal biopsy. Notably, PTHrP expression was detected in the myeloma cells based on immunohistochemistry and in situ hybridization. It is therefore important to examine the PTHrP concentration in MM patients with hypercalcemia.
  • Nakagawa M, Kitazawa R, Kondo T, Ninomiya K, Okita M, Haraguchi R, Kitazawa S
    Virchows Archiv : an international journal of pathology 465 3 253 - 256 2014年09月 [有り][無し]
     研究論文(学術雑誌) 
    Duodenal gastric heterotopia (DGH) is a benign asymptomatic condition assumed to be of congenital origin. Since DGH is often associated with fundic gland polyps (FGPs) that frequently carry a somatic beta-catenin gene mutation, we examined whether DGH, either sporadic or FGP-associated, is attributable to alterations of the Wnt/beta-catenin pathway. Genetic analysis revealed frequent somatic beta-catenin gene mutations in DGH; some of which showed the same mutation pattern as coexisting FGPs. All missense mutations were confined to codons 32, 33, and 37. No such mutations were observed, however, in any of the specimens from focal gastric foveolar metaplasia (GFM). Therefore, DGH is not a mere congenital lesion due to aberrant migration of normal gastric mucosa or a simple reactive metaplasia after regenerative stimuli of the duodenal mucosa, but a distinct condition based upon molecular genetic changes in the Wnt/beta-catenin pathway.
  • Akihiro Inoue, Takanori Ohnishi, Shohei Kohno, Riko Kitazawa, Yosuke Mizuno, Masahiro Nishikawa, Shiro Ohue
    CLINICAL NEUROLOGY AND NEUROSURGERY 124 138 - 141 2014年09月 [有り][無し]
     研究論文(学術雑誌)
  • Mori K, Kitazawa R, Kondo T, Mori M, Hamada Y, Nishida M, Minami Y, Haraguchi R, Takahashi Y, Kitazawa S
    PloS one 9 7 e102797  2014年07月 [有り][無し]
     研究論文(学術雑誌) 
    In diabetics, methylglyoxal (MG), a glucose-derived metabolite, plays a noxious role by inducing oxidative stress, which causes and exacerbates a series of complications including low-turnover osteoporosis. In the present study, while MG treatment of mouse bone marrow stroma-derived ST2 cells rapidly suppressed the expression of osteotrophic Wnt-targeted genes, including that of osteoprotegerin (OPG, a decoy receptor of the receptor activator of NF-kappaB ligand (RANKL)), it significantly enhanced that of secreted Frizzled-related protein 4 (sFRP-4, a soluble inhibitor of Wnts). On the assumption that upregulated sFRP-4 is a trigger that downregulates Wnt-related genes, we sought out the molecular mechanism whereby oxidative stress enhanced the sFRP-4 gene. Sodium bisulfite sequencing revealed that the sFRP-4 gene was highly methylated around the sFRP-4 gene basic promoter region, but was not altered by MG treatment. Electrophoretic gel motility shift assay showed that two continuous CpG loci located five bases upstream of the TATA-box were, when methylated, a target of methyl CpG binding protein 2 (MeCP2) that was sequestered upon induction of 8-hydroxy-2-deoxyguanosine, a biomarker of oxidative damage to DNA. These in vitro data suggest that MG-derived oxidative stress (not CpG demethylation) epigenetically and rapidly derepress sFRP-4 gene expression. We speculate that under persistent oxidative stress, as in diabetes and during aging, osteopenia and ultimately low-turnover osteoporosis become evident partly due to osteoblastic inactivation by suppressed Wnt signaling of mainly canonical pathways through the derepression of sFRP-4 gene expression.
  • Kentaro Nakai, Hideki Fujii, Keiji Kono, Shunsuke Goto, Riko Kitazawa, Sohei Kitazawa, Michinori Hirata, Masami Shinohara, Masafumi Fukagawa, Shinichi Nishi
    AMERICAN JOURNAL OF HYPERTENSION 27 4 586 - 595 2014年04月 [有り][無し]
     研究論文(学術雑誌) 
    BACKGROUND Diabetic nephropathy is a major risk of end-stage kidney disease. Many complex factors relate to the progression of diabetic nephropathy. Using nonobese type 2 diabetes model rats, we confirmed that oxidative stress was a crucial factor. Because recent studies suggest that vitamin D could suppress oxidative stress, we explored whether the active vitamin D analog, maxacalcitol, could also attenuate oxidative stress and prevent the progression of diabetic nephropathy. METHODS Diabetic rats aged 20 weeks were divided into 3 groups and treated with insulin, maxacalcitol, and vehicle. At age 30 weeks, blood and urine analyses, renal histology, immunohistochemistry, real-time polymerase chain reaction, and western blot were performed. RESULTS Although maxacalcitol reduced albuminuria and mesangial matrix expansion, no significant differences were observed in blood pressure and creatinine clearance among the 3 treatment groups. Systemic and intrarenal oxidative stress was reduced by maxacalcitol therapy. Expressions of nuclear factor-kappa B and nicotinamide adenine dinucleotide phosphate oxidase in the kidney also decreased in the insulin-treated and maxacalcitol-treated groups but increased in the vehicle-alone group. In addition, the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) decreased and Kelch-like erythroid cell-derived protein with CNC homology (ECH)-associated protein 1 (Keap1) increased in the vehicle-treated group; however, these expressions were restored in the maxacalcitol- and insulin-treated groups. CONCLUSIONS It is suggested that maxacalcitol attenuates the progression of diabetic nephropathy by suppression of oxidative stress and amelioration of the Nrf2-Keap1 pathway in nonobese type 2 diabetes without significant changes in blood pressure and glomerular filtration rate.
  • Y. Sakai, H. Hoshino, R. Kitazawa, M. Kobayashi
    ANDROLOGY 2 2 282 - 289 2014年03月 [有り][無し]
     研究論文(学術雑誌) 
    Seminoma, the most common testicular malignant neoplasm, originates from germ cells and is characterized by the presence of numerous tumour-infiltrating lymphocytes (TILs). Although it is widely accepted that TILs function in surveillance and cytotoxicity in various tumours including seminoma, detailed mechanisms governing TIL recruitment are not fully understood. It has been shown that high endothelial venule (HEV)-like vessels are induced in inflamed and neoplastic tissues and contribute to lymphocyte recruitment in a manner similar to the way physiological lymphocyte homing occurs in secondary lymphoid organs. Here, we report that HEV-like vessels, which express MECA-79(+) 6-sulfo sialyl Lewis X-capped structures, are induced in TIL aggregates in seminoma, and that such vessels potentially recruit circulating lymphocytes, as an E-selectin center dot IgM chimera bound these vessels in a calcium-dependent manner. These HEV-like vessels express intercellular adhesion molecule 1 (ICAM-1), but not vascular cell adhesion molecule 1 (VCAM-1) or mucosal addressin cell adhesion molecule 1 (MAdCAM-1), which likely contributes to lymphocyte firm attachment. We also found that the number of T cells attached to the luminal surface of HEV-like vessels was greater than the number of B cells (p<0.0001). Interestingly, while CD8(+) cytotoxic T lymphocytes (CTLs) attached to the lumen of HEV-like vessels were scarcely detected, significant numbers of proliferative CTLs were observed outside vessels. These histological findings strongly suggest that TILs, particularly T cells, are recruited to seminoma tissues via HEV-like vessels, and that tumour-infiltrating CTLs then undergo proliferation after transmigration through HEV-like vessels in testicular seminoma.
  • Koto Fujiishi, Riko Kitazawa, Yusa Nagai, Takafumi Watanabe, Kenji Bando, Shinji Kobayashi, Yoshihiro Yakushijin, Ryuma Haraguchi, Sohei Kitazawa
    VIRCHOWS ARCHIV 464 1 121 - 124 2014年01月 [有り][無し]
  • Fujii H, Kono K, Nakai K, Goto S, Kitazawa R, Fukagawa M, Nishi S
    Nephron extra 4 1 18 - 25 2014年01月 [有り][無し]
  • Takafumi Watanabe, Riko Kitazawa, Yosuke Mizuno, Natsumi Kuwahara, Chizu Ito, Atsuro Sugita, Ryuma Haraguchi, Sohei Kitazawa
    ACTA HISTOCHEMICA ET CYTOCHEMICA 47 3 125 - 131 2014年 [有り][無し]
     研究論文(学術雑誌) 
    Analysis of archival formalin-fixed, paraffin-embedded (FFPE) pathological specimens of three case of Epstein-Barr virus (EBV)-positive diffuse large B-cell lymphoma (DLBCL) and three cases of classical Hodgkin lymphoma (CHL) revealed that hypermethylation of the BOB. 1 gene promoter was exclusively observed in CHL. A discrepancy was observed, however, between the methylation status of the BOB. 1 gene promoter and its expression in the EBV-positive mixed cellular CHL (MCCHL). Since MCCHL lacks the typical B-cell phenotype even in the presence of abundant BOB. 1 transcription factors, functional activity of BOB. 1 may be lost or reduced by a mechanism other than epigenetic gene silencing. When some tumor-suppressor gene products have lost their biological function, impact or significance of derepression of such genes may be little. Therefore, when interpreting immunohistochemical results for diagnostic or research purposes, it must be borne in mind that apparent positive immunostaining can merely be the result of chromatin remodeling and that such transient expression often has little functional significance. Any apparent positive immunohistochemical result needs to be interpreted carefully with the help of the hypermethylation status as a molecular marker of gene silencing memory.
  • Munenori Komoda, Riko Kitazawa, Kenji Makita, Keisuke Yoshida, Miyuki Takeji, Yoshiko Soga, Mie Kurata, Ryuma Haraguchi, Sohei Kitazawa
    DIABETES RESEARCH AND CLINICAL PRACTICE 100 2 E59 - E62 2013年05月 [有り][無し]
     研究論文(学術雑誌) 
    Diabetes induces advanced glycation end products (AGEs) that per se are not only a major cause of oxidative stress but also reduce the plasticity of connective tissue by pathological collagen cross-linking. We describe a case of severe pulmonary hypertension manifesting as a major diabetic complication. Impaired pulmonary arteriolar plasticity attributed to pentosidine, together with increased circulation volume by hyperosmotic pressure and reduction in myocardial compliance by multiple patchy fibrosis, may contribute to the clinical manifestation of severe pulmonary hypertension. (C) 2013 Elsevier Ireland Ltd. All rights reserved.
  • S. C. McIver, K. L. Loveland, S. D. Roman, B. Nixon, R. Kitazawa, E. A. McLaughlin
    ANDROLOGY 1 3 517 - 529 2013年05月 [有り][無し]
     研究論文(学術雑誌) 
    Seminoma and non-seminoma tumours increasingly occur within the western population. These tumours originate from carcinoma in situ (CIS) cells, which arise from dysfunctional gonocytes. CXCL12 and its receptors, CXCR4 and CXCR7, have been implicated in migration, proliferation and survival of gonocytes and their precursors and progeny, primordial germ cells and spermatogonial stem cells respectively. We previously found evidence that several miRNA molecules predicted to modulate CXCR4 signalling are differentially expressed during the differentiation of gonocytes into spermatogonia in mice. Bioinformatic analysis predicted these miRNA to modulate CXCR4 signalling, leading us to hypothesize that CXCL12-mediated CXCR4 signalling is involved in the disrupted differentiation of gonocytes that underpins CIS formation. Indeed, we detected CXCL12 in Sertoli cells of normal human testis, and relatively high expression in tumour stroma with concomitant weak staining in dispersed tumour cells. In contrast, CXCR4 was expressed in spermatogonial and meiotic germ cells of normal testis and in the majority of tumour cells. Quantitative RT-PCR identified elevated CXCR4 transcript levels in seminoma compared with normal testis and to non-seminoma, potentially reflecting the higher proportion of dysfunctional germ cells within seminomas. In the normal testis, expression of CXCR4 downstream signalling molecules phospho-MEK1/2 and phospho-ERK1/2 correlated with CXCR4/CXCL12 expression. Strikingly, this correlation was absent in seminoma and non-seminoma samples, suggesting that CXCL12 signalling is disrupted. Proliferation rate and cell survival were not altered by CXCL12 in either seminoma (TCam-2) or non-seminoma (833ke) cell lines. However, CXCL12 exposure induced TCam-2 cell invasion though simulated basement membrane, while in contrast, we provide the novel evidence that CXCR4-expressing non-seminoma cell lines 833ke and NTera2/D1 do not invade in response to CXCL12. These findings indicate that CXCL12 expression in the human testis may selectively influence seminoma migration and metastasis, correlating with its importance in gonocyte and spermatogonial stem cell biology.
  • Sann Sanda Khin, Riko Kitazawa, Kyaw Htet, Hla Min Htike, Than Than Yee, Myint Aung, Ryuma Haraguchi, Sohei Kitazawa
    PATHOLOGY INTERNATIONAL 63 3 193 - 194 2013年03月 [有り][無し]
  • Natsumi Kuwahara, Riko Kitazawa, Koto Fujiishi, Yusa Nagai, Ryuma Haraguchi, Sohei Kitazawa
    World journal of gastroenterology 19 8 1314 - 7 2013年02月 [有り][無し]
     
    Gastritis cystica profunda (GCP) is a rare condition caused by ectopic entrapment of gastric glands, probably secondary to the disruption of muscularis mucosae. GCP is often associated with gastric adenocarcinoma, and loss of the KCNE2 subunit from potassium channel complexes is considered a common primary target molecule leads to both GCP and malignancy. In this study, we, for the first time, analyzed the expression of KCNE2 in surgically excised tissue from human gastric cancer associated with GCP and confirmed that reduced KCNE2 expression correlates with disease formation.
  • Kenji Makita, Riko Kitazawa, Shuho Semba, Koto Fujiishi, Miku Nakagawa, Ryuma Haraguchi, Sohei Kitazawa
    WORLD JOURNAL OF GASTROENTEROLOGY 19 4 536 - 541 2013年01月 [有り][無し]
     研究論文(学術雑誌) 
    AIM: To examine how the expression of caudal type homebox transcription factor 2 (Cdx2) is regulated in the development of malignancy in Barrett's esophagus. METHODS: Cdx2, mucin (MUC) series (MUC2, MUC5AC and MUC6), p53 and E-cadherin expression in Barrett's esophagus and adenocarcinoma specimens were examined by immunostaining. Isolated clusters of cells from (1) MUC2 and Cdx2-positive intestinal metaplastic mucosa; (2) MUC5AC and MUC6-positive, and MUC2 and Cdx2-negative high-grade dysplasia (HD), or intramucosal adenocarcinoma (IMC); and (3) MUC5AC, MUC6 and Cdx2-positive poorly-differentiated invasive adenocarcinoma (PDA) were analyzed by methylation-specific polymerase chain reaction using sets of primers for detecting methylation status of the Cdx2 gene. RESULTS: Most of the non-neoplastic Barrett's esophageal mucosa showing intestinal-type metaplasia with or without low-grade dysplasia was positive for E-cadherin, MUC series and Cdx2, but negative for p53. A portion of the low-grade to HD was positive for E-cadherin, MUC5AC, MUC6 and p53, but negative for MUC2 and Cdx2. The definite IMC area was strongly positive for MUC5AC, MUC6 and p53, but negative for MUC2 and Cdx2. Methylation of the Cdx2 promoter was not observed in intestinal metaplasia, while hypermethylation of part of its promoter was observed in hot dipped and IMC. Hypermethylation of a large fraction of the Cdx2 promoter was observed in PDA. CONCLUSION: Cdx2 expression is restored irrespective of the methylation status of its promoter. Apparent positive immunohistochemical results can be a molecular mark for gene silencing memory. (C) 2013 Baishideng. All rights reserved.
  • Kono K, Fujii H, Nakai K, Goto S, Kitazawa R, Kitazawa S, Shinohara M, Hirata M, Fukagawa M, Nishi S
    American journal of nephrology 37 2 167 - 174 2013年 [有り][無し]
  • Nakagawa M, Kitazawa R, Kuwahara N, Yoshida K, Haraguchi R, Kitazawa S
    Acta histochemica et cytochemica 46 1 19 - 24 2013年 [有り][無し]
     研究論文(学術雑誌) 
    Molecular genetic analyses of archival formalin-fixed, paraffin-embedded (FFPE) pathological specimens taken at biopsy or autopsy, are occasionally compromised because the DNA molecules therein are inevitably degraded. Furthermore, since these tissue samples comprise various cell types, the analyses based on mixtures of such heterogeneous populations often fail to reflect the nature of the affected cells. In the present study, to elucidate the contribution of beta-catenin gene mutation to the fundic gland polyp and the heterotopic gastric mucosa in the duodenum, we successfully introduced an agarose-bead mediated technique as an effectual tool for retrospective morphology-oriented genetic analyses. Microdissected samples were embedded in low-melting agarose, and directly treated with proteinase K. A fragment of the agarose-bead was used as a template for polymerase chain reaction to analyze beta-catenin mutation. Of the six cases of heterotopic gastric mucosa in the duodenum associated with fundic gland polyps, one showed a common 1-bp missense mutation at codon 37 shared by both the fundic gland polyp and the heterotopic gastric mucosa. Alternatively, a 1-bp silent mutation at codon 33 and missense mutation at codon 32 were identified only in the heterotopic gastric mucosa. Agarose-bead mediated technique shows superior sensitivity to the previously described techniques and is an effectual tool for retrospective morphology-oriented genetic analyses using a large number of archival pathological samples stored for long periods in the pathology laboratory.
  • Tomomi Sakuma, Tetsuya Nakamoto, Hiroaki Hemmi, Sohei Kitazawa, Riko Kitazawa, Takuya Notomi, Tadayoshi Hayata, Yoichi Ezura, Teruo Amagasa, Masaki Noda
    JOURNAL OF CELLULAR PHYSIOLOGY 227 7 2807 - 2812 2012年07月 [有り][無し]
     研究論文(学術雑誌) 
    Tumor metastasis to bone is a serious pathological situation that causes severe pain, and deterioration in locomoter function. However, the mechanisms underlying tumor metastasis is still incompletely understood. CIZ/NMP4 is a nucleocytoplasmic shuttling protein and its roles in tumor cells have not been known. We, therefore, hypothesized the role of CIZ/NMP4 in B16 melanoma cells that metastasize to bone. CIZ/NMP4 is expressed in B16 cells. The CIZ/NMP4 expression levels are correlated to the metastatic activity in divergent types of melanoma cells. Overexpression of CIZ/NMP4 increased B16 cell migration in Trans-well assay. Conversely, siRNA-based knockdown of CIZ/NMP4 suppressed migratory activity of these cells. As RANKL promotes metastasis of tumor cells in bone, we tested its effect on CIZ in melanoma cells. RANKL treatment enhanced CIZ/NMP4 expression. This increase of CIZ by RANKL promoted migration. Conversely, we identified CIZ/NMP4 binding site in the promoter of RANKL. Furthermore, luciferase assay indicated that CIZ/NMP4 overexpression enhanced RANKL promoter activities, revealing a positive feedback loop of CIZ/NMP4 and RANKL in melanoma. These observations indicate that CIZ/NMP4 is critical regulator of metastasis of melanoma cells. J. Cell. Physiol. 227: 28072812, 2012. (C) 2012 Wiley Periodicals, Inc.
  • Hitoshi Nishizawa, Genzo Iguchi, Ayumi Murawaki, Hidenori Fukuoka, Yoshitake Hayashi, Hidesuke Kaji, Masaaki Yamamoto, Kentaro Suda, Michiko Takahashi, Yasushi Seo, Yoshihiko Yano, Riko Kitazawa, Sohei Kitazawa, Masafumi Koga, Yasuhiko Okimura, Kazuo Chihara, Yutaka Takahashi
    EUROPEAN JOURNAL OF ENDOCRINOLOGY 167 1 67 - 74 2012年07月 [有り][無し]
     研究論文(学術雑誌) 
    Background: Liver dysfunction in adult hypopituitary patients with GH deficiency (GHD) has been reported and an increased prevalence of nonalcoholic fatty liver disease (NAFLD) has been suggested. Objective: The objective of the present study was to elucidate the pathophysiology of the liver in adult hypopituitary patients with GHD. Patients and methods: We recruited 69 consecutive Japanese adult hypopituitary patients with GHD and examined the prevalence of NAFLD by ultrasonography and nonalcoholic steatohepatitis (NASH) by liver biopsy. Patients had been given routine replacement therapy except for GH. We compared these patients with healthy age-, gender-, and BMI-matched controls. We further analyzed the effect of GH replacement therapy on liver function, inflammation and fibrotic markers, and histological changes. Results: The prevalence of NAFLD in hypopituitary patients with GHD was significantly higher than in controls (77 vs 12%, P<0.001). Of 16 patients assessed by liver biopsy, 14 (21%) patients were diagnosed with NASH. GH replacement therapy significantly reduced serum liver enzyme concentrations in the patients and improved the histological changes in the liver concomitant with reduction in fibrotic marker concentrations in patients with NASH. Conclusions: Adult hypopituitary patients with GHD demonstrated a high NAFLD prevalence. The effect of GH replacement therapy suggests that the NAFLD is predominantly attributable to GHD.
  • Ken-ichiro Tanaka, Yoshifumi Inoue, Geoffrey N. Hendy, Lucie Canaff, Takenobu Katagiri, Riko Kitazawa, Toshihisa Komori, Toshitsugu Sugimoto, Susumu Seino, Hiroshi Kaji
    BONE 51 1 158 - 167 2012年07月 [有り][無し]
     研究論文(学術雑誌) 
    Bone morphogenetic proteins (BMPs) are critical for bone regeneration and induce ectopic bone formation in vivo. The constitutively activating mutation (R206H) of the BMP type 1 receptor, activin A type 1 receptor/activin-like kinase 2 (ACVR1/ALK2), underlies the molecular pathogenesis of fibrodysplasia ossificans progressiva (FOP) in which heterotopic ossification occurs in muscle tissue. In the present study, we performed a comparative DNA microarray analysis between stable empty vector- and ALK2(R206H)-transfected mouse myoblastic C2C12 cells. Forty genes were identified whose expression was increased >3.5 times in the experimental group versus the control. The bone formation-related factor, Tmem119, was included in this group. Osteoblast differentiation markers and mineralization were enhanced in C2C12 cells stably expressing Tmem119. Differentiation of myoblastic cells into myotubes was suppressed but differentiation into chondrocytes was little affected. Transcriptional activity of the BMP-2 signaling molecules, Smad1/5, was increased even in the absence of exogenous BMP-2. Endogenous BMP-2 levels positively correlated with Tmem119 levels. A BMP-2/4 neutralizing antibody and dorsomorphin, an ALK2 inhibitor, antagonized Tmem119-enhanced alkaline phosphatase (ALP) levels. Tmem119 siRNA antagonized the BMP-2-induced ALP and osteocalcin, but not Runx2 and Osterix, mRNAs, in C2C12 cells. In conclusion, Tmem119 levels were increased by the FOP-associated constitutively activating ALK2 mutation in myoblasts. The data show that Tmem119 promotes the differentiation of myoblasts into osteoblasts and the interaction with the BMP signaling pathway likely occurs downstream of Runx2 and Osterix in myoblasts. Tmem119 may play a critical role in the commitment of myoprogenitor cells to the osteoblast lineage. (C) 2012 Elsevier Inc. All rights reserved.
  • Hitoshi Nishizawa, Michiko Takahashi, Hidenori Fukuoka, Genzo Iguchi, Riko Kitazawa, Yutaka Takahashi
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 423 2 295 - 300 2012年06月 [有り][無し]
     研究論文(学術雑誌) 
    The progression to nonalcoholic steatohepatitis (NASH) from simple steatosis is associated with the mitochondrial dysfunction, enhanced oxidative stress, and inflammation. Recently, it has been reported that the prevalence of NAFLD (nonalcoholic fatty liver disease)/NASH is increased in patients with adult growth hormone deficiency (AGHD), suggesting that the deficiencies in GH and insulin-like growth factor (IGF-I) are involved in the development of NAFLD/NASH; however, the precise underlying mechanism remains to be elucidated. To clarify the mechanisms and the specific contribution of GH and IGF-I in these conditions, we examined the liver of a CH-deficient rat model, spontaneous dwarf rat (SDR) and the effect of GH and IGF-I administration. SDR showed steatosis and fibrosis in the liver in line with the phenotype observed in AGHD. Serum AST and ALT levels and triglyceride content in the liver were significantly increased in the SDR compared with the control. Intriguingly, the mitochondrial morphology in the SDR hepatocyte was impaired and the area was significantly decreased. Furthermore, oxidative stress in the SDR liver was enhanced. These changes were improved not only by GH but also by IGF-I administration, suggesting that GH-independent IGF-I action plays an essential role in the liver. In conclusion, we demonstrated that GH-deficient rat exhibits NASH and IGF-I plays an essential role to prevent the development of NASH. The improved mitochondrial function and reduced oxidative stress may contribute the effect of IGF-I in the liver. (C) 2012 Elsevier Inc. All rights reserved.
  • Sohei Kitazawa, Takeshi Kondo, Kiyoshi Mori, Naoki Yokoyama, Masafumi Matsuo, Riko Kitazawa
    HUMAN MUTATION 33 4 651 - 654 2012年04月 [有り][無し]
     研究論文(学術雑誌) 
    We evaluated an autopsy case with severe neonatal respiratory distress, hypoplasia of thymus, thyroid gland and cerebellum, and agenesis of the corpus callosum displaying striking phenotypic similarity to the CrebA knockout mouse. On the assumption that comparable genetic alterations must be present, we checked the whole genomic DNA sequence of cyclic adenosine monophosphate (cAMP) response element binding protein 1 (CREB1), the human counterpart of mouse CrebA, and found a missense c.347A>G mutation corresponding to p.D116G within the kinase-inducible domain (KID) of CREB1. When transcribed in vitro, while Ser-133 phosphorylation of KID was maintained upon forskolin treatment, mutated CREB1 protein failed to associate with the KIX domain of co-activator CREBBP/EP300, and thereby, interrupted cAMP-dependent protein kinase A signal transduction as the dominant-negative form. This is the first report of a sporadic CREB1-related multiple malformation syndrome that, in light of accumulated knowledge of phenotypic features in gene-targeted animals, clearly emphasizes the importance of cross-species translational research. Hum Mutat 33:651654, 2012. (c) 2012 Wiley Periodicals, Inc.
  • Yuying Wang, Wenguang Liu, Ritsuko Masuyama, Ryo Fukuyama, Masako Ito, Quan Zhang, Hisato Komori, Tomohiko Murakami, Takeshi Moriishi, Toshihiro Miyazaki, Riko Kitazawa, Carolina A. Yoshida, Yosuke Kawai, Shinichi Izumi, Toshihisa Komori
    BONE 50 1 409 - 419 2012年01月 [有り][無し]
     研究論文(学術雑誌) 
    Disuse osteoporosis, which occurs commonly in prolonged bed rest and immobilization, is becoming a major problem in modern societies; however, the molecular mechanisms underlying unloading-driven bone loss have not been fully elucidated. The osteocyte network is considered to be an ideal mechanosensor and mechanotransduction system. We searched for the molecules responsible for disuse osteoporosis using BCL2 transgenic mice, in which the osteocyte network was disrupted. Pyruvate dehydrogenase kinase 4 (Pdk4), which inactivates pyruvate dehydrogenase complex (PDC), was upregulated in femurs and tibiae of wild-type mice but not of BCL2 transgenic mice after tail suspension. Bone in Pdk4(-/-) mice developed normally and was maintained. At unloading, however, bone mass was reduced due to enhanced osteoclastogenesis and Rankl expression in wild-type mice but not in Pdk4(-/-) mice. Osteoclast differentiation of Pdk4(-/-) bone marrow-derived monocyte/macrophage lineage cells (BMMs) in the presence of M-CSF and RANKL was suppressed, and osteoclastogenesis was impaired in the coculture of wild-type BMMs and Pdk4(-/-) osteoblasts, in which Rankl expression and promoter activity were reduced. Further, introduction of Pdk4 into Pdk4(-/-) BMMs and osteoblasts enhanced osteoclastogenesis and Rankl expression and activated Rankl promoter. These findings indicate that Pdk4 plays an important role in bone loss at unloading by promoting osteoclastogenesis. (C) 2011 Elsevier Inc. All rights reserved.
  • Yusa Nagai, Riko Kitazawa, Miku Nakagawa, Munenori Komoda, Takeshi Kondo, Ryuma Haraguchi, Sohei Kitazawa
    Case reports in pathology 2012 613180 - 613180 2012年 [有り][無し]
     
    Introduction. Multiple system atrophy (MSA) is a rare and severe adult-onset, sporadic, and progressive neurodegenerative disorder. Here, we describe an autopsy case of MSA in a long-term professional painter. Although typical glial cytoplasmic inclusion (GCI) was not observed in a routine histological examination, strong α-synuclein immunostaining in the nucleus confirmed the diagnosis of MSA. Case Presentation. A 48-year-old Japanese man with a long occupational history of professional painter was sent to the emergency room, where he died of multiple organ failure. The patient had suffered tremors and inarticulateness at age 28, developed diabetes at 42 and was diagnosed with spinocerebellar degeneration at 46. A histopathological examination showed severe neuronal loss, gliosis, and tissue rarefaction in the paleostriatum, striate body of the substantia nigra, the pons, and the olivary nucleus of the upper medulla oblongata, intermediolateral of the spinal gray matter (sacral region). α-synuclein-positive GCI in oligodendroglia was occurred in the cerebral cortex, the midbrain, the medulla oblongata, and the spinal cord. These findings confirmed the presence of multiple-system atrophy (OPCA+SDS). Conclusion. Although the pathogenesis of MSA is still unclear, prolonged, and extensive exposure to organic solvents, together with a hyperglycemic morbidity attributed to diabetes, may have contributed to the onset and clinical course of the present case.
  • Takahashi M, Okimura Y, Iguchi G, Nishizawa H, Yamamoto M, Suda K, Kitazawa R, Fujimoto W, Takahashi K, Zolotaryov FN, Hong KS, Kiyonari H, Abe T, Kaji H, Kitazawa S, Kasuga M, Chihara K, Takahashi Y
    Scientific reports 1 123  2011年10月 [有り][無し]
     研究論文(学術雑誌) 
    Although various function of chemerin have been suggested, its physiological role remains to be elucidated. Here we show that chemerin-deficient mice are glucose intolerant irrespective of exhibiting reduced macrophage accumulation in adipose tissue. The glucose intolerance was mainly due to increased hepatic glucose production and impaired insulin secretion. Chemerin and its receptor ChemR23 were expressed in beta-cell. Studies using isolated islets and perfused pancreas revealed impaired glucose-dependent insulin secretion (GSIS) in chemerin-deficient mice. Conversely, chemerin transgenic mice revealed enhanced GSIS and improved glucose tolerance. Expression of MafA, a pivotal transcriptional factor for beta-cell function, was downregulated in chemerin-deficient islets and a chemerin-ablated beta-cell line and rescue of MafA expression restored GSIS, indicating that chemerin regulates beta-cell function via maintaining MafA expression. These results indicate that chemerin regulates beta-cell function and plays an important role in glucose homeostasis in a tissue-dependent manner.
  • J. C. Young, A. Jaiprakash, S. Mithraprabhu, C. Itman, R. Kitazawa, L. H. J. Looijenga, K. L. Loveland
    INTERNATIONAL JOURNAL OF ANDROLOGY 34 4 E204 - E217 2011年08月 [有り][無し]
     研究論文(学術雑誌) 
    Germ cell testicular cancer is understood to arise during embryogenesis, based on the persistence of embryonic germ cell markers in carcinoma in situ and seminoma. In this study, we examine the potential of the seminoma-derived TCam-2 cell line to be used as representative in functional analyses of seminoma. We demonstrate expression of several early germ cell markers, including BLIMP1, OCT3/4, AP2 gamma, NANOG and KIT. Many TGF-beta superfamily receptors and downstream transcription factors are also present in these cells including the normally foetal ACTRIIA receptor, indicating potential responsiveness to TGF-beta superfamily ligands. Treatment with BMP4 or RA induces a significant increase in ACTRIA, ACTRIIA and ACTRIIB transcripts, whereas activin A decreases ACTRIB. BMP4 and RA each support TCam-2 survival and/or proliferation. In addition, despite increased KIT mRNA levels induced by BMP4, RA and activin A, activin A does not improve survival or proliferation. The capacity for BMP4 and retinoic acid to enhance foetal germ cell survival and proliferation/self-renewal has been demonstrated in mice, but not previously tested in humans. This study is the first to demonstrate a functional response in seminoma cells, using a well-characterized cell line, consistent with their foetal germ cell-like identity.
  • Mukai S, Kitazawa R, Ishii J, Kondo T, Hakozaki A, Horiuchi K, Haraguch R, Mori K, Kitazawa S
    Molecular and cellular biochemistry 350 1-2 29 - 38 2011年04月 [有り][無し]
     研究論文(学術雑誌) 
    Receptor activator of NF-kappa B (RANK) is a member of the tumor necrosis factor receptor (TNFR) family expressed in osteoclast precursors, and RANK-RANK ligand (RANKL) signaling is a key system for differentiation, activation and survival of osteoclasts. Here, we report the identification of a novel alternative splicing variant of mouse RANK gene (vRANK) that contains a new intervening exon between exon 1 and exon 2 of mouse full-length RANK (fRANK) mRNA. Since this novel exon contains the stop codon, vRANK encodes truncated amino acids that have a portion of the signal peptide of fRANK and an additional 19 amino acids that show no homology to previously reported domains. By transient transfection studies with vRANK-GFP and -Flag expressing constructs, vRANK was found localized mostly in the cytoplasm and partly in the cell membrane, but was not secreted into the culture supernatant. Under the stimulation of various factors, the expression of vRANK mRNA was almost parallel to that of fRANK in RAW264.7 cells not treated with M-CSF. Overexpression of vRANK, on the other hand, decreased TRACP (a marker of osteoclasts) mRNA expression as well as the number of TRACP-positive multinucleated giant cells. While the mRNA expression levels of NFATc1 (a master transcriptional factor of the osteoclast differentiation program) were not affected, apoptotic cells increased significantly in vRAN K-transfected cells treated with sRANKL. Taken together, these results suggest that vRANK is a novel osteoclast suppressor that reduces the number of RANKL-induced mature osteoclasts mainly by negating the anti-apoptotic effect of RANKL.
  • Khin SS, Kitazawa R, Kondo T, Idei Y, Fujimoto M, Haraguchi R, Mori K, Kitazawa S
    Cancers 3 1 982 - 93 2011年03月 [有り][無し]
     
    Epigenetic alterations in cancer, especially DNA methylation and histone modification, exert a significant effect on the deregulated expression of cancer-related genes and lay an epigenetic pathway to carcinogenesis and tumor progression. Global hypomethylation and local hypermethylation of CpG islands in the promoter region, which result in silencing tumor suppressor genes, constitute general and major epigenetic modification, the hallmark of the neoplastic epigenome. Additionally, methylation-induced gene silencing commonly affects a number of genes and increases with cancer progression. Indeed, cancers with a high degree of methylation (CpG island methylator phenotype/CIMP) do exist and represent a distinct subset of certain cancers including colorectal, bladder and kidney. On the other hand, signals from the microenvironment, especially those from transforming growth factor-β (TGF-β), induce targeted de novo epigenetic alterations of cancer-related genes. While TGF-β signaling has been implicated in two opposite roles in cancer, namely tumor suppression and tumor promotion, its deregulation is also partly induced by epigenetic alteration itself. Although the epigenetic pathway to carcinogenesis and cancer progression has such reciprocal complexity, the important issue is to identify genes or signaling pathways that are commonly silenced in various cancers in order to find early diagnostic and therapeutic targets. In this review, we focus on the epigenetic alteration by DNA methylation and its role in molecular modulations of the TGF-β signaling pathway that cause or underlie altered cancer-related gene expression in both phases of early carcinogenesis and late cancer progression.
  • Itoko Hisa, Yoshifumi Inoue, Geoffrey N. Hendy, Lucie Canaff, Riko Kitazawa, Sohei Kitazawa, Toshihisa Komori, Toshitsugu Sugimoto, Susumu Seino, Hiroshi Kaji
    JOURNAL OF BIOLOGICAL CHEMISTRY 286 11 9787 - 9796 2011年03月 [有り][無し]
     研究論文(学術雑誌) 
    The mechanisms whereby the parathyroid hormone (PTH) exerts its anabolic action on bone are incompletely understood. We previously showed that inhibition of ERK1/2 enhanced Smad3-induced bone anabolic action in osteoblasts. These findings suggested the hypothesis that changes in gene expression associated with the altered Smad3-induced signaling brought about by an ERK1/2 inhibitor would identify novel bone anabolic factors in osteoblasts. We therefore performed a comparative DNA microarray analysis between empty vector-transfected mouse osteoblastic MC3T3-E1 cells and PD98059-treated stable Smad3-overexpressing MC3T3-E1 cells. Among the novel factors, Tmem119 was selected on the basis of its rapid induction by PTH independent of later increases in endogenous TGF-beta. The levels of Tmem119 increased with time in cultures of MC3T3-E1 cells and mouse mesenchymal ST-2 cells committed to the osteoblast lineage by BMP-2. PTH stimulated Tmem119 levels within 1 h as determined by Western blot analysis and immunocytochemistry in MC3T3-E1 cells. MC3T3-E1 cells stably overexpressing Tmem119 exhibited elevated levels of Runx2, osteocalcin, alkaline phosphatase, and beta-catenin, whereas Tmem119 augmented BMP-2-induced Runx2 levels in mesenchymal cells. Tmem119 interacted with Runx2, Smad1, and Smad5 in C2C12 cells. In conclusion, we identified a Smad3-related factor, Tmem119, that is induced by PTH and promotes differentiation in mouse osteoblastic cells. Tmem119 is an important molecule in the pathway downstream of PTH and Smad3 signaling in osteoblasts.
  • Takayoshi Masuko, Kousaku Souma, Hirofumi Kudo, Yukari Takasaki, Emi Fukui, Reiko Kitazawa, Rikihiro Nishida, Toshimitsu Niida, Teiji Suzuki, Akio Nibe
    ANIMAL SCIENCE JOURNAL 82 4 580 - 586 2011年 [有り][無し]
     研究論文(学術雑誌) 
    Feeding sites for wild Yeso sika deer around Lake Akan, Japan, were established. Effects on the number of deer using the feeding sites, the prevention of bark stripping damage, the amount of feeding, and eating time in a 5-year period (1999-2003) were evaluated. The number of deer using feeding sites increased with years during the feeding period. The damaged tree ratio after the initiation of feeding markedly decreased compared with 16.5% before the initiation of feeding. After the start of feeding, there were no trees with damage the entire circumference. According to tree species, the number of damaged trees of Ulmus laciniata Mayr as a percentage of all investigated trees was high (5.2%). The total amount of beet pulp feeding increased with the feeding year, showing 4.5-fold increase. At feeding sites in deer culling, eating behavior was observed during the night. The preventive effects on bark stripping damage continued during the 5-year feeding period. However, with the course of feeding years, the number of deer using feeding sites and the level of feeding increased.
  • Shunsuke Goto, Hideki Fujii, Keiji Kono, Kentaro Nakai, Yasuhiro Hamada, Hideyuki Yamato, Masami Shinohara, Riko Kitazawa, Sohei Kitazawa, Shinichi Nishi, Masafumi Fukagawa
    AMERICAN JOURNAL OF NEPHROLOGY 34 3 281 - 290 2011年 [有り][無し]
     研究論文(学術雑誌) 
    Background: Diabetic bone disease is a major complication in diabetes mellitus and is characterized by low-turnover bone formation. Recent studies have demonstrated that oxidative stress could be associated with diabetic bone disease and that beta-adrenergic antagonists could increase bone formation. Our study investigated the effect of carvedilol (beta-blocker), possessing an antioxidant effect, on diabetic bone disease. Methods: We used the non-obese, type 2 diabetes model Spontaneously Diabetic Torii (SDT) rats in this study. Sprague-Dawley rats were used as controls (control, n = 6). SDT rats were divided into four groups: diabetic (DM, n = 8), DM+insulin (DM+I, n = 7), DM+carvedilol (DM+C, n = 8), and DM+N-acetylcysteine (DM+N, n = 10) at 20 weeks. The rats were sacrificed at 30 weeks, after which blood and urine samples, bone mineral density, histomorphometry, and oxidative stress were evaluated. Results: The number of 8-hydroxydeoxyguanosine-positive cells in bone tissue was significantly lower in the DM+C and DM+N groups than in the DM group. Mineral apposition rate and bone formation rate per bone surface in the DM+C and DM+N groups were significantly higher than those in the DM group, and these parameters were better in the DM+C group than in the DM+N group. Conclusion: Our data suggest that carvedilol has stronger effects on diabetic low-turnover bone disease beyond that which can be attributed to its antioxidative stress mechanism. Copyright (C) 2011 S. Karger AG, Basel
  • Hamada Y, Kitazawa S, Kitazawa R, Kono K, Goto S, Komaba H, Fujii H, Yamamoto Y, Yamamoto H, Usami M, Fukagawa M
    Endocrine 38 3 369 - 376 2010年12月 [有り][無し]
  • Oka M, Edamatsu H, Kunisada M, Hu L, Takenaka N, Dien S, Sakaguchi M, Kitazawa R, Norose K, Kataoka T, Nishigori C
    Carcinogenesis 31 10 1897 - 1902 2010年10月 [有り][無し]
     研究論文(学術雑誌) 
    Phospholipase C (PLC) epsilon is a phosphoinositide-specific PLC regulated by small guanosine triphosphatases including Ras and Rap. Our previous studies revealed that PLC epsilon gene-knockout (PLC epsilon(-/-)) mice exhibit marked resistance to tumor formation in two-stage skin chemical carcinogenesis using 7,12-dimethylbenz(a)anthracene as an initiator and 12-O-tetradecanoylphorbol-13-acetate as a promoter. In this model, PLC epsilon functions in tumor promotion through augmentation of 12-O-tetradecanoylphorbol-13-acetate-induced inflammation. In this study, we have further assessed the role of PLC epsilon in tumorigenesis using a mouse model of ultraviolet (UV) B-induced skin tumor development. We irradiated PLC epsilon(+/+), PLC epsilon(+/-) or PLC epsilon(-/-) mice with doses of UVB increasing from 1 to 10 kJ/m(2) three times a week for a total of 25 weeks and observed tumor formation for up to 50 weeks. In sharp contrast to the results from the two-stage chemical carcinogenesis study, PLC epsilon(-/-) mice developed a large number of neoplasms including malignant tumors, whereas PLC epsilon(+/+) and PLC epsilon(+/-) mice developed a relatively small number of benign tumors. However, UVB-induced skin inflammation was greatly suppressed in PLC epsilon(-/-) mice, as observed with 12-O-tetradecanoylphorbol-13-acetate-induced inflammation, implying that PLC epsilon's role in the suppression of UVB-induced tumorigenesis is not mediated by inflammation. Studies of the tumor initiation stage revealed that UVB-induced cell death in the skin was markedly suppressed in PLC epsilon(-/-)mice. Our findings identify a novel function for PLC epsilon as a critical molecule regulating UVB-induced cell death and suggest that resistance to UVB-induced cell death conferred by the absence of PLC epsilon is closely related to the higher incidence of skin tumor formation.
  • Yasutaka Mizoro, Yoshiaki Yamaguchi, Rena Kitazawa, Hiroyuki Yamada, Masahiro Matsuo, Jean-Michel Fustin, Masao Doi, Hitoshi Okamura
    PLOS ONE 5 6 e10951  2010年06月 [有り][無し]
     研究論文(学術雑誌) 
    The glutamatergic neurotransmission in the suprachiasmatic nucleus (SCN) plays a central role in the entrainment of the circadian rhythms to environmental light-dark cycles. Although the glutamatergic effect operating via NMDAR (N-methyl D-aspartate receptor) is well elucidated, much less is known about a role of AMPAR (alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor) in circadian entrainment. Here we show that, in the mouse SCN, GluR2 and GluR4 AMPAR subtypes are abundantly expressed in the retinorecipient area. In vivo microinjection of AMPA in the SCN during the early subjective night phase-delays the behavioral rhythm. In the organotypic SCN slice culture, AMPA application induces phase-dependent phase-shifts of core-clock gene transcription rhythms. These data demonstrate that activation of AMPAR is capable of phase-shifting the circadian clock both in vivo and in vitro, and are consistent with the hypothesis that activation of AMPA receptors is a critical step in the transmission of photic information to the SCN.
  • Hirotaka Komaba, Shunsuke Goto, Hideki Fujii, Yasuhiro Hamada, Akira Kobayashi, Koji Shibuya, Yoshihiro Tominaga, Naoki Otsuki, Ken-ichi Nibu, Kimie Nakagawa, Naoko Tsugawa, Toshio Okano, Riko Kitazawa, Masafumi Fukagawa
    KIDNEY INTERNATIONAL 77 3 232 - 238 2010年02月 [有り][無し]
     研究論文(学術雑誌) 
    Fibroblast growth factor 23 (FGF23) exerts its effect by binding to its cognate FGF receptor 1 (FGFR1) in the presence of its co-receptor Klotho. Parathyroid glands express both FGFR1 and Klotho, and FGF23 decreases parathyroid hormone gene expression and hormone secretion directly. In uremic patients with secondary hyperparathyroidism (SHPT), however, parathyroid hormone secretion remains elevated despite extremely high FGF23 levels. To determine the mechanism of this resistance, we measured the expression of Klotho, FGFR1, and the proliferative marker Ki67 in 7 normal and 80 hyperplastic parathyroid glands from uremic patients by immunohistochemistry. All uremic patients had severe SHPT along with markedly high FGF23 levels. Quantitative real-time reverse transcription PCR showed that the mRNA levels for Klotho and FGFR1 correlated significantly with their semi-quantitative immunohistochemical intensity. Compared with normal tissue, the immunohistochemical expression of Klotho and FGFR1 decreased, but Ki67 expression increased significantly in hyperplastic parathyroid glands, particularly in glands with nodular hyperplasia. These results suggest that the depressed expression of the Klotho-FGFR1 complex in hyperplastic glands underlies the pathogenesis of SHPT and its resistance to extremely high FGF23 levels in uremic patients. Kidney International (2010) 77, 232-238; doi:10.1038/ki.2009.414; published online 4 November 2009
  • Hideki Fujii, Keiji Kono, Kentaro Nakai, Shunsuke Goto, Hirotaka Komaba, Yasuhiro Hamada, Masami Shinohara, Riko Kitazawa, Sohei Kitazawa, Masafumi Fukagawa
    AMERICAN JOURNAL OF NEPHROLOGY 31 4 342 - 352 2010年 [有り][無し]
     研究論文(学術雑誌) 
    Background: The role of nitric oxide (NO) is controversial in diabetes nephropathy progression and the mechanisms remain unknown, especially in non-obese type 2 diabetes. To examine mechanisms of nephropathy progression in nonobese type 2 diabetes, we used spontaneously diabetic Torii (SDT) rats, a newly established model of non-obese type 2 diabetes. Methods: Fourteen male Sprague-Dawley rats were used as a control (20 weeks, n = 6; 30 weeks, n = 8), and 20-week-old male SDT rats were divided into 2 groups: diabetic (DM, n = 8) and DM + insulin (n = 8) groups. Twenty-and 36-week-old rats were sacrificed, and blood, urine, and histomorphometric analyses, mRNA expression analysis of endothelial NO synthase (eNOS) and NADPH oxidase, and blood pressure measurement were performed. Results: At 36 weeks, NO metabolites, and 8-hydroxydeoxyguanosine (8-OHdG) were significantly higher in the diabetic group than in the other 2 groups. Further renal studies showed in creased glomerular volume and mesangial area, and intensified eNOS, 8-OHdG, and nitrotyrosine immunostaining in the diabetic group. Oxidative and nitrosative stress were positively associated with increased glomerular volume and mesangial area, which were mostly recovered by insulin therapy. Conclusions: NO and oxidative stress increased in SDT rats, suggesting that these play key roles in nephropathy progression in non-obese type 2 diabetes. Copyright (C) 2010 S. Karger AG, Basel
  • Kondo T, Kitazawa R, Kawata E, Mori K, Kitazawa S
    Cases journal 2 9127  2009年12月 [有り][無し]
  • Kitazawa S, Kitazawa R, Kondo T, Mori K, Matsui T, Watanabe H, Watanabe M
    Cases journal 2 9095  2009年11月 [有り][無し]
  • Ippei Kanazawa, Mika Yamauchi, Shozo Yano, Yasuo Imanishi, Riko Kitazawa, Yoshiki Nariai, Asuka Araki, Keisuke Kobayashi, Masaaki Inaba, Riruke Maruyama, Toru Yamaguchi, Toshitsugu Sugimoto
    BONE 45 3 603 - 608 2009年09月 [有り][無し]
     研究論文(学術雑誌) 
    Malignant transformation of fibrous dysplasia is very rare and has not been previously described in patients with McCune-Albright syndrome in the absence of radiation treatment during gestation. Here, we report a 38-year-old pregnant woman with McCune-Albright syndrome and acromegaly accompanied by osteosarcoma. The patient was in the 6th week of pregnancy, when she visited our hospital. She had Multiple fibrous dysplasia, skin pigmentation, and acromegaly. The markedly high bone turnover rate during pregnancy tended to decrease after a normal delivery. Fibrous dysplasia of the lower jaw rapidly increased in the 37th week of pregnancy, and the tumor was surgically resected after delivery. Pathological examination of the resected tumor revealed fibrous dysplasia admixed with osteosarcoma containing chondroblastic and osteoblastic tissue. We firstly reported a case of osteosarcoma in a patient with McCune-Albright syndrome, which rapidly progressed during pregnancy. (C) 2009 Elsevier Inc. All rights reserved.
  • Kitazawa R, Yamamichi F, Hidaka T, Morishita S, Kondo T, Mori K, Kitazawa S
    Cases journal 2 8326  2009年09月 [有り][無し]
  • Mingzhen Li, Hironori Edamatsu, Riko Kitazawa, Sohei Kitazawa, Tohru Kataoka
    CARCINOGENESIS 30 8 1424 - 1432 2009年08月 [有り][無し]
     研究論文(学術雑誌) 
    Apc(Min/+) mice, carrying an inactivated allele of the adenomatous polyposis coli gene, are widely used as an animal model for human colorectal tumorigenesis, where tumor environment, such as inflammation, is known to play a critical role in tumor progression. We previously demonstrated that phospholipase C (PLC)epsilon, an effector of Ras and Rap small GTPases, plays a crucial role in two-stage skin chemical carcinogenesis using 12-O-tetradecanoyl-phorbor-13-acetate (TPA) as a promoter through augmentation of TPA-induced inflammation. Here, we show that Apc(Min/+) mice lacking PLC epsilon (PLC epsilon(-/-)) exhibit marked resistance to spontaneous intestinal tumorigenesis compared with those with the PLC epsilon(+/+) background. Time course of the development of tumors, which are histopathologically classified into low- and high-grade adenomas with increasing dysplasia and size, and adenocarcinomas indicates that not only the low-grade adenoma formation but also the progression to high-grade adenoma are suppressed in PLC epsilon(-/-);Apc(Min/+) mice. Low-grade adenomas of PLC epsilon(-/-);Apc(Min/+) mice exhibit accelerated apoptosis and reduced cellular proliferation. They also show marked attenuation of tumor angiogenesis and reduction in expression of vascular endothelial growth factor. In contrast, high-grade adenomas of PLC epsilon(-/-);Apc(Min/+) mice exhibit marked attenuation of tumor-associated inflammation without significant differences in apoptosis and proliferation. These results suggest that PLC epsilon plays crucial roles in intestinal tumorigenesis through two distinct mechanisms, augmentation of angiogenesis and inflammation, depending on the tumor stage.
  • Kitazawa S, Mori K, Kondo T, Kitazawa R
    Cases journal 2 8280  2009年08月 [有り][無し]
  • Hideki Fujii, Fuyuhiko Nishijima, Sumie Goto, Mikio Sugano, Hideyuki Yamato, Riko Kitazawa, Sohei Kitazawa, Masafumi Fukagawa
    NEPHROLOGY DIALYSIS TRANSPLANTATION 24 7 2089 - 2095 2009年07月 [有り][無し]
     研究論文(学術雑誌) 
    Methods. Male Lewis rats were administered adriamycin at 8 weeks of age, and the right kidney was removed at 12 weeks of age. From 14 weeks of age, the rats were treated daily with AST-120 (n = 8) or were untreated (control group, n = 8). At 34 weeks of age, the rats were killed and urinary and blood biochemical tests as well as cardiac histological analyses were performed. Results. At 14 weeks of age, there were no significant differences in blood pressure, renal function (creatinine clearance: 1.54 +/- 0.28 mL/min versus 1.60 +/- 0.22 mL/min), oxidative stress markers or other biochemical data between the control and AST-120 groups. At 34 weeks, despite similar blood pressure and renal function (creatinine clearance: 0.78 +/- 0.46 mL/min versus 0.75 +/- 0.54 mL/min), serum concentrations of IS and urinary excretion of 8-hydroxydeoxyguanosine (8-OHdG), acrolein and IS were significantly lower in the AST-120 group than in the control group. Heart volume, left ventricular volume and cardiac fibrosis were significantly smaller in the experimental AST-120 group than in the control group. Immunohistological analysis revealed that the numbers of 8-OHdG- and acrolein-positive cardiomyocytes and the degrees of myocardial and perivascular fibrosis were ameliorated by AST-120 administration. The myocardial fibrosis score was significantly associated with the 8-OHdG- (r = 0.848, P < 0.001) and acrolein-positive (r = 0.812, P < 0.001) cell scores. The perivascular fibrosis score was also significantly associated with the 8-OHdG- (r = 0.906, P < 0.0001) and acrolein-positive (r = 0.789, P < 0.001) cell scores. Conclusions. Oxidative stress is suggested to play a key role in the development of cardiac hypertrophy and fibrosis in CKD. AST-120 may suppress oxidative stress and reduce cardiac damage in CKD.
  • Sann Sanda Khin, Riko Kitazawa, Ne Win, Than Than Aye, Kiyoshi Mori, Takeshi Kondo, Sohei Kitazawa
    INTERNATIONAL JOURNAL OF CANCER 125 2 328 - 338 2009年07月 [有り][無し]
     研究論文(学術雑誌) 
    The bone morphogenetic protein (BMP) and activin membrane-bound inhibitor (BAMBI) is a transmembrane TGFRI/BMPRI-related pseudoreceptor, antagonizes transforming growth factor (TGF)-beta/BMP signaling by inhibiting the formation of functional authentic receptor complexes (TGFRI/BMPRI and TGFRII/BMPRII). On the assumption that BAMBI gene expression is epigenetically altered during human bladder cancer progression, we screened the expression of BAMBI protein by immunohistochemistry and the methylation status of the BAMBI promoter. In the normal or reactive urothelium, BAMBI expression was mostly overlapped with that of BMPRI, and a similar colocalization pattern was noted in low-grade papillary cancers. In high-grade and invasive cancers, however, mainly two reciprocal immunohistochemical expression patterns were observed: BAMBI-low/BMPRI-high, and BAMBI-high/BMPRI-low, indicating that BAMBI expression is controlled such that it does not interfere with the responsiveness of high-grade cancer cells to TGF-beta/BMP signaling. Moreover, methylation of the BAMBI gene correlated significantly with negative BAMBI expression in bladder tumors. Although BAMBI overexpression significantly increased the number of apoptotic cells in T24 line, knock-down small interfering RNA showed no remarkable change. Cell motility assay revealed that on treatment with either TGF-beta 1 or BMP2, T24 and HTB9 lines showed a marked increase in the number of migrated cells which,, however, decreased significantly through the forced expression of BAMBI. Since certain subsets of aggressive tumors often promote cell motility, invasion and survival by inducing epithelial-to-mesenchymal transition through TGF-beta/BMP in an autocrine and paracrine manner, hypermethylation of the BAMBI gene promoter that leads to BAMBI gene suppression may be one of the epigenetic events affecting the invasiveness or aggressiveness of bladder cancers. (C) 2009 UICC
  • Yasuhiro Hamada, Hideki Fujii, Riko Kitazawa, Junji Yodoi, Sohei Kitazawa, Masafumi Fukagawa
    BONE 44 5 936 - 941 2009年05月 [有り][無し]
     研究論文(学術雑誌) 
    Diabetes mellitus is associated with increased risk of osteopenia and bone fracture. However, the mechanisms accounting for diabetic bone disorder are unclear. We have previously reported that streptozotocin-induced diabetic mice develop low turnover osteopenia associated with increased oxidative stress in the diabetic condition. To determine the role of oxidative stress in the development of diabetic osteopenia, we presently investigated the effect of overexpression of thioredoxin-1 (TRX), a major intracellular antioxidant, on the development of diabetic osteopenia, using TRX transgenic mice (TRX-Tg). TRX-Tg are C57BL/6 mice that carry the human TRX transgene under the control of beta-actin promoter. Eight-week-old male TRX-Tg mice and wild type (WT) littermates were intraperitoneally injected with either streptozotocin or vehicle. Mice were grouped as 1) non-diabetic WT, 2) non-diabetic TRX-Tg, 3) diabetic WT, and 4) diabetic TRX-Tg. After 12 weeks of streptozotocin treatment, oxidative stress on the whole body and bone was evaluated, and the physical properties of the femora, and histomorphometry parameters of the tibiae were assessed. TRX overexpression did not affect either body weight or hemoglobin A1c levels. There were no significant differences in renal function and in serum levels of calcium, phosphate, and intact parathyroid hormone among the four groups. Oil the other hand, urinary excretion of 8-hydroxydeoxyguanosine (8-OHdG), a marker of oxidative DNA damage, was significantly elevated in diabetic WT and attenuated in diabetic TRX-Tg. Immunohistochemical staining for 8-OHdG revealed marked intensity in the bone tissue of diabetic WT compared with non-diabetic WT while staining was attenuated in diabetic TRX-Tg. TRX overexpression partially restored reduced bone mineral density and prevented the suppression of bone formation observed in diabetic WT. Increased oxidative stress in diabetic condition contributes to the development of diabetic osteopenia. Suppression of increased oxidative stress by TRX induction could be a potential therapeutic approach for diabetic osteopenia. (C) 2008 Elsevier Inc. All rights reserved.
  • Riko Kitazawa, Kiyoshi Mori, Akira Yamaguchi, Takeshi Kondo, Sohei Kitazawa
    JOURNAL OF CELLULAR BIOCHEMISTRY 105 5 1289 - 1297 2008年12月 [有り][無し]
     研究論文(学術雑誌) 
    The expression of receptor activator of nuclear factor-kappa B ligand (RANKL) is regulated by bone-seeking hormones such as PTH and 1 alpha,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)). Runx2, a master gene for osteoblastic differentiation, also modulates osteoclastogenesis by regulating the RANKL gene. To elucidate the mechanism whereby runx2 and 1,25(OH)(2)D(3) regulate RANKL expression, we studied the function of runx2 on the chromatin structure and on the proximal binding sites using osteoblastic cell lines derived from normal (ST2) and runx2-deficient mice (RD-C6). Although the expression of RANKL in the steady-state was higher in RD-C6 than in ST2, 1,25(OH)(2)D(3)-treatment of the cells increased it 20-fold in ST2 but only 1.8-fold in RD-C6. Transient transfection studies with proximal RANKL 2kb promoter, runx2 knock-down in ST2, and forced expression of runx2 in RD-C6 all confirmed that runx2 set the steady-state expression of the RANKL gene at a low level, but exerted a positive effect on enhanced transcriptional activity in response to 1,25(OH)(2)D(3). Also, assessment of the acetylation status of the area spanning 40 kb upstream of the basic promoter in ST2 and RD-C6 by ChIP assay revealed that whereas H3 and H4 historic acetylation was detected even in the steady-state in RD-C6, it was detected only with 1,25(OH)(2)D(3) in ST2. In the steady-state, runx2 may suppress RANKL gene by condensing the chromatin structure; however, it exerts a positive effect on 1,25(OH)(2)D(3)-induced RANKL transcription when the proximal runx2 sites are accessible. Thus, RANKL expression in stromal/osteoblastic cells is keenly regulated by 1,25(OH)(2)D(3) which transactivates the gene at two different levels. J. Cell. Biochem. 105: 1289-1297, 2008. (c) 2008 Wiley-Liss, Inc.
  • Junko Ishii, Riko Kitazawa, Kiyoshi Mori, Kevin P. McHugh, Eiichi Morii, Takeshi Kondo, Sohei Kitazawa
    JOURNAL OF CELLULAR BIOCHEMISTRY 105 3 896 - 904 2008年10月 [有り][無し]
     研究論文(学術雑誌) 
    Receptor activator of NF-kappa B (RANK) is a receptor for RANK ligand (RANKL), and signals transduced by RANK-RANKL interaction are prerequisite for the differentiation and activation of osteoclasts. We cloned and characterized a 6-kb fragment containing the 5'-flanking region of the mouse RANK gene. A fragment of 1-kb from the transcription start sites containing four Sp-1 sites and putative binding sites for MITF, CRE/AP-1, and PU.1 was ligated to the pGL3-basic vector, and the promoter activity was confirmed by transfection studies. By electrophoretic gel motility shift assay, both PU.1 and proximal MITF binding site showed specific DNA-protein binding. Co-transfection studies with MITF- and PU.1-expression vectors revealed that MITF and PU.1 increased RANK promoter activity three- and twofold, respectively, and sixfold synergistically. Taken together, these results show that RANK transcription is positively regulated by both PU.1 and MITF. The effect of lipopolysaccharide (LPS) on RANK gene expression, analyzed by in situ hybridization using mouse bone tissue, showed that LPS decreased RANK transcripts of both precursor and mature osteoclasts. Furthermore, LPS treatment of RAW.264.7 cells decreased their RANK mRNA expression by 70%, mirroring the decrease of PU.1 and MITF mRNA. Short-term treatment with LPS decreased the promoter activity of pGL3-WT by 70%. Although LPS has been reported to promote osteoclastogenesis in chronic and local pyogenic inflammation, we speculate that LPS per se may directly suppress RANK expression in the osteoclastic cell lineage by down-regulating the expression of PU.1 and MITF genes in acute and systemic severe endotoxemia, such as in septic shock. J. Cell. Biochem. 105: 896-904, 2008. (C) 2008 Wiley-Liss, Inc.
  • Masanori Fuse, Norihide Yokoi, Masami Shinohara, Taku Masuyama, Riko Kitazawa, Sohei Kitazawa, Susumu Seino
    PHYSIOLOGICAL GENOMICS 35 1 96 - 105 2008年09月 [有り][無し]
     研究論文(学術雑誌) 
    Fuse M, Yokoi N, Shinohara M, Masuyama T, Kitazawa R, Kitazawa S, Seino S. Identification of a major locus for islet inflammation and fibrosis in the spontaneously diabetic Torii rat. Physiol Genomics 35: 96-105, 2008. First published July 8, 2008; doi: 10.1152/physiolgenomics.90214.2008.-The pathogenesis of inflammation and fibrosis in the pancreatic islets in diabetes is largely unknown. Spontaneously diabetic Torii (SDT) rats exhibit inflammation and fibrosis in and around the islets during the development of the disease. We investigated genetic factors for diabetes, islet inflammation, and fibrosis in the SDT rat. We produced F1 and F2 rats by intercross between SDT and F344 rats, examined the onset of diabetes, glucose tolerance, and histology of the pancreas, and performed genetic analysis of these traits. We then established a congenic strain carrying the SDT allele at the strongest diabetogenic locus on the F344 genetic background and characterized glucose tolerance and histology of the pancreas. F1 rats showed glucose intolerance and inflammatory changes mainly in the islets. Genetic analysis of diabetes identified a major locus on chromosome 3, designated Dmsdt1, at which a dominantly acting SDT allele was involved. Quantitative trait locus (QTL) analysis of glucose tolerance revealed, in addition to Dmsdt1 [logarithm of odds (LOD) 5.3 near D3Mit12], three other loci, designated Dmsdt2 (LOD 4.2 at D8Rat46), Dmsdt3 (LOD 3.8 near D13Arb5), and Dmsdt4 (LOD 5.8 at D14Arb18). Analysis of a congenic strain for Dmsdt1 indicates that the dominantly acting SDT allele induces islet inflammation and fibrosis. Thus we have found a major locus on chromosome 3 for islet inflammation and fibrosis in the SDT rat. Identification of the genes responsible should provide insight into the pathogenesis of diabetes.
  • Mori K, Kitazawa R, Kondo T, Kitazawa S
    Cases journal 1 1 162  2008年09月 [有り][無し]
  • Takeshi Kondo, Riko Kitazawa, Sohei Kitazawa
    DIGESTIVE DISEASES AND SCIENCES 53 8 2287 - 2289 2008年08月 [有り][無し]
     研究論文(学術雑誌) 
    An invasive micropapillary carcinoma ( IMPC) is defined as a carcinoma composed of small clusters of tumor cells lying within clear spaces simulating vascular channels [ 1]. It is a histological variant of invasive breast carcinoma with poor clinical prognosis [2, 3]. This distinct histological pattern has been described in various organs, including the urinary bladder, lung, ovary, and major salivary glands [4 - 8]. Although rarely observed as a pure histological component, IMPC is usually mixed with otherwise conventional carcinoma [3] and is therefore often referred to as carcinoma with a micropapillary component. In cases of adenocarcinoma with a micropapillary component, an abrupt transition is usually seen between the invasive micropapillary component and conventional adenocarcinoma [3]. IMPCs are all invariably associated with high aggressiveness, extensive lymphovascular invasion, extensive lymph node metastases, and poor prognosis [1-3]. We describe the first case of primary IMPC originating in the stomach as a histologic subcomponent of recurrent gastric adenocarcinoma.
  • Naonobu Nishino, Yoshikazu Tamori, Sanshiro Tateya, Takayuki Kawaguchi, Tetsuro Shibakusa, Wataru Mizunoya, Kazuo Inoue, Riko Kitazawa, Sohei Kitazawa, Yasushi Matsuki, Ryuji Hiramatsu, Satoru Masubuchi, Asako Omachi, Kazuhiro Kimura, Masayuki Saito, Taku Amo, Shigeo Ohta, Tomohiro Yamaguchi, Takashi Osumi, Jinglei Cheng, Toyoshi Fujimoto, Harumi Nakao, Kazuki Nakao, Atsu Aiba, Hitoshi Okamura, Tohru Fushiki, Masato Kasuga
    JOURNAL OF CLINICAL INVESTIGATION 118 8 2808 - 2821 2008年08月 [有り][無し]
     研究論文(学術雑誌) 
    White adipocytes are unique in that they contain large unilocular lipid droplets that occupy most of the cytoplasm. To identify genes involved in the maintenance of mature adipocytes, we expressed dominant-negative PPAR gamma in 3T3-L1 cells and performed a microarray screen. The fat-specific protein of 27 kDa (FSP27) was strongly downregulated in this context. FSP27 expression correlated with induction of differentiation in cultured preadipocytes, and the protein localized to lipid droplets in murine white adipocytes in vivo. Ablation of FSP27 in mice resulted in the formation of multilocular lipid droplets in these cells. Furthermore, FSP27-deficient mice were protected from diet-induced obesity and insulin resistance and displayed an increased metabolic rate due to increased mitochondrial biogenesis in white adipose tissue (WAT). Depletion of FSP27 by siRNA in murine cultured white adipocytes resulted in the formation of numerous small lipid droplets, increased lipolysis, and decreased triacylglycerol storage, while expression of FSP27 in COS cells promoted the formation of large lipid droplets. Our results suggest that FSP27 contributes to efficient energy storage in WAT by promoting the formation of unilocular lipid droplets, thereby restricting lipolysis. In addition, we found that the nature of lipid accumulation in WAT appears to be associated with maintenance of energy balance and insulin sensitivity.
  • Hepatic failure and enhanced oxidative stress in mitochondrial diabetes.
    Takahashi Y, Iida K, Takeno R, Kitazawa R, Kitazawa S, Kitamura H, Fujioka Y, Yamada H, Kanda F, Ohta S, Nishimaki K, Fujimoto M, Kondo T, Iguchi G, Takahashi K, Kaji H, Okimura Y, Chihara K
    Endocrine journal 55 3 509 - 514 2008年07月 [有り][無し]
  • Taurine administration after appearance of proteinuria retards progression of diabetic nephropathy in rats.
    Higo S, Miyata S, Jiang QY, Kitazawa R, Kitazawa S, Kasuga M
    The Kobe journal of medical sciences 54 1 E35 - 45 2008年05月 [有り][無し]
  • Sopaporn Niemhom, Sohei Kitazawa, Riko Kitazawa, Sakan Maeda, Juvady Leopairat
    CANCER DETECTION AND PREVENTION 32 2 127 - 134 2008年 [有り][無し]
     研究論文(学術雑誌) 
    Background: Epstein-Barr virus (EBV) is documented as the important etiologic agent of nasopharyngeal carcinoma (NPC) but the mechanism of development and pathogenesis induced by EBV is presently unclear. Hypermethylation of epithelial-cadherin (E-cadherin) promoter has been shown to be induced in NPC cell line by EBV LMP1 via DNA methyltransferase activation. EBV genomes and hypermethylation of E-cadherin promoter were investigated in NPC tissues to evaluate the role of EBV in the hypermethylation and pathogenesis of NPC. Methods: Methylation-specific polymerase chain reaction (MSP) was performed to detect E-cadherin promoter hypermethylation in paraffin embedded tissues from patients with NPC and normal nasopharyngeal tissues. EBV genomes were detected by PCR in the tissue samples. Results: Hypermethylation of E-cadherin promoter and EBV were predominantly detected in undifferentiated and non-keratinizing NPC compared to those in squamous cell NPC. Hypermethylation of E-cadherin was found in 28 of 38 (73.7%) patient samples. EBV was detected in 22 of the 28 (78.6%) NPC samples demonstrating E-cadherin hypermethylation. EBV genomes and hypermethylation were not detected in normal nasopharyngeal tissues. Significant association was found between E-cadherin hypermethylation and EBV genomes (p < 0.001; Fisher's exact test). Hypermethylation of E-cadherin was more frequently detected in advanced stages compared to those in early stages of NPC (p = 0.036; Fisher's exact test). Conclusions: The high incidence of EBV with the consistency of E-cadherin hypermethylation, particularly in undifferentiated and non-keratinizing NPC suggests the role of EBV in the hypermethylation. EBV exists at early stage of NPC that induces the hypermethylation and contributes to progression of the disease to the advanced stage of NPC. (C) 2008 International Society for Preventive Oncology. Published by Elsevier Ltd. All rights reserved.
  • Agus Darwanto, Riko Kitazawa, Kiyoshi Mori, Takeshi Kondo, Sohei Kitazawa
    ACTA HISTOCHEMICA ET CYTOCHEMICA 41 5 135 - 142 2008年 [有り][無し]
     研究論文(学術雑誌) 
    Hypermethylation-dependent silencing of the gene is achieved by recruiting methyl-CpG binding proteins ( MeCPs). Among the MeCPs, MeCP2 is the most abundantly and ubiquitously expressed in various types of cells. We first screened the distribution and expression pattern of MeCP2 in adult and developing rat tissues and found strong MeCP2 expression, albeit rather ubiquitously among normal tissues, in ganglion cells and intestinal epithelium in the small intestine, in Purkinje cells and neurons in the brain, in spermatogonia and in epithelial cells in the epididymal duct of the testis. We then assessed the expression and the methylation pattern of the promoter region of cyclin D1 by immunohistochemistry and sodium bisulfite mapping, and found that cyclin D1 expression in the epididymal duct decreased rapidly during rat development: strong in newborn rats and very weak or almost negative in 7-day-old rats. Mirroring the decrease of cyclin D1 expression, methylated cytosine at both CpG and non-CpG loci in the cyclin D1 promoter was frequently observed in the epididymal duct of 7-day-old rats but not in that of newborn rats. Interestingly, MeCP2 expression also increased concomitant with the increase of methylation. Cyclin D1 expression in the epididymal duct may be efficiently regulated by the epigenetic mechanism of the cooperative increase of MeCP2 expression and promoter methylation.
  • [Regulatory mechanism of bone morphogenetic protein gene expression].
    Kitazawa S, Kitazawa R
    Nihon rinsho. Japanese journal of clinical medicine 63 Suppl 10 409 - 413 2005年10月 [有り][無し]
  • Menin inactivation leads to loss of transforming growth factor beta inhibition of parathyroid cell proliferation and parathyroid hormone secretion
    H Sowa, H Kaji, R Kitazawa, S Kitazawa, T Tsukamoto, S Yano, T Tsukada, L Canaff, GN Hendy, T Sugimoto, K Chihara
    CANCER RESEARCH 64 6 2222 - 2228 2004年03月 [有り][無し]
     研究論文(学術雑誌) 
    Primary hyperparathyroidism is a common endocrine disorder caused by parathyroid gland enlargement and excessive parathyroid hormone (PTH) secretion. However, the precise mechanisms of tumorigenesis of the parathyroids are unknown. Here we have investigated the roles of transforming growth factor (TGF)-beta and menin, the product of the multiple endocrine neoplasia type 1 (Men1) gene, in the proliferation and PTH production of parathyroid cells from either patients with secondary hyperparathyroidism or Men1. TGF-beta was expressed in the parathyroid endocrine cells. Addition of TGF-beta to parathyroid cells from patients with secondary hyperparathyroidism inhibited their proliferation and PTH secretion. These responses to TGF-beta were lost when menin was specifically inactivated by antisense oligonucleotides. Moreover, TGF-beta did not affect the proliferation and PTH production of parathyroid cells from a Men1 patient. These results indicate that menin is required for TGF-beta action in the parathyroid. We conclude that TGF-beta is an important autocrine/paracrine negative regulator of parathyroid cell proliferation and PTH secretion and that loss of TGF-beta signaling due to menin inactivation contributes to parathyroid tumorigenesis.
  • Imaging of fine structure of bone sample with high coherent X-ray beam and high spatial resolution detector.
    Hirano M, Yamasaki K, Kitazawa R, Kitazawa S, Okada H, Katafuchi T, Sakurai T, Kondoh T, Ohbayashi C, Maeda S, Sugimura K, Tamura S
    Radiation medicine 22 1 56 - 59 2004年01月 [有り][無し]
  • [Not Available].
    Chen Q, Kaji H, Iu MF, Nomura R, Sawa H, Yamauchi M, Tsukamoto T, Yamaguchi T, Sugimoto T, Chihara K, Kitazawa R, Kobayashi A
    Clinical calcium 13 4 496 - 499 2003年04月 [有り][無し]
  • Influence of heart surgery on magnesium concentrations in pediatric patients.
    Hoshino K, Ogawa K, Hishitani T, Kitazawa R
    Pediatrics international : official journal of the Japan Pediatric Society 45 1 39 - 44 2003年02月 [有り][無し]
  • S Kitazawa, R Kitazawa
    JOURNAL OF PATHOLOGY 198 2 228 - 236 2002年10月 [有り][無し]
     研究論文(学術雑誌) 
    Breast cancer is frequently associated with osteolytic bone metastasis, where osteoclasts play a major role in bone destruction. Recently, osteoclast differentiation factor (RANKL) has been identified as a prerequisite for the formation and maintenance of osteoclasts from haematopoietic precursors. To elucidate the mechanism of osteoclastogenesis and bone destruction in bone-residing breast cancer, PTHrP-producing (MCF-7) and -non-producing (MCF-7UP) human breast cancer cells were subcutaneously injected into the forehead of nude mice maintained without oestrogen supplement. One, two, and three weeks thereafter, the expression of RANKL and PTHrP mRNA, and osteoclastogenesis were analysed by in situ hybridization and TRAP staining. In MCF-7 cells, at early stages, spindle-shaped stromal cells and osteoblasts on the bone surface expressed RANKL, then numerous osteoclasts were induced on the periosteal bone surface. Three weeks after the transplantation, MCF-7 cancer cells migrated onto the eroded bone surface, where they survived apoptosis. At all stages, RANKL expression was confined to the stromal/osteoblastic cells, whereas PTHrP was confined to the MCF-7 breast cancer cells. On the other hand, PTHrP was negative in MCF-7UP cells at all stages, and neither induction of osteoclasts nor infiltrative growth of cancer cells was observed. Moreover, in vitro treatment with PTHrP resulted in increased RANKL mRNA expression and transcription activity in the MC3T3-E1 mouse osteoblastic cell line. Thus PTHrP induces osteoclastic bone resorption through the transactivation of the RANKL gene on stromal/osteoblastic cells, affording a bone microenvironment conducive to the survival of PTHrP-producing cancer cells. Copyright (C) 2002 John Wiley Sons, Ltd.
  • Atsushi Ishida, Naoya Fujita, Riko Kitazawa, Takashi Tsuruo
    The Journal of biological chemistry 277 29 26217 - 24 2002年07月 [有り][無し]
     
    Vascular endothelial cells in bone are thought to have significant roles on pathological bone resorption such as bone metastasis and hypercalcemia because this resorption is often seen where blood vessels are abundant. However, the detailed mechanisms have not yet been elucidated. Here, we focused on transforming growth factor-beta (TGF-beta) and studied its effects on vascular endothelial cells because TGF-beta is abundantly stored in bone matrix and is released and activated during bone resorption. We found that TGF-beta up-regulated the expression of receptor activator of NF-kappa B ligand (RANKL) mRNA and protein in bone marrow-derived endothelial cells and in primary vascular endothelial cells but not in osteoblasts. Further analysis revealed that TGF-beta promoted phosphorylation of cAMP response element-binding protein and p38. Protein kinase A inhibitor KT5720 and p38 inhibitor SB203580 significantly reduced the TGF-beta-induced RANKL expression. Moreover, we found two CRE-like domains in murine RANKL promoter region that were critical for TGF-beta-dependent RANKL expression. Therefore, protein kinase A and p38 signaling pathways are involved in TGF-beta-induced RANKL expression by stimulating transcription factors that bind to the CRE-like domains. Our findings indicate that TGF-beta stimulates osteoclastogenesis by promoting RANKL expression in endothelial cells under pathological conditions.
  • S Kitazawa, R Kitazawa
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 293 1 126 - 131 2002年04月 [有り][無し]
     研究論文(学術雑誌) 
    Receptor activator of NF-kappaB ligand (RANKL) is a membrane-bound signal transducer requisite for differentiation and maintenance of osteoclasts. RANKL expression on stromal/osteoblastic cells is tightly regulated to maintain physiological serum calcium levels and bone mass. These stromal/osteoblastic cells, however, comprise a rather heterogeneous population ranging from immature mesenchymal cells to mature osteoblasts and also respond differently to bone resorptive stimuli. In the mouse coculture system, we also have demonstrated the passage-dependent difference of cultured mouse stromal cells in supporting osteoclastogenesis due to altered RANKL gene expression. To address the issue of what molecular mechanism gives the diversity of RANKL gene expression to stromal/osteoblastic cells, we characterized the mouse RANKL gene promoter that contains two CpG clustering regions; one around the transcription start site, and the other downstream of the vitamin D response element (VDRE). Using earlier- and later-passage mouse ST2 cells. we analyzed the CpG methylation status by sodium bisulfite mapping and found that CpG loci around the transcription start site (-66/+246) were predominantly methylated in later-passage ST2 cells. Moreover, earlier- and later-passage ST2 cells transfected with a RANKL promoter construct showed the same steady-state level of luciferase activity and of the inducible effect of 1.25(OH)(2)D-3. Furthermore, the introduction of methylation to the promoter construct silenced promoter activity. The results suggest that CpG methylation around the transcription start site of the mouse RANKL gene is an important epigenetic event, and that its heterogeneity might cause the diversity of the stromal/osteoblastic cells in RANKL gene expression. (C) 2002 Elsevier Science (USA). All rights reserved.
  • H Mori, R Kitazawa, S Mizuki, M Nose, S Maeda, S Kitazawa
    HISTOCHEMISTRY AND CELL BIOLOGY 117 3 283 - 292 2002年03月 [有り][無し]
     研究論文(学術雑誌) 
    Rheumatoid arthritis (RA) is a systemic disorder characterized by synovial inflammation and subsequent destruction and deformity of synovial joints. The articular lesions start with synovitis, focal erosion of unmineralized cartilage, and then culminate in the destruction of subarticular bone by pannus tissue. Periarticular osteopenia and systemic osteoporosis follow as late complications of RA. Osteoclasts, specialized cells that resorb bone, play a central role in developing these osteolytic lesions. To elucidate the mechanism of osteoclastogenesis and bone destruction in autoimmune arthritis, we investigated the expression of RANK ligand (RANKL), RANK, and osteoprotegerin (OPG) mRNA in a mouse type II collagen-induced arthritis (CIA) model by in situ hybridization. The results indicated that most of the TRAP-positive mono- and multinucleated cells in the inflamed and proliferating synovium and in the pannus were RANK-positive authentic osteoclasts and their precursors. In the inflamed synovium and pannus of the mouse CIA model, synovial fibroblastic cells around these RANK-positive cells were strongly positive for RANKL. Moreover, RANKL-positive osteoblasts on the endosteal bone surface, at a distance from the affected synovial joints, increased significantly in the mouse CIA model prior to periarticular osteopenia and systemic osteoporosis. These data indicated that the RANKL-RANK system plays an important role for osteoclastogenesis in both local and systemic osteolytic lesions in autoimmune arthritis, and can therefore be a good target for therapeutic intervention.
  • R Kitazawa, S Kitazawa
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 290 2 650 - 655 2002年01月 [有り][無し]
     研究論文(学術雑誌) 
    Receptor activator of NF-B-kappa ligand (RANKL) is a membrane-bound signal transducer necessary for the induction and maintenance of osteoclasts. To clarify the molecular mechanism by which 1,25-dihydroxyvitamin D-3 (1,25-(OH)(2)D-3) augments osteoclasts, we characterized the promoter region of the mouse RANKL gene. Mirroring in vitro osteoclastogenesis demonstrated by a coculture of bone marrow macrophages with ST2 stromal cells, Northern blot, and nuclear run-on analyses showed that 1,25-(OH)(2)D-3 upregulate RANKL gene expression at the transcriptional level. Using a series of deletion mutants of mouse RANKL promoter-luciferase reporter gene constructs, transient transfection studies revealed that the inductive effect of 1,25-(OH)(2)D-3 was abolished when the region up to -723 was deleted. Am electrophoretic motility shift assay demonstrated that the VDR-RXRbeta heterodimer bound to (AGGTCA) under bar GCC (TGGTTCA) under bar (-937/-922), and VDRE/nuclear protein super-shift complexes that bound to anti-VDR and -RXRbeta antibodies were detected in the nuclear extract of 1,25-(OH)(2)D-3-treated ST2 cells. Furthermore, induction of mutation to the putative VDRE also diminished the inductive effect of 1,25-(OH)(2)D-3. We therefore concluded that mouse RANKL gene is one of the target genes of 1,25-(OH)(2)D-3 containing a functional VDRE in the promoter region. (C) 2002 Elsevier Science.
  • R Kitazawa, S Kitazawa, S Maeda, A Kobayashi
    HISTOLOGY AND HISTOPATHOLOGY 17 1 179 - 184 2002年 [有り][無し]
     研究論文(学術雑誌) 
    Parathyroid hormone-related protein (PTHrP), a factor responsible for malignancy associated hypercalcemia, plays a physiological roles such as bone development and placental calcium transport. The expression of PTHrP in adult human parathyroid tissues under normal and pathological conditions was analyzed. By immunohistochemistry, PTHrP expression was detected in 86% of normal parathyroid (12/14 cases), 74% of adenomas (14/19) and 89% of hyperplasia secondary to chronic renal failure (16/18). PTHrP protein was observed mainly in the cytoplasm of oxyphil cells, consistent with the localization of its mRNA demonstrated by in situ hybridization. The rate of PTHrP-positive cells was higher in areas consisting of oxyphil cells than in those of non-oxyphil cells, regardless of whether the parathyroid was normal or pathological. In the normal parathyroid, an age-related increase in PTHrP expression was observed with a relative increase in oxyphil cells, reflecting aging and deterioration of parathyroid tissue. In adenoma, cases with a predominance of oxyphil cells expressed PTHrP, whereas clear cell adenoma did not. In secondary hyperplasia, the rate of PTHrP-expressing cells was higher than in normal parathyroid or adenoma, with varying levels of expression among nodules. We speculate that PTHrP could act through the paracrine/ autocrine mechanism to regulate proliferation and differentiation of normal and neoplastic parathyroid cells.
  • K Hoshino, K Ogawa, T Hishitani, R Kitazawa
    PEDIATRIC CARDIOLOGY 23 1 41 - 48 2002年01月 [有り][無し]
     研究論文(学術雑誌) 
    We studied the role of magnesium (Mg) in congenital long QT syndrome (LQTS). Twenty-two congenital LQTS patients and 30 control subjects were included in this study. We measured serum Mg (SMg) level and Mg retention (MgR) level, and evaluated the role of Mg (a high MgR level reflects Mg deficiency in the body). The influence of intravenous Mg infusion on Mg level was evaluated. Relatively low SMg level and high MgR level (LQTS:control = 53:33%, p < 0.01) were recognized in congenital LQTS patients, but there was an overlap with controls. Mg supplementation did not shorten QT interval and there was no significant correlation between Mg levels and QTc interval. Patients with syncopal history showed a higher MgR level (syncope (+):syncope (-) = 70:46%, p - 0.01) and intravenous Mg infusion improved Mg deficiency. These results suggest that some (not all) congenital LQTS patients are in a Mg-deficient state, which may be associated with syncope, and Mg supplementation may prevent recurrent syncope in these patients. Because there are several subtypes of congenital LQTS, perhaps with genetic testing Mg deficiency may be identified as a significant cofactor in some forms, whereas in other forms it is not relevant.
  • Expression of parathyroid hormone-related protein (PTHrP) in multiple myeloma
    R Kitazawa, S Kitazawa, K Kajimoto, H Sowa, T Sugimoto, T Matsui, K Chihara, S Maeda
    PATHOLOGY INTERNATIONAL 52 1 63 - 68 2002年01月 [有り][無し]
     研究論文(学術雑誌) 
    Multiple myeloma is a plasma cell neoplasia often associated with multiple skeletal lesions and hypercalcemia. Several cytokines, including interleukin (IL)-1, IL-6 and tumor necrosis factor-beta (TNF-beta), derived from myeloma cells are thought to accelerate osteoclastic bone resorption and cause hypercalcemia through a paracrine mechanism. We report on a case of a 69-year-old man with multiple myeloma associated with hypercalcemia and advanced osteolytic lesions. After bisphosphonate treatment and MP (melphalan and prednisolone) therapy, the patient's serum calcium level was successfully but transiently recovered to the normal range. Biochemical analysis showed a remarkable increase in serum parathyroid hormone-related protein (PTHrP; 3.7 pmol/L) and IL-6 (22.0 pg/mL). On the other hand, parathyroid hormone and 1alpha,25(OH)(2) vitamin D-3 were suppressed. By immunohistochemistry and in situ hybridization on aspiration-biopsied bone marrow clot sections, PTHrP mRNA and protein were detected in the cytoplasm of myeloma cells. The rate of PTHrP-positive myeloma cells was estimated to be at least one-third. Since PTHrP can, as an endocrine factor, systemically act on bone and kidney, hypercalcemia in this case might have been caused through both local osteolytic hypercalcemia and humoral hypercalcemia of malignancy mechanisms.
  • [X-ray refraction contrast imaging].
    Yamazaki K, Matsui J, Kagoshima Y, Tusaka Y, Suzuki Y, Hirano M, Ohbayashi C, Kitazawa S, Kitazawa R, Maeda S, Fukushima K, Tamura S, Hishikawa Y, Nagai H, Katabuchi T, Sugiyama K
    Igaku butsuri : Nihon Igaku Butsuri Gakkai kikanshi = Japanese journal of medical physics : an official journal of Japan Society of Medical Physics 22 1 8 - 12 2002年 [有り][無し]
  • Characteristics of improved NOVA magnesium ion-selective electrode: changes of ionized magnesium values and reference interval in healthy children
    K Hoshino, K Ogawa, T Hishitani, R Kitazawa
    MAGNESIUM RESEARCH 14 3 203 - 210 2001年09月 [有り][無し]
     研究論文(学術雑誌) 
    Following the report of interference between the thiocyanate ion (SCN-) and NOVA's previous ion-selective electrode (ISE) for ionized magnesium (iMg(2+)), NOVA has developed a new ISE which eliminates the effect of SCN-. Two hundred and sixty healthy children were divided into two groups; those who had presented when using NOVA's previous ISE (group A; n=160) and those using NOVA's new ISE (group B; n=100). The mean iMg(2+) value and the mean iMg(2+) percent fraction (iMg(2+)/Serum Mg) were significantly higher in group B than in group A (0.59 +/-0.03 vs 0.54 +/-0.03 mmol/L for iMg(2+); p<0.001 and 64.8<plus/minus>3.1 vs 58.2 +/-4.1 per cent for iMg(2+) percent fraction; p<0.001). The mean serum SCN- level was 0.023<plus/minus>0.008 mmol/L in group A (n=8) and 0.0.21 +/-0.007 mmol/L in group B (n=12), and was not significantly different between the two groups. The suspected change of iMg(2+) value interfered by SCN- was 0.037mmol/L in group A. The difference of iMg(2+) percent fraction between two groups was higher at high serum magnesium (SMg) levels. The reference interval of iMg(2+) was 0.56-0.62 rnmo/L in healthy children with the NOVA's new ISE, and was constant irrespective of growth. The NOVA's previous iMd(2+) ISE may be interfered with mainly by SCN-. The newly designed ISE eliminated these effects especially at higher SMg levels.
  • Interleukin-1 beta stimulates transendothelial mobilization of human peripheral blood mononuclear cells with a potential to differentiate into osteoclasts in the presence of osteoblasts
    Y Tokukoda, S Takata, H Kaji, R Kitazawa, T Sugimoto, K Chihara
    ENDOCRINE JOURNAL 48 4 443 - 452 2001年08月 [有り][無し]
     研究論文(学術雑誌) 
    There is accumulating evidence that interleukin-1 (IL-1) levels are increased locally at the site of active bone resorption in a variety of diseases including osteoporosis, periodontal disease and rheumatoid arthritis. However, the pathogenic role of IL-1 in bone loss remains to be fully elucidated. We present here additional evidence that IL-1 beta enhances endothelial activation and thereby stimulates mobilization of peripheral blood mononuclear cells (PBMCs) from luminal to abluminal spaces across the endothelium. Furthermore, IL-1 beta stimulates the differentiation of PBMCs into osteoclast-like cells with bone-resorbing activity in the presence of human osteoblastic SaOS-2 cells without systemic hormones. These findings provide circumstantial evidence for the hypothesis that IL-1 beta generated in the bone microenviroment plays a stimulatory role in PBMC mobilization from the peripheral circulation and their subsequent differentiation into osteoclast-like cells in the bone tissue. In addition, the present study supports the notion that osteoclast progenitor cells might be derived from the peripheral circulating blood mononuclear cells in human.
  • H Tamada, R Kitazawa, K Gohji, S Kitazawa
    JOURNAL OF BONE AND MINERAL RESEARCH 16 3 487 - 496 2001年03月 [有り][無し]
     研究論文(学術雑誌) 
    Bone morphogenetic proteins (BMPs), belonging to the transforming growth factor-beta (TGF-beta) superfamily, are multifunctional molecules that regulate bone induction and organ development. Among BMPs, BMP-6 has been shown to be overexpressed in prostate cancer and is speculated to be associated with bone-forming skeletal metastasis. We investigated the regulatory mechanism of the BMP-6 gene expression in prostate cancer cell lines DU-145, LNCaP, PC-3, and PC-3M with regard to the methylation status of the CpG island in the 5' Banking region of the human BMP-6 gene. By sequence-specific analysis of methylated cytosines, we show here that the methylation status of the CpG loci around the Sp1 site of the BMP-6 promoter is related to its steady-state expression and an alternative splicing of messenger RNA (mRNA) in prostate cancer cell lines. Furthermore, a study of clinical cases of benign and malignant prostate lesion by in situ hybridization showed that BMP-6 expression was high at both primary and secondary sites in cases of advanced cancer with metastasis. Demethylation of the CPG loci around the Spl binding site was shown in cases with high BMP-6 expression by sequencing analysis of the methylated cytosine from paraffin-embedded materials, Our results suggested that during cancer progression, besides inactivation of tumor suppressor genes by hypermethylation, activation of certain genes like BMP-6 by selective demethylation was a common epigenetic event giving a variable character to the invading and metastasizing cancer cells.
  • Methylation of CpG loci in 5 '-flanking region alters steady-state expression of adenomatous polyposis coli gene in colon cancer cell lines
    Y Sakamoto, R Kitazawa, S Maeda, S Kitazawa
    JOURNAL OF CELLULAR BIOCHEMISTRY 80 3 415 - 423 2001年 [有り][無し]
     研究論文(学術雑誌) 
    The APC genetic locus has been linked to the tumorigenesis and progression of colorectal cancer, although the precise mechanism of its involvement in this disease remains unknown. We used high sensitivity mapping of the methylated cytosine, Northern blot analysis and immunocytochemical staining in six colorectal cancer cell lines (DLD-1, SW480, Colo320, HT29, WiDr, and Colo201) to examine the relationship between the methylation status of the CpG loci in the 5'-flanking region of the APC gene and its expression. APC mRNA expression levels determined by Northern blot analysis correlated well with APC protein levels visualized by immunocytochemistry. In these colorectal cancer cell lines, no major genetic alterations of the APC gene, such as amplification or deletion, were detected. Analysis of the epigenetic control of APC gene expression in these lines revealed that methylation of the CpG loci in the 5'-untranslated region of APC mRNA repressed steady-state expression of the gene. Furthermore, epigenetic alteration of the APC gene was independent of the APC protein truncation and CpG methylation of the hMLH1 promoter. Although less eminent than protein truncation by point mutation within the coding region of the APC gene, epigenetic alteration suppressing APC gene expression may significantly contribute to oncogenesis and the progression of colorectal cancer. (C) 2001 Wiley-Liss, Inc.
  • K Takeuchi, K Murata, K Funaki, S Kitazawa, R Kitazawa
    GYNECOLOGIC ONCOLOGY 79 3 504 - 507 2000年12月 [有り][無し]
     研究論文(学術雑誌) 
    Background Humoral hypercalcemia of malignancy is a cancer-related hypercalcemia caused by the production of humoral factors by malignant cells in patients without bone metastases. Squamous cell carcinomas are the tumors most frequently associated with humoral hypercalcemia of malignancy, and parathyroid hormone-related protein (PTH-rP) is the main humoral factor implicated. Squamous cell carcinoma arising from mature cystic teratoma is a rare diagnosis itself, much less the description of associated hypercalcemia, despite the fact that the normal keratinocytes produce parathyroid hormone-related protein. Case We present a well-documented case of squamous cell carcinoma arising from mature cystic teratoma of the ovary, complicated by hypercalcemia in a patient with high levels of plasma parathyroid hormone-related protein and immunohistochemical evidence of parathyroid hormone-related protein expression by the tumor cells. Conclusion, in this case, the carcinoma cells had already produced PTH-rP in the primary tumor although the serum calcium levels had not been significantly high at surgery. It is therefore suggested that hypercalcemia may have occurred after PTH-rP production had overcome the homeostatic level during the terminal stage. (C) 2000 Academic Press.
  • S Yano, T Sugimoto, T Tsukamoto, K Chihara, A Kobayashi, S Kitazawa, S Maeda, R Kitazawa
    KIDNEY INTERNATIONAL 58 5 1980 - 1986 2000年11月 [有り][無し]
     研究論文(学術雑誌) 
    Background The down-regulation of both calcium-sensing receptor (CaSR) and vitamin D receptor (VDR) in parathyroid (PT) glands of secondary hyperparathyroidism (HPT) caused by chronic renal failure has been associated with PT hormone hypersecretion as well as PT hypergrowth. To clarify the predominance of decreased expression of CaSR and VDR in the high proliferative activity of PT glands, we examined the relationship between the expression of both receptors and proliferative activity in human PT glands. Methods. Serial sections of 56 PT glands, including 52 glands from secondary HPT and 4 normal PT glands resected together with thyroid carcinoma, were examined immunohistochemically with specific antibodies against CaSR, VDR, and Ki67. The Ki67-positive cell number was counted and expressed as the Ki67 score. The CaSR and VDR expressions were semiquantitatively analyzed. Results. The expressions of both CaSR and VDR were markedly decreased in PT glands of secondary HPT, while the Ki67 score was significantly higher than it was in normal controls. When hyperplastic glands were classified into two subgroups, with [N(+)] or without [N(-)] nodular formation, CaSR expression was significantly decreased in N(+), while VDR expression was not different. Multiple regression analyses revealed that the decreased expression of CaSR could contribute significantly to the high proliferative activity, even if VDR expression was taken into account. Conclusion. The decrease in CaSR expression is associated with the high proliferative activity of PT glands in secondary HPT, independently of the decreased VDR expression. These findings provide a new insight into the pathogenesis of PT hyperplasia, which is refractory to vitamin D therapy in patients with severe secondary HPT.
  • Identification of methylated cytosine from archival formalin-fixed paraffin-embedded specimens.
    Kitazawa S, Kitazawa R, Maeda S
    Laboratory investigation; a journal of technical methods and pathology 80 2 275 - 276 2000年02月 [有り][無し]
  • H Ueno, R Yoneda, W Ogawa, S Yoon, S Kitazawa, R Kitazawa, M Kasuga
    ACTA HAEMATOLOGICA 104 4 212 - 216 2000年 [有り][無し]
     研究論文(学術雑誌) 
    A 59-year-old man with progressive and advanced agnogenic myeloid metaplasia, also called idiopathic my elofibrosis, had complications showing bilateral interstitial pneumonic shadows. Pathological assessment of transbronchial biopsy revealed pulmonary perivascular fibrosis and infiltration of megakaryocytes. Autopsy 3 months later showed extramedullary megakaryopoiesis and fibrosis in lung, pleura, kidney, liver and spleen. Histopathological analysis for platelet-derived growth factor (PDGF) and PDGF-receptor revealed an abnormally high expression of the PDGF-receptor-beta gene in pulmonary fibroblasts. This is the first description of an association between pulmonary fibrosis and PDGF in idiopathic myelofibrosis. Copyright (C) 2001 S. Karger AG, Basel.
  • Transcriptional regulation of rat cyclin D1 gene by CpG methylation status in promoter region
    S Kitazawa, R Kitazawa, S Maeda
    JOURNAL OF BIOLOGICAL CHEMISTRY 274 40 28787 - 28793 1999年10月 [有り][無し]
     研究論文(学術雑誌) 
    Cyclin D1, a G(1)/S cell cycle-regulating oncogene, is known to be transcriptionally regulated by numerous growth factors. We cloned and characterized the rat cyclin D1 gene 5'-flanking region and, by species- and subspecies-matched transient transfection studies, found that a basic promoter structure with a cAMP response element and two continuous Spl-binding sites was crucial for the steady-state expression of the cyclin D1 gene. Furthermore, the methylation status especially around two continuous Spl-binding sites was found to be an important epigenetical mechanism determining the steady-state expression level in rat leukemic cell lines K4D, K4DT, and K4D16. Whether or not epigenetic control of the cyclin D1 gene existed among normal rat tissues was further examined by high sensitivity mapping of the methylated cytosine, In normal rat tissues, the methylated cytosines at non-CpG loci within two continuous Spl-binding sites were observed in uterine stromal cells of the basal layer and found to be demethylated in the functioning layer, possibly by a passive demethylation mechanism through cell division. Since in the passive demethylation process Spl-binding sites remain methylated in a part of the cell population, methylated cytosines at Spl-binding sites may be essential for keeping a number of the stromal cells in the basal layer live against estrogen-induced proliferation that leads to either apoptosis or compaction.
  • Expression of platelet-derived growth factor proteins and their receptor alpha and beta mRNAs during fracture healing in the normal mouse.
    Fujii H, Kitazawa R, Maeda S, Mizuno K, Kitazawa S
    Histochemistry and cell biology 112 2 131 - 138 1999年08月 [有り][無し]
  • In situ demonstration of parathyroid hormone-related protein mRNA in sclerosing hepatic carcinoma
    R Kitazawa, S Kitazawa, S Yoon, M Kasuga, S Maeda
    VIRCHOWS ARCHIV-AN INTERNATIONAL JOURNAL OF PATHOLOGY 435 2 137 - 142 1999年08月 [有り][無し]
     研究論文(学術雑誌) 
    A 69-year-old man had a hepatic tumour occupying the left and half of the right lobe, with portal vein thrombus. There were hypercalcaemia and hypophosphataemia with increased nephrogenous cyclic adenosine monophosphate; bone metastases were excluded. Serum parathyroid hormone-related protein (PTHrP) was elevated, but no increase in intact parathyroid hormone (PTH) or vitamin Dg metabolites was found. At autopsy the histological features were typical of sclerosing hepatic carcinoma. By immunohistochemistry PTHrP was detected in cancer cell nests but not in the fibrous stroma. PTHrP transcripts were demonstrated by in situ hybridization using a polymerase chain reaction (PCR)-derived single-stranded DNA probe. Tumour cells expressed AE1 and CA19-9 (markers for cholangioepithelium) and CEA (for bile canaliculi). Electron microscopy revealed microvilli on the apical surface, and secretory granules 100 nm in diameter were observed. These findings indicate that this case is one of cholangiocellular sclerosing hepatic carcinoma. The interaction between cancer and stromal cells may be the cause of PTHrP overexpression.
  • Promoter structure of mouse RANKL/TRANCE/OPGL/ODF gene.
    Kitazawa R, Kitazawa S, Maeda S
    Biochimica et biophysica acta 1445 1 134 - 141 1999年04月 [有り][無し]
  • Hypoplastic left heart syndrome: Duration of survival without surgical intervention
    K Hoshino, K Ogawa, T Hishitani, R Kitazawa, R Uehara
    AMERICAN HEART JOURNAL 137 3 535 - 542 1999年03月 [有り][無し]
     研究論文(学術雑誌) 
    Background Mortality rate for heart transplantation for patients with hypoplastic left heart syndrome (HLHS) has improved, but there is a considerable wait until a suitable donor is available. Thus it is important to examine the duration of survival and risk factors for early death in patients with HLHS who did not undergo surgical intervention. Methods and Results Twenty-six consecutive patients were studied retrospectively. Duration of survival and the 14 following variables were investigated: date of birth, body weight at birth,cardiothoracic ratio, ascending aorta diameter, interatrial communication size, coarctation of the aorta, tricuspid regurgitation, anatomic subtype (patency) of mitral and aortic valve, arterial blood gas Findings (pH, PaO2, SaO(2), PaCO2, base excess), and ST depression in the electrocardiogram. Twenty patients survived <60 days (group A) and 6 patients survived beyond 60 days (group B). The duration of survival (mean [SD]) was 60 (151) days overall (1 patient is currently alive at 783 days). The long-term survivors (beyond 60 days) increased significantly after 1991 (P <.05). Coarctation of the aorta was a significant risk of early death (<60 days) (P <.05). Interatrial communication size was significantly smaller in group B than in group A (P <.05). The mean pH and base excess were significantly lower in group A than in group B. The other 9 variables showed no significant difference between the 2 groups. Conclusions There was a significant correlation of long-term survival with stabilized ductal blood flow without coarctation of the aorta, adequate restriction of interatrial communication without severe hypoxemia, and no metabolic acidosis.
  • In situ hybridization with polymerase chain reaction-derived single-stranded DNA probe and S1 nuclease.
    Kitazawa S, Kitazawa R, Maeda S
    Histochemistry and cell biology 111 1 7 - 12 1999年01月 [有り][無し]
  • Promoter structure of human sonic hedgehog gene.
    Kitazawa S, Kitazawa R, Tamada H, Maeda S
    Biochimica et biophysica acta 1443 3 358 - 363 1998年12月 [有り][無し]
  • Analysis of 5'-flanking region of human Smad4 (DPC4) gene.
    Minami R, Kitazawa R, Maeda S, Kitazawa S
    Biochimica et biophysica acta 1443 1-2 182 - 185 1998年11月 [有り][無し]
  • Undifferentiated pancreatic cancer associated with humoral hypercalcemia of malignancy
    S Kakizaki, N Ohya, T Yoshinaga, T Higuchi, R Kitazawa, H Takayama, H Takagi, T Nagamine, M Mori
    JAPANESE JOURNAL OF CLINICAL ONCOLOGY 28 9 563 - 566 1998年09月 [有り][無し]
     研究論文(学術雑誌) 
    We present a case of undifferentiated pancreatic cancer associated with humoral hypercalcemia of malignancy (HHM) in which parathyroid hormone-related protein (PTH-rP) is identified as the causative factor of hypercalcemia. A 61-year-old man was hospitalized with right hypochondralgia. Ultrasound examination and computed tomography demonstrated a large mass in the pancreatic head with liver metastases. Biopsy of the pancreatic tumor demonstrated undifferentiated carcinoma. Serum calcium level and PTH-rP were elevated. Bone scan with technetium-99 demonstrated no accumulation in the bones. Immunohistochemical staining for PTH-rP was weakly positive in the tumor cells. We considered that PTH-rP was the causative factor of HHM in this case from laboratory data and immunohistochemical findings. This rare case was successfully treated with pamidronate disodium, which is a type of bisphosphonate derivative. We compared this case with previously reported cases.
  • Type IIa early gastric cancer with proliferation of xanthoma cells
    A Muraoka, Suehiro, I, M Fujii, H Ueno, S Hayashi, K Shimizu, R Kitazawa, S Kitazawa, K Murakami
    JOURNAL OF GASTROENTEROLOGY 33 3 326 - 329 1998年06月 [有り][無し]
     研究論文(学術雑誌) 
    We report a type IIa early gastric cancer associated with xanthoma cell proliferation in a 61-year-old man. The patient was admitted to our hospital because of a gastric polyp detected at a medical checkup. An irregular protruding lesion with xanthoma cell proliferation was detected endoscopically. Histological examination showed a well differentiated tubular adenocarcinoma in the mucosa associated with xanthoma cell proliferation. The distribution of the xanthoma cells in the stroma corresponded closely with that of the cancer cells. Neither atypism nor mitotic figures were recognized in the xanthoma cells. In an immunohistochemical study, almost all the xanthoma cells were stained positive for alpha(1)-antitrypsin, while relatively few exhibited positive S-100 protein staining. Specific monocyte chemotactic and activating factor im munoreactivity was present only in the xanthoma cells, and not in the cancer cells. On the basis of these findings, it was speculated that the gastric cancer cells may have caused the xanthoma cell proliferation via an autocrine mechanism i.e., by a chemical mediator acting in a paracrine or juxtacrine manner.
  • M Kanatani, T Sugimoto, Y Takahashi, H Kaji, R Kitazawa, K Chihara
    JOURNAL OF BONE AND MINERAL RESEARCH 13 5 854 - 862 1998年05月 [有り][無し]
     研究論文(学術雑誌) 
    Several lines of evidence indicate that estrogen inhibits parathyroid hormone (PTH)-induced bone resorption in vivo and in vitro, However, its precise mechanism remains unknown, The present study was performed to investigate whether osteoclast precursor cells possess the receptors for PTH/PTH-related protein (PTHrP) and/or estrogen and to clarify the mechanism by which estrogen affects PTH-induced osteoclast-like cell (Ocl) formation. The polymerase chain reaction (PCR) product corresponding in size to the mouse PTH/PTHrP receptor cDNA was detected in mouse hemopoietic blast cells supported by granulocyte-macrophage colony-stimulating factor (GM-CSF) as well as in osteoblastic MC3T3-E1 cells. The nucleotide sequence of the PTH/PTHrP receptor PCR product of hemopoietic blast cells was found to be 95.4% identical to that of PTH/PTHrP receptor cDNA of rat osteoblastic ROS cells. The PCR product corresponding in size to the mouse estrogen receptor cDNA was detected in mouse hemopoietic blast cells supported by GM-CSF as web as in MC3T3-E1 cells, The nucleotide sequence of the estrogen receptor PCR product of hemopoietic blast cells was completely identical to that of mouse estrogen receptor cDNA. 17 beta-estradiol (17 beta-E-2) but not 17 alpha-E-2, dose dependently antagonized Ocl formation stimulated by human (h) PTR(1-34) at a minimal effective concentration of 10(-10) M in the hemopoietic blast cell culture. 17 beta-E-2 also significantly inhibited Ocl formation stimulated by 10(-8) M hPTHrP(1-34), while it did not affect 1,25-dihydroxyvitamin D-3-induced Ocl formation. EIolvever, 10(-8) M 17 beta-E-2 significantly inhibited Ocl formation stimulated by dibutyryladenosine cAMP (10(-4) M) and Sp-cAMPS (10(-4) M), an activator of cAMP-dependent protein kinase (PKA) as well as forskolin (10(-5) M). In contrast, 17 beta-E-2 did not affect Ocl formation by either phorbol myristate acetate (10(-7) hi), an activator of protein kinase C (PKC), or A23187 (10(-7) hi), a calcium ionophore, The pretreatment with 17 beta-E-2 significantly inhibited Ocl formation induced by the combined treatment with PTH and PKC inhibitors (H7 or staurosporine), while it did not affect Ocl formation stimulated by the combined treatment with RID and Rp-cAMPS, a PKA inhibitor. The present data indicate that estrogen inhibits RID-stimulated Ocl formation by directly acting on hemopoietic blast cells, possibly through blocking a PRA pathway but not a calcium/PKC pathway.
  • Ionized magnesium level in whole blood of healthy Japanese children.
    Hoshino K, Ogawa K, Kitazawa R, Nakamura Y, Uehara R
    Acta paediatrica Japonica : Overseas edition 40 2 116 - 121 1998年04月 [有り][無し]
  • Molecular cloning and analysis of the 5'-flanking region of the human bone morphogenetic protein-6 (BMP-6).
    Tamada H, Kitazawa R, Gohji K, Kamidono S, Maeda S, Kitazawa S
    Biochimica et biophysica acta 1395 3 247 - 251 1998年02月 [有り][無し]
  • K Nishida, R Kitazawa, K Mizuno, S Maeda, S Kitazawa
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 241 2 258 - 263 1997年12月 [有り][無し]
     研究論文(学術雑誌) 
    The integrin alpha 6 subunit associates with either the beta 1 or beta 4 subunit to form receptors for laminin, a major component of the basement membrane. Here, we characterized basal promoter of the human integrin alpha 6 subunit gene. The transcription start site, mapped by primer extension analysis, was 208 bp upstream of the translation start site. The promoter region lacked canonical TATA and GC boxes, but did contain a TATA-like sequence (GATAAA) 23 bp upstream of the major transcription start site. Consensus binding sites for Sp1 and the NF-kappa B complex were also present in the promoter region. A putative glucocorticoid/progesterone receptor responsive element (GRE/PRE), together with the Ap1 and c-myc binding sites located around 350-360 bp upstream of the transcription start site, represented positive regulatory sequences. Our current study showed a molecular model by which progesterone promotes tumor cell invasion through the basement membrane by up-regulating the laminin receptor. (C) 1997 Academic Press.
  • In situ detection of parathyroid hormone-related protein in ovarian clear cell carcinoma
    R Kitazawa, S Kitazawa, T Matui, S Maeda
    HUMAN PATHOLOGY 28 3 379 - 382 1997年03月 [有り][無し]
     研究論文(学術雑誌) 
    A case of ovarian clear fell carcinoma associated with hypercalcemia is reported. A 67-year-old woman developed the lung metastasis 8 months after primary surgery. The patient manifested symptoms of humoral hypercalcemia of malignancy (HHM) during the last 3 months of her clinical course. Serum and urinary C-terminus parathyroid hormone-related protein (PTHrP) levels were remarkably high. No increase in interleukin (IL) 1 beta, tumor necrosis factor (TNF) alpha, vitamin D-3 metabolites or intact PTH was detected. Pamidronate disodium treatment was transiently suppressed her serum calcium level. The patient died despite seven courses of chemotherapy. Autopsy showed multiorgan metastases and accelerated osteoclastic bone resorption; skeletal metastasis was not detected. Immunohistochemical analysis clearly showed the localization of PTHrP at both the primary and metastatic sites. The transcripts of PTHrP at pulmonary metastatic sites were revealed by in situ hybridization and the reverse transcription polymerase chain reaction (RT-PCR) method. PTHrP was the causative factor for HHM in this case. It is therefore suggested that hypercalcemia may have occurred after PTHrP production had overcome the homeostatic level during the terminal stage, although PTHrP production continued irrespective of the patient's serum calcium level. Copyright (C) 1997 by W.B. Saunders Company.
  • Rapid enlargement of cardiac rhabdomyoma during corticotropin therapy for infantile spasms
    T Hishitani, K Hoshino, K Ogawa, R Uehara, R Kitazawa, S Hamano, T Nara, Y Ogawa
    CANADIAN JOURNAL OF CARDIOLOGY 13 1 72 - 74 1997年01月 [有り][無し]
     研究論文(学術雑誌) 
    OBJECTIVE: To investigate the influence of corticotropin therapy on cardiac rhabdomyoma. DESIGN: Analysis of data from echocardiography performed on in-patients. PATIENTS: Six patients with rhabdomyoma who were admitted to the authors' medical centre with either convulsion (Ave cases) or prematurity (one case) between 1955 and 1995. Five had tuberous sclerosis. INTERVENTION: Size of cardiac rumours of each patient was measured by echocardiography, and volume index was calculated as the ratio of the tumour volume to its initial volume. MAIN RESULT: Increase in size of some of the rumours was found during corticotropin therapy on follow-up echocardiography. Maximum volume indexes of rumours in the case of patients (n=4) who did not receive corticotropin therapy was 1.2 to 3.7, whereas those of patients (n=2) who received therapy was 9.1 to 12; one of the latter patients died. CONCLUSION: Corticotropin may contribute to the enlargement of cardiac rhabdomyoma. The size of cardiac rhabdomyomas must he carefully followed when patients are treated with corticotropin.
  • M Sakaue, S Kitazawa, K Nishida, R Kitazawa, S Maeda
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 221 3 768 - 772 1996年04月 [有り][無し]
     研究論文(学術雑誌) 
    The bone morphogenic proteins (BMPs) constitute a novel subfamily of the transforming growth factor type beta (TGF-beta) supergene family and play a critical role in modulating mesenchymal differentiation and inducing the processes of cartilage and bone formation. In this study we isolated the 5'-flanking region of the human BMP-5 gene. Nucleotide sequencing, primer extension, DNase I foot printing and functional analysis by transient expression showed that the cloned 1.5 kb promoter region contains two transcription start sites, a canonical TATA box (-17 similar to -12) and a number of consensus recognition sequences including GATA-1 (-582 similar to -576) and engrailed (-549 similar to -541). (C) 1996 Academic Press, Inc.
  • R KITAZAWA, RB KIMBLE, JL VANNICE, VT KUNG, R PACIFICI
    JOURNAL OF CLINICAL INVESTIGATION 94 6 2397 - 2406 1994年12月 [有り][無し]
     研究論文(学術雑誌) 
    To investigate the contribution of IL-1, IL-6, and TNF to the increased osteoclastogenesis induced by estrogen deficiency, ovariectomized (ovx) mice were treated with either IL-1 receptor antagonist (IL-1ra), a competitive inhibitor of IL-1, TNF binding protein (TNFbp), an inhibitor of TNF, or the anti-IL-6 antibody (Ab) 20F3 for the first 2 wk after surgery. ovx increased the bone marrow cells secretion of IL-1 and TNF, but not IL-6, and the formation of TRAP-positive osteoclast-like multinucleated cells (MNCs) in bone marrow cultures treated with 1,25(OH)(2)D-3. The increase in MNC formation induced by ovx was prevented by in vivo treatment with either 17 beta estradiol, IL-1ra, TNFbp, or anti IL-6 Ab. However, the percent change in MNC formation induced by the anti-IL-6 Ab was similar in ovx and sham-operated animals, whereas IL-1ra and TNFbp were effective only in ovx mice. MNC formation was also decreased by in vitro treatment of bone marrow cultures with IL-1ra and TNFbp, but not with anti-IL-6 Ab. Ovx also increased bone resorption in vivo and in vitro, as assessed by the urinary excretion of pyridinoline cross links and the formation of resorption pits, respectively. IL-1ra, TNFbp and estrogen decreased bone resorption in vivo and in vitro whereas the anti-IL-6 Ab inhibited bone resorption in vitro but not in vivo. In conclusion, these data indicate that IL-1 and TNF play a direct role in mediating the effects of ovx on osteoclastogenesis and bone resorption. The data also suggest that IL-6 is not essential for increasing bone resorption in the early postovariectomy period.
  • PERSISTENT BONE-SPARING EFFECT OF INTERLEUKIN-1 RECEPTOR ANTAGONIST - A HYPOTHESIS ON THE ROLE OF IL-1 IN OVARIECTOMY-INDUCED BONE LOSS
    RB KIMBLE, R KITAZAWA, JL VANNICE, R PACIFICI
    CALCIFIED TISSUE INTERNATIONAL 55 4 260 - 265 1994年10月 [有り][無し]
     研究論文(学術雑誌) 
    The recent finding that treatment with the interleukin-l (IL-I) inhibitor, interleukin-1 receptor antagonist (IL-1ra) decreases bone loss and bone resorption in ovariectomized rats, strongly suggested that IL-1 mediates, at least in part, the effects of estrogen deficiency on bone resorption. Although in vitro studies have shown that IL-1 activates mature osteoclasts and stimulates osteoclastogenesis, the two main mechanisms by which estrogen deficiency stimulates bone resorption, it is still unclear whether IL-1 mediates both effects of estrogen deficiency in vivo. To investigate this matter, we have examined the changes in bone mineral density (BMD) which occur in ovariectomized rats after completion of 1 month of estrogen or IL-1ra treatment begun at the time of ovariectomy. Ovariectomy caused a marked decreased in BMD which was blocked by 17 beta estradiol and decreased by IL-1ra. Cessation of estrogen therapy was followed by a rapid induction of bone loss, indicating that estrogen blocks the activation and utilization of mature osteoclasts without depleting the bone microenvironment of osteoclast precursors and mature, inactive osteoclasts. In contrast, ovariectomized rats treated with IL-1ra maintained a stable bone density for the first 4 weeks after completion of the treatment. In these rats, bone loss resumed not earlier than 6 weeks after discontinuation of the IL-1ra treatment. Estrogen deficiency was necessary to unveil the bone-sparing effect of IL-1ra because in a control experiment in which rats were treated with IL-1ra for the 4 weeks before ovariectomy, BMD began to decrease immediately after ovariectomy. Based on these results we propose the hypothesis that in conditions of estrogen deficiency, the main effect of IL-1ra is to block the proliferation and differentiation of osteoclast precursors, an event that results in the depletion of mature, rapidly responsive osteoclasts. We also suggest that estrogen may have important direct effects on the regulation of osteoclast activity.
  • Parathyroid Hormone-related Protein in Breast Cancer Tissues: Relationship between Primary and Metastatic Sites.
    Kohno N, Kitazawa S, Sakoda Y, Kanbara Y, Furuya Y, Ohashi O, Kitazawa R
    Breast cancer (Tokyo, Japan) 1 1 43 - 49 1994年07月 [有り][無し]
  • PARATHYROID HORMONE-RELATED PROTEIN IN GASTRIC CANCERS WITH HETEROTOPIC OSSIFICATION
    Y YAMAMURAIDEI, S KITAZAWA, R KITAZAWA, T FUJIMORI, T CHIBA, S MAEDA
    CANCER 72 6 1849 - 1852 1993年09月 [有り][無し]
     研究論文(学術雑誌) 
    Background. Parathyroid hormone-related protein (PTHrP) has been regarded as one of the substances causing humoral hypercalcemia of malignancy. Methods. The immunohistochemical localization of PTHrP was investigated in 33 cases of gastric cancer (4 with heterotopic ossification and 29 without heterotopic ossification) to clarify the role of PTHrP in heterotopic ossification by using the anti-PTHrP monoclonal antibody, 4B3. Results. The four cases with heterotopic ossification showed positive staining at primary or metastatic sites, and in one case fibroblasts in the stroma surrounding the heterotopic ossifying foci also showed positive. On the other hand, of the 29 cases without heterotopic ossification, only 5 showed positive staining. Conclusions. The presence of PTHrP in ossifying gastric carcinomas at a relatively high rate indicates that PTHrP also might be related to heterotopic ossification associated with malignancies. It is speculated that PTHrP would contribute to heterotopic ossification by facilitating the process of mineralization.
  • In situ hybridization using bromodeoxyuridine labelled DNA probe and RNase H.
    Kitazawa S, Kitazawa R, Gotoh A, Fujimori T, Maeda S
    Biotechnic & histochemistry : official publication of the Biological Stain Commission 68 3 137 - 141 1993年05月 [有り][無し]
  • COMMON EXPRESSION OF PARATHYROID HORMONE-RELATED PROTEIN AND NO CORRELATION OF CALCIUM LEVEL IN RENAL-CELL CARCINOMAS
    A GOTOH, S KITAZAWA, Y MIZUNO, A TAKENAKA, S ARAKAWA, O MATSUMOTO, R KITAZAWA, T FUJIMORI, S MAEDA, S KAMIDONO
    CANCER 71 9 2803 - 2806 1993年05月 [有り][無し]
     研究論文(学術雑誌) 
    Background. Parathyroid hormone-related protein (PTHrP) is the predominant cause of humoral hypercalcemia of malignancy (HHM). Methods. Using a PTHrP-specific monoclonal antibody (MoAb), 4B3, the authors investigated the immunohistochemical localization of PTHrP in formalin-fixed and paraffin-embedded sections of normal human kidney tissues and tissues from 42 human renal cell carcinomas obtained at operation or autopsy. Results. In normal renal tissues, the distal tubules and collecting ducts showed positive immunostaining. PTHrP was detected in 40 of 42 renal cell carcinoma tissues (95%). Histopathologically, the granular cell subtypes of renal cell carcinomas tended to be more strongly positive than the clear cell subtypes. There was no significant correlation between the level of immunostaining and each patient's serum calcium level. Conclusion. PTHrP was commonly observed in renal cell carcinomas, and no significant correlation was seen between the intensity of PTHrP staining and the serum calcium level.
  • [Immunohistological evaluation of parathyroid hormone-related protein in breast cancer with and without calcification on mammography].
    Kanbara Y, Kono N, Nakaya M, Ishikawa Y, Fujiwara O, Kitazawa R, Kitazawa S
    Nihon Geka Gakkai zasshi 94 4 394 - 399 1993年04月 [有り][無し]
  • A MEMBRANE-BOUND METALLOENDOPEPTIDASE FROM RAT-KIDNEY - ITS IMMUNOLOGICAL CHARACTERIZATION
    T YAMAGUCHI, H KIDO, R KITAZAWA, S KITAZAWA, M FUKASE, N KATUNUMA
    JOURNAL OF BIOCHEMISTRY 113 3 299 - 303 1993年03月 [有り][無し]
     研究論文(学術雑誌) 
    The structure and location of a membrane-bound metallo-endopeptidase, previously purified from rat kidney [Yamaguchi et al. (1991) Eur. J. Biochem. 200, 563-571], were examined by immunochemical and immunohistochemical methods with a rabbit polyclonal antibody against the purified enzyme. On treatment with endoglycosidase F, the subunit of the purified enzyme (molecular mass = 88 kDa) was converted to a smaller form (78.5 kDa), indicating that the enzyme contained at least 11% N-linked carbohydrate. Treatment of kidney membranes with papain resulted in release of the enzyme, as shown by Western blotting analysis of the solubilized fraction. Immunoassays of rat tissues showed that only the kidney, and small and large intestine expressed significant amounts of the antigen. Moreover, immunohistochemical studies showed that the antigen was confined to the luminal surfaces of the proximal renal tubules and the intestinal villi. Thus, like another kidney membrane metallo-endopeptidase, meprin [Kounnas et al. (1991) J. Biol. Chem. 266, 17350-17357], the purified enzyme is shown to be a glycoprotein that is probably anchored in the plasma membrane, and located in the luminal surface of microvillar membranes of the kidney and intestine. These results indicate that our enzyme and meprin have clear structural and topological similarities.
  • THE EXPRESSION OF PARATHYROID HORMONE-RELATED PROTEIN (PTHRP) IN NORMAL PARATHYROID - HISTOCHEMISTRY AND INSITU HYBRIDIZATION
    R KITAZAWA, S KITAZAWA, M FUKASE, T FUJITA, A KOBAYASHI, K CHIHARA, S MAEDA
    HISTOCHEMISTRY 98 4 211 - 215 1992年11月 [有り][無し]
     研究論文(学術雑誌) 
    The expression and localization of parathyroid hormone-related protein (PTHrP), a major factor responsible for humoral hypercalcemia of malignancy (HHM), was investigated in 14 cases of surgically resected normal parathyroid glands. For light microscopic immunohistochemistry, formalin-fixed and paraffin-embedded specimens were stained with avidin-biotin-peroxidase complex (ABC) using the anti-PTHrP monoclonal antibody (MoAb), 4B3. Four percent paraformaldehyde (PFA)-fixed and OCT compound-embedded specimens were used for pre-embedded immunoelectron microscopy. For in situ hybridization, 4% PFA-fixed, frozen sections were studied using a bromodeoxyuridine (BrdU)-labeled PTHrP cDNA probe. Immunohistochemically, 12 of the 14 cases were positive for PTHrP, which was observed mainly in the oxyphil and transitional oxyphil cells. The chief and clear cells, on the other hand, were faintly positive. Electron microscopically, secretory granules positive for PTHrP were observed in cells containing abundant mitochondria. Consistent with the PTHrP immunoreactivity, transcripts of PTHrP were observed also in the oxyphilic cells by in situ hybridization. Thus the production and secretion of PTHrP was shown by the oxyphil cell lineage in the normal parathyroid glands.
  • IMMUNOHISTOCHEMICAL EVALUATION OF PARATHYROID HORMONE-RELATED PROTEIN (PTHRP) IN THE UTERINE CERVIX
    S KITAZAWA, R KITAZAWA, M FUKASE, T FUJIMORI, S MAEDA
    INTERNATIONAL JOURNAL OF CANCER 50 5 731 - 735 1992年03月 [有り][無し]
     研究論文(学術雑誌) 
    We investigated the immunohistochemical localization of parathyroid hormone-related protein (PTHrP), a major factor responsible for the humoral hypercalcemia of malignancy, in uterine cervical lesions. Formalin-fixed paraffin-embedded specimens from 16 cases of normal and reactive conditions, 45 cases of cervical intra-epithelial neoplasm (CIN) and 63 cases of invasive cancer were studied immunohistochemically by the avidine-biotin-peroxidase method, using an anti-PTHrP monoclonal antibody (MAb), 4B3. In normal and reactive conditions, PTHrP was positive in parabasal cells, squamous metaplasia, and hyperplastic reserve cells. In neoplastic conditions, 96% (43/45) of invasive squamous-cell carcinomas were positive for PTHrP, regardless of the patients' serum calcium levels. Two cases with hypercalcemia were strongly positive for PTHrP and showed prominent stromal interaction of the scirrhous type. In CIN, including koilocytic atypia, 76% (32/42) of cases were positive for PTHrP. In contrast, 91% (10/11) of adenocarcinomas were negative for PTHrP. In conclusion, we found, first, that in non-neoplastic conditions, the presence of PTHrP was correlated with the transformation of progenitor cells into squamous epithelia and with the maturation of keratinocytes and, second, that in squamous-cell carcinoma, the degree of keratinization and stromal interaction was higher in direct proportion to the apparent incidence of detectable PTHrP.
  • EFFECTS OF CONTINUOUS INFUSION OF PARATHYROID-HORMONE AND PARATHYROID HORMONE-RELATED PEPTIDE ON RAT BONE INVIVO - COMPARATIVE-STUDY BY HISTOMORPHOMETRY
    R KITAZAWA, Y IMAI, M FUKASE, T FUJITA
    BONE AND MINERAL 12 3 157 - 166 1991年03月 [有り][無し]
     研究論文(学術雑誌) 
    We have investigated the actions of parathyroid hormone (PTH) and PTH-related peptide (PTHrP) on the bones of parathyroidectomized (PTX) rats by histomorphometric analysis. Miniosmotic pumps filled with either human PTH (hPTH)(1-34), hPTHrP(1-34) or vehicle were subcutaneously implanted on the backs of the rats. The peptides were continuously infused for 6 days at a rate of 15 nmole/kg/day. PTH and PTHrP exhibited similar hypercalcemic and hypophosphatemic actions on these PTX rats. No significant differences were noted in bone weight or calcium and phosphorus contents of the ashed bone among the 3 groups. By quantitative histomorphometric analysis, hPTH(1-34) and hPTHrP(1-34) were found similarly to enhance both bone formation and resorption. Peritrabecular fibrosis was observed only in the PTH-infused animals. PTHrP thus mimics the actions of PTH, but is not as effective in promoting mesenchymal cell proliferation along the bone trabeculae.
  • IMMUNOHISTOLOGIC EVALUATION OF PARATHYROID HORMONE-RELATED PROTEIN IN HUMAN LUNG-CANCER AND NORMAL TISSUE WITH NEWLY DEVELOPED MONOCLONAL-ANTIBODY
    S KITAZAWA, M FUKASE, R KITAZAWA, A TAKENAKA, A GOTOH, T FUJITA, S MAEDA
    CANCER 67 4 984 - 989 1991年02月 [有り][無し]
     研究論文(学術雑誌) 
    With a newly developed monoclonal anti-PTHrP antibody, 4B3, the immunohistochemical localization of the parathyroid hormone-related protein (PTHrP) was studied on the formalin-fixed and paraffin-embedded sections of normal human tissues and various subtypes of lung cancer. Among normal epithelial tissues, keratinocytes in squamous epithelia, transitional and bronchial epithelia with squamous metaplasia, meningoepithelial cells, and mammary ductal cells with lactating changes showed positive immunoreactivity. Also, among endocrine tissues, cells in the parathyroid gland, pancreatic islets, adrenal cortex, pituitary gland, and testis were sporadically positive for PTHrP. These distribution patterns suggested that in a physiologic condition, PTHrP was closely related to keratinization and local secretion and/or the metabolism of calcium in specifically differentiated tissues. In lung cancer, however, PTHrP was detected in all cases of well-differentiated and moderately differentiated squamous cell carcinoma and in most cases of small cell carcinoma, irrespective of the patients' serum calcium level. However, PTHrP was not detected in two of five cases of poorly differentiated squamous cell carcinoma and in all cases of adenocarcinoma. Consequently, it was found that PTHrP was commonly produced by squamous cell carcinomas of the differentiated type, and that humoral hypercalcemia of malignancy could be induced when the PTHrP transgressed the homeostatic mechanisms.

MISC

  • Kaoru Funaki, Katsuhiro Sawada, Hidenobu Fukunishi, Riko Kitazawa, Sohei Kitazawa INTERNATIONAL JOURNAL OF GYNECOLOGY & OBSTETRICS その他 100 (3) 282 -283 2008年03月 [無し][無し]
  • Michiko Takahashi, Yutaka Takahashi, Kenichi Takahashi, Fyodor N. Zolotaryov, Kyoung Su Hong, Riko Kitazawa, Keiji Iida, Yasuhiko Okimura, Hidesuke Kaji, Sohei Kitazawa, Masato Kasuga, Kazuo Chihara FEBS LETTERS 582 (5) 573 -578 2008年03月 [無し][無し]
     
    To explore a novel adipokine, we screened adipocyte differentiation-related gene and found that TIG2/chemerin was strongly induced during the adipocyte differentiation. Chemerin was secreted by the mature 3T3-L1 adipocytes and expressed abundantly in adipose tissue in vivo as recently described. Intriguingly, the expression of chemerin was differently regulated in the liver and adipose tissue in db/db mice. In addition, serum chemerin concentration was decreased in db/db mice. Chemerin and its receptor/ChemR23 were expressed in mature adipocytes, suggesting its function in autocrine/paracrine fashion. Finally, chemerin potentiated insulin-stimulated glucose uptake concomitant with enhanced insulin signaling in the 3T3-L1 adipocytes. These data establish that chemerin is a novel adipokine that regulates adipocyte function. (C) 2008 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
  • Kaoru Funaki, Katsuhiro Sawada, Hidenobu Fukunishi, Riko Kitazawa, Sohei Kitazawa INTERNATIONAL JOURNAL OF GYNECOLOGY & OBSTETRICS その他 100 (3) 282 -283 2008年03月 [無し][無し]
  • Michiko Takahashi, Yutaka Takahashi, Kenichi Takahashi, Fyodor N. Zolotaryov, Kyoung Su Hong, Riko Kitazawa, Keiji Iida, Yasuhiko Okimura, Hidesuke Kaji, Sohei Kitazawa, Masato Kasuga, Kazuo Chihara FEBS LETTERS 582 (5) 573 -578 2008年03月 [無し][無し]
     
    To explore a novel adipokine, we screened adipocyte differentiation-related gene and found that TIG2/chemerin was strongly induced during the adipocyte differentiation. Chemerin was secreted by the mature 3T3-L1 adipocytes and expressed abundantly in adipose tissue in vivo as recently described. Intriguingly, the expression of chemerin was differently regulated in the liver and adipose tissue in db/db mice. In addition, serum chemerin concentration was decreased in db/db mice. Chemerin and its receptor/ChemR23 were expressed in mature adipocytes, suggesting its function in autocrine/paracrine fashion. Finally, chemerin potentiated insulin-stimulated glucose uptake concomitant with enhanced insulin signaling in the 3T3-L1 adipocytes. These data establish that chemerin is a novel adipokine that regulates adipocyte function. (C) 2008 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
  • Takeshi Kondo, Riko Kitazawa, Akira Yamaguchi, Sohei Kitazawa JOURNAL OF CELLULAR BIOCHEMISTRY 103 (1) 335 -345 2008年01月 [無し][無し]
     
    Increased bone fragility attributed to osteopenia is a serious side effect of glucocorticoid treatment. Glucocorticoid-induced bone loss is caused primarily by hypofunction and apoptosis of osteoblasts, and secondarily by accelerated bone resorption. To explore the mechanism whereby dexamethasone (Dex) stimulates osteoclastogenesis in the coculture system, we analyzed the effect of Dex on the expression of both mouse osteoprotegerin (OPG) and receptor activator of NF-kappa B ligand (RANKL). Dex reduced OPG transcripts and OPG protein secretion by the ST2 osteoblastic cells. Since mainly the c-Jun homodimer maintains the steady-state transcription of the OPG gene, we examined the effect of Dex on c-Jun signaling in ST2 cells. Western blotting disclosed that Dex decreased the amount of phospho-c-Jun protein (p-c-Jun) and, correspondingly, the amount of the phosphorylated p46 isoform of Jun N-terminal kinase (JNK). The amount of phospho-SEK1 also decreased after Dex treatment, while the amounts of phospho-ERK and p38 remained constant. Among mitogen-activated protein (MAP) kinase inhibitors, the JNK inhibitor mimicked the inhibitory effect of Dex on OPG promoter activity. On the other hand, Dex treatment per se showed a nominal increase of RANKL gene expression. A part of Dex-mediated OPG gene suppression was achieved by the suppression of beta-catenin signaling. We speculate therefore that the bone resorptive action of Dex is mediated mainly by the inhibition of OPG by transrepressing the OPG gene through the AP-1 site, with a reduction (mediated mainly by the decrease in the p46 isoform of JNK) in the proportion of p-c-Jun in a JNK-dependent manner.
  • Takeshi Kondo, Riko Kitazawa, Akira Yamaguchi, Sohei Kitazawa JOURNAL OF CELLULAR BIOCHEMISTRY 103 (1) 335 -345 2008年01月 [無し][無し]
     
    Increased bone fragility attributed to osteopenia is a serious side effect of glucocorticoid treatment. Glucocorticoid-induced bone loss is caused primarily by hypofunction and apoptosis of osteoblasts, and secondarily by accelerated bone resorption. To explore the mechanism whereby dexamethasone (Dex) stimulates osteoclastogenesis in the coculture system, we analyzed the effect of Dex on the expression of both mouse osteoprotegerin (OPG) and receptor activator of NF-kappa B ligand (RANKL). Dex reduced OPG transcripts and OPG protein secretion by the ST2 osteoblastic cells. Since mainly the c-Jun homodimer maintains the steady-state transcription of the OPG gene, we examined the effect of Dex on c-Jun signaling in ST2 cells. Western blotting disclosed that Dex decreased the amount of phospho-c-Jun protein (p-c-Jun) and, correspondingly, the amount of the phosphorylated p46 isoform of Jun N-terminal kinase (JNK). The amount of phospho-SEK1 also decreased after Dex treatment, while the amounts of phospho-ERK and p38 remained constant. Among mitogen-activated protein (MAP) kinase inhibitors, the JNK inhibitor mimicked the inhibitory effect of Dex on OPG promoter activity. On the other hand, Dex treatment per se showed a nominal increase of RANKL gene expression. A part of Dex-mediated OPG gene suppression was achieved by the suppression of beta-catenin signaling. We speculate therefore that the bone resorptive action of Dex is mediated mainly by the inhibition of OPG by transrepressing the OPG gene through the AP-1 site, with a reduction (mediated mainly by the decrease in the p46 isoform of JNK) in the proportion of p-c-Jun in a JNK-dependent manner.
  • H. Fukunishi, K. Funaki, K. Ikuma, Y. Kaji, K. Sugimura, R. Kitazawa, S. Kitazawa INTERNATIONAL JOURNAL OF GYNECOLOGICAL CANCER 17 (3) 724 -728 2007年05月 [無し][無し]
     
    Uterine leiomyosarcoma, initially diagnosed as leiomyoma on magnetic resonance (MR) images, was disclosed after focused ultrasound surgery (FUS). The tumor did not display high signal intensity on either T1- or T2-weighted images on the patient's first visit. Four months thereafter, T2-weighted images revealed a high signal intensity area within the tumor, while T1-weighted images showed low signal intensity. Six months after FUS, the nonperfused volume calculated on meglumine gadoterate-enhanced T1-weighted images decreased markedly and an intermediate signal intensity in a circular area on T2-weighted images appeared to be atypically increasing in volume. After laparoscopic myomectomy, this tumor was diagnosed as uterine leiomyosarcoma coexistent with leiomyoma. The early stages of uterine leiomyosarcoma are clinically difficult to diagnose; therefore, both careful monitoring during FUS and close follow-up after the procedure are vital.
  • Yasuhiro Hamada, Sohei Kitazawa, Riko Kitazawa, Hideki Fujii, Masato Kasuga, Masafumi Fukagawa BONE 40 (5) 1408 -1414 2007年05月 [無し][無し]
     
    Diabetic osteopenia causes an increase in bone fracture and a delay in healing of fractures, and affects the quality of life. However, the mechanisms responsible for the disease have not been clearly identified. Oxidative stress may be a potential candidate for the pathogenesis, since it is increased under diabetic conditions and is known to induce cellular dysfunction in a wide variety of cell types. Although in vitro studies have shown that oxidative stress inhibits osteoblastic differentiation and induces osteoblast insults and apoptosis, the relationship between diabetic osteopenia and oxidative stress remains unclear. To explore these issues, analysis of a mouse model that represents the diabetic osteopenia as seen in patients with diabetes is necessary. However, there are few reports of such a model. Therefore, we focused on the streptozotocin (STZ)-induced diabetic mouse, one of the most common animal models of type I diabetes. Eight-week-old male C57BL/6 mice were randomly assigned to the following three groups: 1) control group, 2) diabetic group, and 3) insulin-treated diabetic group. After 12 weeks of STZ treatment, the physical properties of the femora, and the static and dynamic parameters of bone histomorphometry of the tibiae from STZ-induced diabetic mice (STZ-mice) were assessed, and oxidative stress in the whole body and bone of the mice was evaluated. Renal function was comparable in all three groups at the end of the experimental period. In addition, no significant difference in serum PTH, Ca, and P was found among the three groups. In contrast, radiological analysis demonstrated a significant decrease in trabecular bone volume, and histomorphometric analyses confirmed that parameters for both bone formation (OV/BV, OS/BS, and BFR/BS) and bone resorption (ES/BS and Oc.S/BS) were also significantly lower in STZ-mice. In addition, urinary excretion of 8-hydroxydeoxyguano sine, a marker of oxidative DNA damage, was elevated in STZ-mice. Further immunohistological studies showed intensified immunostaining of an oxidative stress marker in bone tissue including the osteoblasts of diabetic mice. Here, we demonstrated that STZ-mice exhibit low-turnover osteopenia associated with increased oxidative stress. (c) 2006 Elsevier Inc. All rights reserved.
  • H. Fukunishi, K. Funaki, K. Ikuma, Y. Kaji, K. Sugimura, R. Kitazawa, S. Kitazawa INTERNATIONAL JOURNAL OF GYNECOLOGICAL CANCER 17 (3) 724 -728 2007年05月 [無し][無し]
     
    Uterine leiomyosarcoma, initially diagnosed as leiomyoma on magnetic resonance (MR) images, was disclosed after focused ultrasound surgery (FUS). The tumor did not display high signal intensity on either T1- or T2-weighted images on the patient's first visit. Four months thereafter, T2-weighted images revealed a high signal intensity area within the tumor, while T1-weighted images showed low signal intensity. Six months after FUS, the nonperfused volume calculated on meglumine gadoterate-enhanced T1-weighted images decreased markedly and an intermediate signal intensity in a circular area on T2-weighted images appeared to be atypically increasing in volume. After laparoscopic myomectomy, this tumor was diagnosed as uterine leiomyosarcoma coexistent with leiomyoma. The early stages of uterine leiomyosarcoma are clinically difficult to diagnose; therefore, both careful monitoring during FUS and close follow-up after the procedure are vital.
  • Yasuhiro Hamada, Sohei Kitazawa, Riko Kitazawa, Hideki Fujii, Masato Kasuga, Masafumi Fukagawa BONE 40 (5) 1408 -1414 2007年05月 [無し][無し]
     
    Diabetic osteopenia causes an increase in bone fracture and a delay in healing of fractures, and affects the quality of life. However, the mechanisms responsible for the disease have not been clearly identified. Oxidative stress may be a potential candidate for the pathogenesis, since it is increased under diabetic conditions and is known to induce cellular dysfunction in a wide variety of cell types. Although in vitro studies have shown that oxidative stress inhibits osteoblastic differentiation and induces osteoblast insults and apoptosis, the relationship between diabetic osteopenia and oxidative stress remains unclear. To explore these issues, analysis of a mouse model that represents the diabetic osteopenia as seen in patients with diabetes is necessary. However, there are few reports of such a model. Therefore, we focused on the streptozotocin (STZ)-induced diabetic mouse, one of the most common animal models of type I diabetes. Eight-week-old male C57BL/6 mice were randomly assigned to the following three groups: 1) control group, 2) diabetic group, and 3) insulin-treated diabetic group. After 12 weeks of STZ treatment, the physical properties of the femora, and the static and dynamic parameters of bone histomorphometry of the tibiae from STZ-induced diabetic mice (STZ-mice) were assessed, and oxidative stress in the whole body and bone of the mice was evaluated. Renal function was comparable in all three groups at the end of the experimental period. In addition, no significant difference in serum PTH, Ca, and P was found among the three groups. In contrast, radiological analysis demonstrated a significant decrease in trabecular bone volume, and histomorphometric analyses confirmed that parameters for both bone formation (OV/BV, OS/BS, and BFR/BS) and bone resorption (ES/BS and Oc.S/BS) were also significantly lower in STZ-mice. In addition, urinary excretion of 8-hydroxydeoxyguano sine, a marker of oxidative DNA damage, was elevated in STZ-mice. Further immunohistological studies showed intensified immunostaining of an oxidative stress marker in bone tissue including the osteoblasts of diabetic mice. Here, we demonstrated that STZ-mice exhibit low-turnover osteopenia associated with increased oxidative stress. (c) 2006 Elsevier Inc. All rights reserved.
  • Yutaka Takahashi, Keiji Iida, Kentaro Takahashi, Shiro Yoshioka, Hidenori Fukuoka, Ryoko Takeno, Mari Imanaka, Hitoshi Nishizawa, Michiko Takahashi, Yasushi Seo, Yoshitake Hayashi, Takuma Kondo, Yasuhiko Okimura, Hidesuke Kaji, Riko Kitazawa, Sohei Kitazawa, Kazuo Chihara GASTROENTEROLOGY 132 (3) 938 -943 2007年03月 [無し][無し]
     
    Background & Aims: Nonalcoholic steatohepatitis (NASH) is an emerging progressive hepatic disease and demonstrates steatosis, inflammation, and fibrosis. Insulin resistance is a common feature in the development of NASH. Molecular pathogenesis of NASH consists of 2 steps: triglyceride accumulation in hepatocytes with insulin resistance and an enhanced oxidative stress caused by reactive oxygen species. Interestingly, NASH demonstrates a striking similarity to the pathologic conditions observed in adult growth hormone deficiency (AGHD). AGHD is characterized by decreased lean body mass, increased visceral adiposity, abnormal lipid profile, and insulin resistance. Moreover, liver dysfunctions with hyperlipidemia and nonalcoholic fatty liver disease (NAFLD) are frequently observed in patients with AGHD, and it is accompanied by metabolic syndrome. Methods: We studied a case diagnosed as NASH with hyperlipidemia in AGHD. The effect of GH-replacement therapy on the patient was analyzed. Results: Six months of GH-replacement therapy in the patient drastically ameliorated NASH and the abnormal lipid profile concomitant with a marked reduction in oxidative stress. Conclusions: These results suggest that GH plays an essential role in the metabolic and redox regulation in the liver.
  • Yutaka Takahashi, Keiji Iida, Kentaro Takahashi, Shiro Yoshioka, Hidenori Fukuoka, Ryoko Takeno, Mari Imanaka, Hitoshi Nishizawa, Michiko Takahashi, Yasushi Seo, Yoshitake Hayashi, Takuma Kondo, Yasuhiko Okimura, Hidesuke Kaji, Riko Kitazawa, Sohei Kitazawa, Kazuo Chihara GASTROENTEROLOGY 132 (3) 938 -943 2007年03月 [無し][無し]
     
    Background & Aims: Nonalcoholic steatohepatitis (NASH) is an emerging progressive hepatic disease and demonstrates steatosis, inflammation, and fibrosis. Insulin resistance is a common feature in the development of NASH. Molecular pathogenesis of NASH consists of 2 steps: triglyceride accumulation in hepatocytes with insulin resistance and an enhanced oxidative stress caused by reactive oxygen species. Interestingly, NASH demonstrates a striking similarity to the pathologic conditions observed in adult growth hormone deficiency (AGHD). AGHD is characterized by decreased lean body mass, increased visceral adiposity, abnormal lipid profile, and insulin resistance. Moreover, liver dysfunctions with hyperlipidemia and nonalcoholic fatty liver disease (NAFLD) are frequently observed in patients with AGHD, and it is accompanied by metabolic syndrome. Methods: We studied a case diagnosed as NASH with hyperlipidemia in AGHD. The effect of GH-replacement therapy on the patient was analyzed. Results: Six months of GH-replacement therapy in the patient drastically ameliorated NASH and the abnormal lipid profile concomitant with a marked reduction in oxidative stress. Conclusions: These results suggest that GH plays an essential role in the metabolic and redox regulation in the liver.
  • Nephrology Dialysis Transplantation Vol. 22, No. 6, pp. 1547-57 2007年 [無し][無し]
  • Riko Kitazawa, Sohei Kitazawa MOLECULAR ENDOCRINOLOGY 21 (1) 148 -158 2007年01月 [無し][無し]
     
    Receptor activator of nuclear factor-kappa B ligand (RANKL) expression is tissue specific and limited to certain subsets of T-lymphocytes and stromal/osteoblastic cells. Even among osteoblasts, RANKL is expressed on about 20% of osteoblasts of the normal mouse. To clarify the mechanism of population-specific RANKL expression, we analyzed the effect of CpG methylation on its transcription, mRNA and protein expression as well as on osteoclastogenesis. Subpopulations of ST2 cells were used: P9, which expresses RANKL and supports osteoclastogenesis, and P16, which does not. By sodium bisulfite mapping, the rate of CpG methylation of the -65/+350 region, especially of CpG locus no. 1 three bases upstream of the TATA-box, was higher in P16 than in P9 ST2 cells. ChIP and gel shift assay showed that methylated CpG locus no. 1 was a target of MeCP2 binding that, in turn, blocked the binding of the TATA-box binding protein to the TATA-box. In vitro methylation by SssI of the promoter construct reduced its transcriptional activity at the steady state and its response to 1 alpha,25(OH)(2) vitamin D-3. Conversely, treatment with DNA methylase inhibitor, 5-aza-2'-deoxycytidine, significantly restored RANKL expression and osteoclastogenesis in P16 cells. Except for primary cultured osteoblasts, CpG locus no. 1 was frequently methylated in various normal mouse tissues. We propose that the methylation status of the CpG locus three bases upstream of the TATA-box modulates the control of cell- and tissue-specific expression of RANKL gene and osteoclastogenesis. The heterogeneity of stromal/osteoblastic cells in response to bone-resorbing stimuli may be attributed, in part, to the methylation status of the RANKL gene promoter.
  • Nephrology Dialysis Transplantation Vol. 22, No. 6, pp. 1547-57 2007年 [無し][無し]
  • Sohei Kitazawa, Atsushi Takenaka, Takeshi Kondo, Akira Mizoguchi, Riko Kitazawa HISTOCHEMISTRY AND CELL BIOLOGY 126 (6) 665 -677 2006年12月 [無し][無し]
     
    We established a monoclonal antibody (MAb), 5G9, with the use of a fixed seminoma tissue from an archival paraffin-embedded specimen, as an immunogen. Without antigen retrieval, positive 5G9-immunohistochemical staining was confined mostly to primordial germ cells, spermatogonia and various germ cell tumors. 5G9 recognized a mitochondrial 32-kD protein with an isoelectric point of pH 4.2, identified as a multifunctional ubiquitous protein, receptor for globular head of C1q (gC1qR), whose epitope was mapped in a disordered loop connecting the beta 3 and the beta 4 strands. Reflecting the ubiquitous distribution of gC1qR, with antigen retrieval, 5G9 was found reactive to a wide range of normal and tumor tissues. Since several co-precipitated and phosphorylated bands were observed in various human cell lines but not in germ cell tumor cell lines by in vitro phosphorylation assay, we speculate that the epitope of gC1qR is specifically unmasked in the germ cell lineage. By reducing gC1qR by siRNA, a significant increase was observed in the number of apoptotic cells in ITO-II and TCam-2 cell lines, but to a lesser extent in the Colo201 colon cancer cell line, showing an antiapoptotic property of gC1qR in the germ cells. Since protein-protein interaction is partially preserved by fixation, archival paraffin-embedded specimens can be a valuable source of immunogens for generating monoclonal antibodies (MAbs) that recognize tissue-specific protein conformation.
  • Riko Kitazawa, Sohei Kitazawa, Yoshihiro Nishimura, Takeshi Kondo, Chiho Obayashi PATHOLOGY INTERNATIONAL 56 (8) 449 -452 2006年08月 [無し][無し]
     
    Pulmonary carcinosarcoma, consisting of both carcinoma and sarcoma with a heterologous element, is a rare subtype, comprising approximately 0.3% of primary lung neoplasia. A 57-year-old man was admitted because of severe dyspnea. A tumor wholly occupying the right thorax was biopsied and diagnosed as pleomorphic sarcoma. The tumor did not respond to chemotherapy, and the patient died of respiratory failure and sepsis. At autopsy, pleomorphic sarcoma was histologically dominant and contained a liposarcoma element confirmed by histocytological and electron microscopic analysis. Adenocarcinoma component with papillary and tubular patterns was confined to the medial lesion of the right lower lobe (3 x 8 cm), which was found in the chest X-ray 3 years before admission, and had continuously merged with the sarcomatous lesion through the histological transition of both components. Aggressive and rapid growth of the sarcoma derived from the earlier adenocarcinoma became prevalent and contributed to the severe clinical outcome. This is the first documented case of primary lung carcinosarcoma with a liposarcoma element.
  • Kiyoshi Mori, Riko Kitazawa, Takeshi Kondo, Sakan Maeda, Akira Yamaguchi, Sohei Kitazawa JOURNAL OF CELLULAR BIOCHEMISTRY 98 (6) 1629 -1644 2006年08月 [無し][無し]
     
    Runx2 regulates the target genes characteristic of osteoblastic phenotypes, while exerting diverse and sometimes controversial effects on osteoblastic cells depending on their differentiation stage. Receptor activator of nuclear factor-KB (RANK) ligand (RANKL) is a membrane bound cytokine essential for osteo(chondro)clastogenesis. During endochondral ossification, while Runx2-positive hypertrophic chondrocytes express RANKL, the steady-state expression of the RANKL gene in osteoblastic cells is, at later stages, kept at a relatively low level to sustain the established bone. The aim of this study was to elucidate the mechanism whereby Runx2 and the protein kinase A (PKA) pathway modulate RANKL expression, especially from the viewpoint of their functions in RANKL basic promoter activity and in chromatin structural changes in osteoblastic/stromal cells. Osteoblastic/stromal cell lines derived from normal and Runx2-deficient mice were used to analyze endogenous RANKL gene expression by real-time reverse transcription (RT)-PCR, the acetylation status of the H3 and H4 histone proteins associated with the 5'-flanking region of the RANKL gene by chromatin immunoprecipitation, and the exogenously transfected RANKL gene promoter activity both in the steady-state and under PKA-activated conditions. Here, we demonstrate that Runx2 suppresses steady-state RANKL gene expression by condensing chromatin, while showing a slightly positive effect on RANKL basic promoter activity. Besides acting through the CRE-like region (-0.96 kb) of the RANKL gene promoter, forskolin (FK) treatment transactivates the RANKL gene by antagonizing the function of Runx2, by reducing Runx2 mRNA expression and by opening the chromatin conformation far upstream (more than 40 kb) of the RANKL gene.
  • Riko Kitazawa, Sohei Kitazawa, Yoshihiro Nishimura, Takeshi Kondo, Chiho Obayashi PATHOLOGY INTERNATIONAL 56 (8) 449 -452 2006年08月 [無し][無し]
     
    Pulmonary carcinosarcoma, consisting of both carcinoma and sarcoma with a heterologous element, is a rare subtype, comprising approximately 0.3% of primary lung neoplasia. A 57-year-old man was admitted because of severe dyspnea. A tumor wholly occupying the right thorax was biopsied and diagnosed as pleomorphic sarcoma. The tumor did not respond to chemotherapy, and the patient died of respiratory failure and sepsis. At autopsy, pleomorphic sarcoma was histologically dominant and contained a liposarcoma element confirmed by histocytological and electron microscopic analysis. Adenocarcinoma component with papillary and tubular patterns was confined to the medial lesion of the right lower lobe (3 x 8 cm), which was found in the chest X-ray 3 years before admission, and had continuously merged with the sarcomatous lesion through the histological transition of both components. Aggressive and rapid growth of the sarcoma derived from the earlier adenocarcinoma became prevalent and contributed to the severe clinical outcome. This is the first documented case of primary lung carcinosarcoma with a liposarcoma element.
  • Kiyoshi Mori, Riko Kitazawa, Takeshi Kondo, Sakan Maeda, Akira Yamaguchi, Sohei Kitazawa JOURNAL OF CELLULAR BIOCHEMISTRY 98 (6) 1629 -1644 2006年08月 [無し][無し]
     
    Runx2 regulates the target genes characteristic of osteoblastic phenotypes, while exerting diverse and sometimes controversial effects on osteoblastic cells depending on their differentiation stage. Receptor activator of nuclear factor-KB (RANK) ligand (RANKL) is a membrane bound cytokine essential for osteo(chondro)clastogenesis. During endochondral ossification, while Runx2-positive hypertrophic chondrocytes express RANKL, the steady-state expression of the RANKL gene in osteoblastic cells is, at later stages, kept at a relatively low level to sustain the established bone. The aim of this study was to elucidate the mechanism whereby Runx2 and the protein kinase A (PKA) pathway modulate RANKL expression, especially from the viewpoint of their functions in RANKL basic promoter activity and in chromatin structural changes in osteoblastic/stromal cells. Osteoblastic/stromal cell lines derived from normal and Runx2-deficient mice were used to analyze endogenous RANKL gene expression by real-time reverse transcription (RT)-PCR, the acetylation status of the H3 and H4 histone proteins associated with the 5'-flanking region of the RANKL gene by chromatin immunoprecipitation, and the exogenously transfected RANKL gene promoter activity both in the steady-state and under PKA-activated conditions. Here, we demonstrate that Runx2 suppresses steady-state RANKL gene expression by condensing chromatin, while showing a slightly positive effect on RANKL basic promoter activity. Besides acting through the CRE-like region (-0.96 kb) of the RANKL gene promoter, forskolin (FK) treatment transactivates the RANKL gene by antagonizing the function of Runx2, by reducing Runx2 mRNA expression and by opening the chromatin conformation far upstream (more than 40 kb) of the RANKL gene.
  • K Kishimoto, R Kitazawa, M Kurosaka, S Maeda, S Kitazawa HISTOCHEMISTRY AND CELL BIOLOGY 125 (5) 593 -602 2006年05月 [無し][無し]
     
    Based on developmental fate and function, cartilage tissue is broadly classified into transient cartilage (e.g. growth plate, GP) and permanent cartilage (e.g. articular cartilage, AC). The former eventually disappears and is replaced by bone during the endochondral ossification process, whereas the latter retains its permanency. Osteo(chondro)clasts, multinucleated giant cells of the monocyte/macrophage lineage, are selectively induced in the GP during endochondral ossification and play central roles in the resorption of cartilagenous matrices. The aim of this study was to investigate the factors determining the GP-specific recruitment of osteo(chondro)clasts. We especially focused on the expression pattern of the receptor activator of NF-kappa B ligand (RANKL), an essential factor for osteo(chondro)clast differentiation, and on that of epigenetic and transcriptional factors affecting RANKL gene expression. Knee joints of male BALB/c mice aged 8 weeks were dissected and subjected to immunohistochemical analysis using anti-RANKL, Runx2, Dlx5 and Msx2 antibodies. The methylation status of the mouse RANKL gene promoter in both the GP and the AC was analyzed by sodium bisulfite mapping using microdissected mouse tissue. The expression of BMP-2, -3, -4, -6 and type X collagen mRNA was examined by in situ hybridization (ISH). At the boundary between the calcifying cartilage and the hypertrophic chondrocytes of the GP, RANKL-expressing chondrocytes overlapped those expressing Runx2, Dlx5 and Msx2, near numerous osteo(chondro)clasts. Although similar BMP-2 and -4 expression was observed in chondrocytes in both the GP and the AC as well as in maturing osteoblasts, a rather restricted BMP-6 expression pattern was observed in resting and proliferating chondrocytes in the GP. On the other hand, sodium bisulfite mapping showed that mostly non-CpG methylation was similarly scattered in a non-specific manner in chondrocytes in the GP and the AC. Taken together with the fact that putative Runx2 binding elements are located in the RANKL promoter, our data suggest that Runx2, an essential transcription factor for skeletal development, is also a key regulator of RANKL expression in chondrocytes in the GP. Furthermore, a selective and sequential expression of a subset of BMP and of transcription factors may define the expression pattern of RANKL through Runx2.
  • K Kishimoto, R Kitazawa, M Kurosaka, S Maeda, S Kitazawa HISTOCHEMISTRY AND CELL BIOLOGY 125 (5) 593 -602 2006年05月 [無し][無し]
     
    Based on developmental fate and function, cartilage tissue is broadly classified into transient cartilage (e.g. growth plate, GP) and permanent cartilage (e.g. articular cartilage, AC). The former eventually disappears and is replaced by bone during the endochondral ossification process, whereas the latter retains its permanency. Osteo(chondro)clasts, multinucleated giant cells of the monocyte/macrophage lineage, are selectively induced in the GP during endochondral ossification and play central roles in the resorption of cartilagenous matrices. The aim of this study was to investigate the factors determining the GP-specific recruitment of osteo(chondro)clasts. We especially focused on the expression pattern of the receptor activator of NF-kappa B ligand (RANKL), an essential factor for osteo(chondro)clast differentiation, and on that of epigenetic and transcriptional factors affecting RANKL gene expression. Knee joints of male BALB/c mice aged 8 weeks were dissected and subjected to immunohistochemical analysis using anti-RANKL, Runx2, Dlx5 and Msx2 antibodies. The methylation status of the mouse RANKL gene promoter in both the GP and the AC was analyzed by sodium bisulfite mapping using microdissected mouse tissue. The expression of BMP-2, -3, -4, -6 and type X collagen mRNA was examined by in situ hybridization (ISH). At the boundary between the calcifying cartilage and the hypertrophic chondrocytes of the GP, RANKL-expressing chondrocytes overlapped those expressing Runx2, Dlx5 and Msx2, near numerous osteo(chondro)clasts. Although similar BMP-2 and -4 expression was observed in chondrocytes in both the GP and the AC as well as in maturing osteoblasts, a rather restricted BMP-6 expression pattern was observed in resting and proliferating chondrocytes in the GP. On the other hand, sodium bisulfite mapping showed that mostly non-CpG methylation was similarly scattered in a non-specific manner in chondrocytes in the GP and the AC. Taken together with the fact that putative Runx2 binding elements are located in the RANKL promoter, our data suggest that Runx2, an essential transcription factor for skeletal development, is also a key regulator of RANKL expression in chondrocytes in the GP. Furthermore, a selective and sequential expression of a subset of BMP and of transcription factors may define the expression pattern of RANKL through Runx2.
  • 頸管ポリープとして経過観察されていた子宮腺肉腫の1例
    診断病理 23巻, 2号, pp. 136-139 2006年 [無し][無し]
  • PHOTO REPORT くも膜下出血で発症した左後頭蓋窩線維性髄膜腫
    内科 98巻, 2号, pp. 352-352 2006年 [無し][無し]
  • The Journal Of Clinical Investigation Vol. 116, No. 6, pp. 1494-1505 2006年 [無し][無し]
  • 【骨形成・骨吸収の最近のトピックス】 骨吸収・骨形成の相互作用 RANKL,OPGの転写制御
    THE BONE 20巻, 3号, pp. 321-326 2006年 [無し][無し]
  • Pathology International Vol. 56, No. 9, pp. 554-7 2006年 [無し][無し]
  • S Kitazawa, R Kitazawa JOURNAL OF STRUCTURAL BIOLOGY 153 (1) 64 -72 2006年01月 [無し][無し]
     
    In situ hybridization (ISH) is a widely applied technique used for visualizing specific nucleic acid sequences at chromosomal, cytologic, and histologic levels. It sometimes fails, however, to demonstrate precise cell identity, early stages of gene expression and variants of altNSernative splicing because of its limited resolution. To overcome this shortcoming, we have developed an improved ISH technique at the electron microscopic (EM) level by conducting en bloc hybridization before embedding (pre-embedding) and immuno-EM detection after ultra-thin sectioning (post-embedding). We applied this technique to demonstrate both the dynamic expression of interleukin (IL)-6 mRNA immediately after lipopolysaccharide (LPS) treatment, and the static expression of osteonectin mRNA in a differentiating osteoblastic cell linage. Tissue samples were diced into 1 mm cubes, fixed with 4% paraformaldehyde, and then successively hybridized en bloc with the digoxigenin (DIG)-labeled single-stranded probe measuring 200-300 bp with the aid of microwave treatment. After washing, for EM observation, the cubes were embedded in epon for ultra-thin sectioning, and a gold-colloid-labeled anti-DIG antibody was used for post-embedding immuno-EM; some of the cubes was directly incubated with anti-DIG antibody and developed en bloc for stereoscopic and light microscopic observation. IL-6 mRNA during and immediately after transcription was demonstrated in the nuclei of the alveolar macrophages and in neutrophils of mouse lung tissue as early as 15 min after LPS treatment, which was of better sensitivity than that by Northern blot or nuclear run-on techniques. Moreover, in mouse calvaria tissue, osteonectin mRNA both in the nucleus and the cytoplasm was observed in a differentiating osteoblastic cell linage in a differentiation-specific manner. This technique is useful in identifying specific cell types during and immediately after transcribing specific mRNA based on ultrastructural morphology. (C) 2005 Elsevier Inc. All rights reserved.
  • The Journal Of Clinical Investigation Vol. 116, No. 6, pp. 1494-1505 2006年 [無し][無し]
  • Pathology International Vol. 56, No. 9, pp. 554-7 2006年 [無し][無し]
  • Histochemistry And Cell Biology Vol. 126, No. 6, pp. 665-77 2006年 [無し][無し]
  • CS Shin, SJ Her, JA Kim, DH Kim, SW Kim, SY Kim, HS Kim, KH Park, JG Kim, R Kitazawa, SL Cheng, R Civitelli JOURNAL OF BONE AND MINERAL RESEARCH 20 (12) 2200 -2212 2005年12月 [無し][無し]
     
    We studied the effects of dominant negative N-cadherin (NCad Delta C) expression in ST2 cells on their ability to support osteoclastogenesis. Expression of NCad Delta C in ST2 cells did not decrease cell-to-cell adhesion but significantly reduced osteoclast formation when co-cultured with BMMs. NCad Delta C inhibited beta-catenin/TCF signaling, resulting in decreased RANKL expression, which could contribute to the reduced osteoclast formation.
  • CS Shin, SJ Her, JA Kim, DH Kim, SW Kim, SY Kim, HS Kim, KH Park, JG Kim, R Kitazawa, SL Cheng, R Civitelli JOURNAL OF BONE AND MINERAL RESEARCH 20 (12) 2200 -2212 2005年12月 [無し][無し]
     
    We studied the effects of dominant negative N-cadherin (NCad Delta C) expression in ST2 cells on their ability to support osteoclastogenesis. Expression of NCad Delta C in ST2 cells did not decrease cell-to-cell adhesion but significantly reduced osteoclast formation when co-cultured with BMMs. NCad Delta C inhibited beta-catenin/TCF signaling, resulting in decreased RANKL expression, which could contribute to the reduced osteoclast formation.
  • CS Shin, SJ Her, JA Kim, DH Kim, SW Kim, SY Kim, HS Kim, KH Park, JG Kim, R Kitazawa, SL Cheng, R Civitelli JOURNAL OF BONE AND MINERAL RESEARCH 20 (12) 2200 -2212 2005年12月 [無し][無し]
     
    We studied the effects of dominant negative N-cadherin (NCad Delta C) expression in ST2 cells on their ability to support osteoclastogenesis. Expression of NCad Delta C in ST2 cells did not decrease cell-to-cell adhesion but significantly reduced osteoclast formation when co-cultured with BMMs. NCad Delta C inhibited beta-catenin/TCF signaling, resulting in decreased RANKL expression, which could contribute to the reduced osteoclast formation.
  • CS Shin, SJ Her, JA Kim, DH Kim, SW Kim, SY Kim, HS Kim, KH Park, JG Kim, R Kitazawa, SL Cheng, R Civitelli JOURNAL OF BONE AND MINERAL RESEARCH 20 (12) 2200 -2212 2005年12月 [無し][無し]
     
    We studied the effects of dominant negative N-cadherin (NCad Delta C) expression in ST2 cells on their ability to support osteoclastogenesis. Expression of NCad Delta C in ST2 cells did not decrease cell-to-cell adhesion but significantly reduced osteoclast formation when co-cultured with BMMs. NCad Delta C inhibited beta-catenin/TCF signaling, resulting in decreased RANKL expression, which could contribute to the reduced osteoclast formation.
  • S Kitazawa, R Kitazawa, C Obayashi, T Yamamoto JOURNAL OF BONE AND MINERAL RESEARCH 20 (8) 1472 -1477 2005年08月 [無し][無し]
     
    Introduction: Desmoid-type fibromatosis, originating from mesenchymal cells with myofibroblastic features, is a locally aggressive and frequently recurring infiltrative lesion. One such sporadic case with metaplastic ossification in the chest wall is presented. Materials and Methods: A 43-year-old man was referred to the hospital with a gradually enlarging hard mass in the left anterolateral chest wall. A thoracotomy was carried out, and histopathological specimens were used for immunohistochemical, genetic, and methylation studies. Results: Accumulation of altered P-catenin associated with a somatic heterozygous activating mutation in codon 41 was detected in the typical desmoid-type fibromatosis and at the ossifying focus. Among factors related to bone formation and the classical wnt-beta-catenin signaling pathway, BMP and activin membrane-bound inhibitor (BAMBI) expression was specifically downregulated at the ossifying focus. Hypermethylation of the BAMBI promoter was observed in microdissected tissue from the ossifying focus but not in that from the typical desmoid-type fibromatosis. Conclusions: Because both BMP and classical Wnt/beta-catenin/LEF1 signaling cooperatively and mutually induce differentiation of mesenchymal cells into osteoblastic cells and promote bone formation, the epigenetic event leading to the enhanced responsiveness to BMP signaling may play a crucial role in the formation of metaplastic bone.
  • M Fujimoto, R Kitazawa, S Maeda, S Kitazawa ONCOGENE 24 (32) 5108 -5118 2005年07月 [無し][無し]
     
    Nerve growth factor and its high-affinity receptor TrkA are thought to be involved in the progression of various cancers. This study investigated the mechanism that regulates aberrant or increased TrkA expression in various cancer cell lines and in the course of pancreatic cancer progression. W e found that the negative cis-acting AP-1-like sequence TGAGCGA was located in the 5'-untranslated region of the TrkA gene. Sodium bisulfite mapping revealed that steady-state TrkA expression correlated positively with the accumulation of methylated CpG around the AP-1-like site. Electrophoretic mobility shift assay showed that the AP-1- like site was bound mainly by c-Jun homodimers; the binding was directly blocked by Sss I methylase-induced methylation or by an excess of oligonucleotides containing consensus AP-1 sequences. Consequently, activation of TrkA gene expression by methylation was considered to be caused by the direct interference of c-Jun binding to the negatively regulating AP-1- like site. Further more, the accumulation of methylated CpG around the AP-1- like site was also observed with increased TrkA immunohistochemical staining in cases of advanced pancreatic adenocarcinoma with extensive perineural invasion. Unlike global methylation at CpG islands that leads to gene silencing, specific methylation at non-CpG islands would play a crucial epigenetic role in the versatility and plasticity of TrkA expression during cancer progression.
  • M Fujimoto, R Kitazawa, S Maeda, S Kitazawa ONCOGENE 24 (32) 5108 -5118 2005年07月 [無し][無し]
     
    Nerve growth factor and its high-affinity receptor TrkA are thought to be involved in the progression of various cancers. This study investigated the mechanism that regulates aberrant or increased TrkA expression in various cancer cell lines and in the course of pancreatic cancer progression. W e found that the negative cis-acting AP-1-like sequence TGAGCGA was located in the 5'-untranslated region of the TrkA gene. Sodium bisulfite mapping revealed that steady-state TrkA expression correlated positively with the accumulation of methylated CpG around the AP-1-like site. Electrophoretic mobility shift assay showed that the AP-1- like site was bound mainly by c-Jun homodimers; the binding was directly blocked by Sss I methylase-induced methylation or by an excess of oligonucleotides containing consensus AP-1 sequences. Consequently, activation of TrkA gene expression by methylation was considered to be caused by the direct interference of c-Jun binding to the negatively regulating AP-1- like site. Further more, the accumulation of methylated CpG around the AP-1- like site was also observed with increased TrkA immunohistochemical staining in cases of advanced pancreatic adenocarcinoma with extensive perineural invasion. Unlike global methylation at CpG islands that leads to gene silencing, specific methylation at non-CpG islands would play a crucial epigenetic role in the versatility and plasticity of TrkA expression during cancer progression.
  • MEN1遺伝子の変異を同定できない乳癌合併MEN1症例におけるMeninの発現及び機能解析
    2005年 [無し][無し]
  • 乳癌合併とガストリノーマ肝転移を認めたMENI遺伝子変異を同定できないMENIの一例
    2005年 [無し][無し]
  • 多発性内分泌腺腫症I型及び続発性副甲状腺機能亢進症患者副甲状腺細胞におけるMeninの発現及び機能の検討
    2005年 [無し][無し]
  • 骨軟骨研究領域における再生医学の動向と病理学
    2005年 [無し][無し]
  • 異所骨形成を認めた胸壁デスモイド腫瘍の1症例
    2005年 [無し][無し]
  • 外陰部転移で発見されたmicropapillary componentを伴う甲状腺乳頭癌の1剖検例
    2005年 [無し][無し]
  • 単クローン抗体5G9の認識するヒト胚細胞特異抗原の解析
    2005年 [無し][無し]
  • 副腎皮質ホルモンによるマウス破骨細胞抑制因子(OPG)発現調節機構の解析
    2005年 [無し][無し]
  • レジオネラ感染症の2例
    2005年 [無し][無し]
  • DNAメチル化による破骨細胞分化因子(RANKL)発現制御
    2005年 [無し][無し]
  • 11B3モノクローナル抗体を用いた種々の癌腫における免疫組織学的検討
    2005年 [無し][無し]
  • MEN遺伝子の変異を同定できない乳癌合併MEN1症例におけるmeninの発現及び機能解析
    2005年 [無し][無し]
  • 胚細胞および胚細胞由来腫瘍を認識する単クローン抗体5G9の認識する抗原解析
    2005年 [無し][無し]
  • 遺伝子プロモータ領域CpGメチル化による破骨細胞分化因子(RANKL)の転写抑制機構
    2005年 [無し][無し]
  • 副腎皮質ホルモンによるマウス破骨細胞抑制因子(OPG)発現調節機構の解析
    2005年 [無し][無し]
  • Runx2はマウス骨芽細胞/間質細胞における破骨細胞分化因子(RANKL)発現を抑制的に調節する
    2005年 [無し][無し]
  • Runx2 Modulates RANKL Expression by Repressing Its Steady-state Level in Mouse Osteoblastic/Stromal Cells.
    2005年 [無し][無し]
  • Regulation of Mouse Osteoprotegerin Gene Expression by Glucocorticoids.
    2005年 [無し][無し]
  • Mouse RANKL Gene Transcription Is Reversibly Suppressed by CpG Methylation of its Promoter Region.
    2005年 [無し][無し]
  • 抗酸化ストレス物質チオレドキシンの糖尿病性腎症に及ぼす効果
    日本臨床分子医学会記録 41巻, pp.44-44 2005年 [無し][無し]
  • 【臨床分子内分泌学 甲状腺・副甲状腺・骨内分泌代謝系】 BMPs 基礎研究の進展 BMP遺伝子発現調節
    日本臨床 63巻, 増刊10, pp.409-413 2005年 [無し][無し]
  • MEN1遺伝子の変異を同定できない乳癌合併MEN1症例におけるmeninの発現及び機能解析
    日本内分泌学会雑誌 81巻, 1号, pp.83-83 2005年 [無し][無し]
  • 急速破壊性股関節症病態を示したオクロノーシス(組織黒変症)の1例
    診断病理 22巻, 3号, pp.193-195 2005年 [無し][無し]
  • 肺梗塞を起こして死亡した潰瘍性大腸炎の1例
    診断病理 22巻, 2号, pp.113-116 2005年 [無し][無し]
  • 脊髄アスペルギルス症と脊髄癆を合併した統合失調症の1例
    診断病理 23巻, 1号, pp.52-54 2005年 [無し][無し]
  • 脳室上衣腫合併MEN1型の一例と副甲状腺におけるMeninの検討
    日本内分泌学会雑誌 81巻, Suppl., pp.101-103 2005年 [無し][無し]
  • 多発性内分泌腺腫症I型及び続発性副甲状腺機能亢進症患者副甲状腺細胞におけるMeninの発現及び機能の検討
    日本骨代謝学会学術集会プログラム抄録集 pp.183-183 2005年 [無し][無し]
  • Ray tracing analysis of overlapping objects in refraction contrast imaging
    Radiation Medicine Vol. 23, No. 5, pp. 386-389 2005年 [無し][無し]
  • Estimation of contrast of refraction contrast imaging compared with absorption imaging-basic approach
    Radiation Medicine Vol. 23, No. 2, pp. 89-96 2005年 [無し][無し]
  • Neuropathology Vol. 25, No. 3, pp. 274-279 2005年 [無し][無し]
  • Desmoid tumor with ossification in chest wall: possible involvement of BAMBI promoter hypermethylation in metaplastic bone formation
    Journal Of Bone And Mineral Research Vol. 20, No. 8, pp. 1472-1477 2005年 [無し][無し]
  • Runx2 Modulates RANKL Expression by Repressing Its Steady-state Level in Mouse Osteoblastic/Stromal Cells.
    2005年 [無し][無し]
  • Regulation of Mouse Osteoprotegerin Gene Expression by Glucocorticoids.
    2005年 [無し][無し]
  • Mouse RANKL Gene Transcription Is Reversibly Suppressed by CpG Methylation of its Promoter Region.
    2005年 [無し][無し]
  • Ray tracing analysis of overlapping objects in refraction contrast imaging
    Radiation Medicine Vol. 23, No. 5, pp. 386-389 2005年 [無し][無し]
  • Estimation of contrast of refraction contrast imaging compared with absorption imaging-basic approach
    Radiation Medicine Vol. 23, No. 2, pp. 89-96 2005年 [無し][無し]
  • Neuropathology Vol. 25, No. 3, pp. 274-279 2005年 [無し][無し]
  • Journal Of Structural Biology Vol. 153, No. 1, pp. 64-72 2005年 [無し][無し]
  • Desmoid tumor with ossification in chest wall: possible involvement of BAMBI promoter hypermethylation in metaplastic bone formation
    Journal Of Bone And Mineral Research Vol. 20, No. 8, pp. 1472-1477 2005年 [無し][無し]
  • T Kondo, R Kitazawa, S Maeda, S Kitazawa JOURNAL OF BONE AND MINERAL RESEARCH 19 (9) 1411 -1419 2004年09月 [無し][無し]
     
    1alpha,25(OH)(2)D-3 rapidly and transiently suppressed OPG gene expression both by accelerating the degradation of mRNA and by suppressing promoter activity. The latter process was mediated through the AP-1 binding site by a reduction in the proportion of phospho-c-Jun in a JNK-independent manner. Introduction: Osteoclastogenesis is regulated by an integrated network of numerous bone metabolic factors, among which 1alpha,25-dihydroxyvitamin D-3 [1alpha,25(OH)(2)D-3] promotes osteoclastogenesis by reciprocally upregulating the expression of RANKL and downregulating that of osteoprotegerin (OPG). Materials and Methods: To analyze the mechanism by which 1alpha,25(OH)(2)D-3 suppresses OPG, we characterized cis-acting elements of the Mouse OPG gene and assessed the post-transcriptional modifications by actinomycin D assays. Results: 1alpha,25(OH)(2)D-3 rapidly and transiently suppressed OPG expression and shortened the half-life of OPG rnRNA; additionally, the c-Jun homodinier bound to the AP-1 binding site (TGACTGA, -293/-287) and maintained steady-state transcription of the OPG gene. Furthermore, mutation of the AP-1 site negated 1alpha,25(OH)(2)D-3-driven OPG Suppression. Moreover, 1alpha,25(OH)(2)D-3 treatment of ST2 cells decreased the amount of phosphorylated C-Jun protein (phospho-c-Jun), while the total amount of c-Jun remained constant; however, the amount of phosphorylated Jun N-terminal kinase (JNK) was nearly unchanged by 1alpha,25(OH)(2)D-3 treatment. Conclusion: Taken together with the observation that the OPG promoter has no consensus negative vitamin D-responsive elements. these data suggest that 1alpha,25(OH)(2)D-3 transrepresses mouse OPG by reducing the proportion of phospho-c-Jun in a JNK-independent manner. Our data indicated that short-term treatment with 1alpha,25(OH)(2)D-3 effectively downregulated OPG expression both by accelerating the degradation of OPG mRNA and by transrepressing the OPG gene through its AP-1 binding site in the catabolic phase. The OPG gene became insensitive to 1alpha,25(OH)(2)D-3 treatment. however, and reverted to its steady-state expression level over time, leading to the anabolic phase of the effect of 1alpha,25(OH)(2)D-3 on bone.
  • J Zhang, Y Lu, JL Dai, Z Yao, R Kitazawa, S Kitazawa, XP Zhao, DE Hall, KJ Pienta, ET Keller PROSTATE 59 (4) 360 -369 2004年06月 [無し][無し]
     
    BACKGROUND. Current animal models of prostate cancer (CaP) bone metastasis do not allow measurement of either tumor growth in bone over time or activation of gene promoters in intraosseous tumors. To develop these methods, we used bioluminescent imaging (BLI) to determine if expression of receptor activator of NF-kappaB ligand (RANKL), a pro-osteoclastogenic factor that promotes CaP bone metastases, is modulated by the bone matrix protein transforming growth factor-beta (TGF-beta) in vivo. METHODS. C4-2B human CaP cells were treated with TGF-beta in vitro and RANKL mRNA and protein production were Measured by polymerase chain reaction (PCR) and ELISA, respectively. Then C4-2B cells stably transfected with the RANKL promoter driving luciferase (lux) were injected intra-tibially into severe combined immundeficient (SCID) mice. Tumors were subjected to BLI every 2 weeks for 6 weeks and serum prostate specific antigen (PSA) was measured using ELISA. Vehicle (V), 1,25 dihydroxyvitamin D (VitD), or TGF-beta was administered to mice with established tumors and BLI to measure RANKL promoter activity was performed. Tumors were then subjected to immunohistochemistry for lux and assayed for RANKL mRNA levels. RESULTS. TGF-beta induced RANKL protein and mRNA expression and activated the RANKL promoter activity in a dose-dependent manner in vitro. BLI demonstrated an increase in intraosseous tumor size over time, which correlated with serum PSA levels. Administration of TGF-beta and VitD to mice with established intraosseous tumors increased lux activity compared to V. Intratibial tumor RANKL mRNA expression paralleled the increased promoter activity. Immunohistochemistry confirmed the presence of lux in the intraosseous tumors. CONCLUSIONS. These results demonstrate the ability to measure intraosseous tumor growth over time and gene promoter activation in an established intraosseous tumor in vivo and also demonstrate that TGF-beta induces activates the RANKL promoter. These results provide a novel method to explore the biology of CaP bone metastases. (C) 2003 Wiley-Liss, Inc.
  • J Zhang, Y Lu, JL Dai, Z Yao, R Kitazawa, S Kitazawa, XP Zhao, DE Hall, KJ Pienta, ET Keller PROSTATE 59 (4) 360 -369 2004年06月 [無し][無し]
     
    BACKGROUND. Current animal models of prostate cancer (CaP) bone metastasis do not allow measurement of either tumor growth in bone over time or activation of gene promoters in intraosseous tumors. To develop these methods, we used bioluminescent imaging (BLI) to determine if expression of receptor activator of NF-kappaB ligand (RANKL), a pro-osteoclastogenic factor that promotes CaP bone metastases, is modulated by the bone matrix protein transforming growth factor-beta (TGF-beta) in vivo. METHODS. C4-2B human CaP cells were treated with TGF-beta in vitro and RANKL mRNA and protein production were Measured by polymerase chain reaction (PCR) and ELISA, respectively. Then C4-2B cells stably transfected with the RANKL promoter driving luciferase (lux) were injected intra-tibially into severe combined immundeficient (SCID) mice. Tumors were subjected to BLI every 2 weeks for 6 weeks and serum prostate specific antigen (PSA) was measured using ELISA. Vehicle (V), 1,25 dihydroxyvitamin D (VitD), or TGF-beta was administered to mice with established tumors and BLI to measure RANKL promoter activity was performed. Tumors were then subjected to immunohistochemistry for lux and assayed for RANKL mRNA levels. RESULTS. TGF-beta induced RANKL protein and mRNA expression and activated the RANKL promoter activity in a dose-dependent manner in vitro. BLI demonstrated an increase in intraosseous tumor size over time, which correlated with serum PSA levels. Administration of TGF-beta and VitD to mice with established intraosseous tumors increased lux activity compared to V. Intratibial tumor RANKL mRNA expression paralleled the increased promoter activity. Immunohistochemistry confirmed the presence of lux in the intraosseous tumors. CONCLUSIONS. These results demonstrate the ability to measure intraosseous tumor growth over time and gene promoter activation in an established intraosseous tumor in vivo and also demonstrate that TGF-beta induces activates the RANKL promoter. These results provide a novel method to explore the biology of CaP bone metastases. (C) 2003 Wiley-Liss, Inc.
  • 副甲状腺腫瘍におけるTGFβ及びmeninの役割 MEN1患者検体を用いた解析
    2004年 [無し][無し]
  • 膵癌及び大腸癌細胞株におけるTrkAの発現調節機構
    2004年 [無し][無し]
  • 感染症対応剖検施設の建築 教育重視の観点から
    2004年 [無し][無し]
  • 感染症対応剖検施設の建築 安全性の観点から
    2004年 [無し][無し]
  • 感染症対応剖検施設の建築ー教育重視の観点から
    2004年 [無し][無し]
  • 感染症対応剖検施設の建築ー安全性の観点から
    2004年 [無し][無し]
  • ステロイドホルモンによるマウス破骨細胞抑制因子(OPG)発現調節機構の解析
    2004年 [無し][無し]
  • Pena-Shokeir Syndrome Type Ⅰの1剖検例
    2004年 [無し][無し]
  • 副甲状腺腫瘍におけるTGF beta 及びmeninの役割 MEN I患者検体を用いた解析
    2004年 [無し][無し]
  • DNAメチル化による破骨細胞分化因子(RANKL)発現制御
    2004年 [無し][無し]
  • 11B3モノクローナル抗体を用いた種々の癌腫における免疫組織学的検討
    2004年 [無し][無し]
  • 硬組織ISH 破骨細胞分化因子 (RANKL) 発現局在について
    2004年 [無し][無し]
  • 成長軟骨と関節軟骨における破骨細胞分化因子(RANKL)の発現
    2004年 [無し][無し]
  • ステロイドホルモンによるマウス破骨細胞抑制因子(OPG)発現調節機構の解析
    2004年 [無し][無し]
  • DNAメチル化による破骨細胞分化因子(RANKL)発現制御
    2004年 [無し][無し]
  • 脳室上衣腫合併MEN1型の一例と副甲状腺におけるMeninの検討
    2004年 [無し][無し]
  • Thioredoxin-1 overexpression attenuates diabetic nephropathy in streptozotocin-induced diabetic mice.
    2004年 [無し][無し]
  • Thioredoxin-1 Overexpression Attenuates Diabetic Nephropathy in Streptozotocin-Induced Diabetic Mice
    2004年 [無し][無し]
  • Regulation of Mouse Osteoprotegerin Gene Expression by Steroid Hormones.
    2004年 [無し][無し]
  • Expression of Mouse Receptor Activator of NF-kB Ligand (RANKL) in Growth Plate and Articular Cartilage.
    2004年 [無し][無し]
  • Epigenetic Regulation of Mouse Receptor Activator NF-KappaB Ligand (RANKL) Gene.
    2004年 [無し][無し]
  • Methylation around Negatively Regulating AP-1 Binding Site in TrkA Promoter Is Positively Associated with TrkA Gene Expression during Cancer Progression.
    2004年 [無し][無し]
  • In Situ Hybridization at Electron Microscopic Level for Detecting mRNA Immediately after Transcription.
    2004年 [無し][無し]
  • 鼻の巨細胞性線維芽細胞腫の1例(原著論文)
    診断病理 21巻, 4, pp. 300-302 2004年 [無し][無し]
  • Pena-Shokeir症候群I型の1剖検例
    診断病理 22巻, 1号, pp. 64-67 2004年 [無し][無し]
  • 異所骨形成を認めた胸壁デスモイド腫瘍の1症例
    日本病理学会会誌 94巻, 1号, pp. 281-281 2004年 [無し][無し]
  • 脳室上衣腫合併MEN1型の一例と副甲状腺におけるメニンの検討
    日本内分泌学会雑誌 80巻, 3号, pp. 663-663 2004年 [無し][無し]
  • 副甲状腺腫瘍におけるTGFβ及びmeninの役割 MEN1患者検体を用いた解析
    日本内分泌学会雑誌 80巻, 1号, pp. 90-90 2004年 [無し][無し]
  • T-138C Polymorphism of Matrix Gla Protein Promoter Alters Its Expression but is not Directly Associated with Atherosclerotic Vascular Calcification
    The Kobe Journal Of Medical Sciences Vol. 50, No. 3, pp. 69-81 2004年 [無し][無し]
  • Study on General Bacterial Contamination in the Nursery Environment─Focused on objects in playrooms of infants under 1 year of age─
    Medicine and Biology Vol. 148, No. 8, pp. 16-24 2004年 [無し][無し]
  • Acta Histochemica Et Cytochemica Vol. 38, No. 2, pp. 143-149 2004年 [無し][無し]
  • 病理診断における分子生物学 / 転写因子
    文光堂 2004年 [無し][無し]
  • 組織細胞化学2004 / In situ hybridization法の再生医学への応用
    学際企画 2004年 [無し][無し]
  • Thioredoxin-1 overexpression attenuates diabetic nephropathy in streptozotocin-induced diabetic mice.
    2004年 [無し][無し]
  • Thioredoxin-1 Overexpression Attenuates Diabetic Nephropathy in Streptozotocin-Induced Diabetic Mice
    2004年 [無し][無し]
  • Regulation of Mouse Osteoprotegerin Gene Expression by Steroid Hormones.
    2004年 [無し][無し]
  • Expression of Mouse Receptor Activator of NF-kB Ligand (RANKL) in Growth Plate and Articular Cartilage.
    2004年 [無し][無し]
  • Epigenetic Regulation of Mouse Receptor Activator NF-KappaB Ligand (RANKL) Gene.
    2004年 [無し][無し]
  • Methylation around Negatively Regulating AP-1 Binding Site in TrkA Promoter Is Positively Associated with TrkA Gene Expression during Cancer Progression.
    2004年 [無し][無し]
  • In Situ Hybridization at Electron Microscopic Level for Detecting mRNA Immediately after Transcription.
    2004年 [無し][無し]
  • T-138C Polymorphism of Matrix Gla Protein Promoter Alters Its Expression but is not Directly Associated with Atherosclerotic Vascular Calcification
    The Kobe Journal Of Medical Sciences Vol. 50, No. 3, pp. 69-81 2004年 [無し][無し]
  • Journal Of Bone And Mineral Research Vol. 19, No. 9, pp. 1411-1419 2004年 [無し][無し]
  • Acta Histochemica Et Cytochemica Vol. 38, No. 2, pp. 143-149 2004年 [無し][無し]
  • M Saijo, R Kitazawa, M Nakajima, M Kurosaka, S Maeda, S Kitazawa HISTOCHEMISTRY AND CELL BIOLOGY 120 (6) 493 -503 2003年12月 [無し][無し]
     
    Bone fracture healing takes place through endochondral ossification where cartilaginous callus is replaced by bony callus. Vascular endothelial growth factor (VEGF) is a requisite for endochondral ossification, where blood vessel invasion of cartilaginous callus is crucial. Heparanase is an endoglucuronidase that degrades heparan sulfate proteoglycans (HSPG) and releases heparin-binding growth factors including VEGF as an active form. To investigate the role of heparanase in VEGF recruitment during fracture healing, the expression of heparanase mRNA and VEGF, and vessel formation were examined in mouse fractured bone. On days 5 and 7 after the fracture, when mesenchymal cells proliferated and differentiated into chondrocytes, heparanase mRNA was detected in osteo(chondro)clasts and their precursors, but not in the inflammatory phase (day 3). On day 10, both VEGF and HSPG were produced by hypertrophic chondrocytes of the cartilaginous callus and by osteoblasts of the bony callus; numerous osteo(chondro)clasts resorbing the cartilage expressed strong heparanase signals. Adjacent to the cartilage resorption sites, angiogenesis with CD31-positive endothelial cells and osteogenesis with osteonectin-positive osteoblasts were observed. On days 14 and 21, osteoclasts in the woven bone tissue expressed heparanase mRNA. These data suggest that by producing heparanase osteo(chondro)clasts contribute to the recruitment of the active form of VEGF. Thus osteo(chondro)clasts may promote local angiogenesis as well as callus resorption in endochondral ossification during fracture healing.
  • M Saijo, R Kitazawa, M Nakajima, M Kurosaka, S Maeda, S Kitazawa HISTOCHEMISTRY AND CELL BIOLOGY 120 (6) 493 -503 2003年12月 [無し][無し]
     
    Bone fracture healing takes place through endochondral ossification where cartilaginous callus is replaced by bony callus. Vascular endothelial growth factor (VEGF) is a requisite for endochondral ossification, where blood vessel invasion of cartilaginous callus is crucial. Heparanase is an endoglucuronidase that degrades heparan sulfate proteoglycans (HSPG) and releases heparin-binding growth factors including VEGF as an active form. To investigate the role of heparanase in VEGF recruitment during fracture healing, the expression of heparanase mRNA and VEGF, and vessel formation were examined in mouse fractured bone. On days 5 and 7 after the fracture, when mesenchymal cells proliferated and differentiated into chondrocytes, heparanase mRNA was detected in osteo(chondro)clasts and their precursors, but not in the inflammatory phase (day 3). On day 10, both VEGF and HSPG were produced by hypertrophic chondrocytes of the cartilaginous callus and by osteoblasts of the bony callus; numerous osteo(chondro)clasts resorbing the cartilage expressed strong heparanase signals. Adjacent to the cartilage resorption sites, angiogenesis with CD31-positive endothelial cells and osteogenesis with osteonectin-positive osteoblasts were observed. On days 14 and 21, osteoclasts in the woven bone tissue expressed heparanase mRNA. These data suggest that by producing heparanase osteo(chondro)clasts contribute to the recruitment of the active form of VEGF. Thus osteo(chondro)clasts may promote local angiogenesis as well as callus resorption in endochondral ossification during fracture healing.
  • S Srivastava, M Matsuda, ZY Hou, JP Bailey, R Kitazawa, MP Herbst, ND Horseman JOURNAL OF BIOLOGICAL CHEMISTRY 278 (46) 46171 -46178 2003年11月 [無し][無し]
     
    Prolactin (PRL) is the primary hormone that, in conjunction with local factors, leads to lobuloalveolar development during pregnancy. Recently, receptor activator of NF-kappaB ligand (RANKL) has been identified as one of the effector molecules essential for lobuloalveolar development. The molecular mechanisms by which PRL may induce RANKL expression have not been carefully examined. Here we report that RANKL expression in the mammary gland is developmentally regulated and dependent on PRL and progesterone, whereas its receptor RANK (receptor activator of NF-kappaB) and decoy receptor osteoprotegerin (OPG) are constitutively expressed at all stages in both normal (PRL+/-) and prolactin knockout (PRL-/-) mice. In vitro, PRL markedly increased RANKL expression in primary mammary epithelial cells and RANKL-luciferase reporter activity in CHOD6 cells, which constitutively express the PRL receptor. We identified a gamma-interferon activation sequence (GAS) in the region between residues -965 to -725 of the RANKL promoter, which conferred a PRL response. Using dominant negative mutants of recombinant Jak2 and Stat5 in CHOD6 cells, and by reconstituting the Jak2/Stat5 pathway in COS7 cells, we determined that Jak2 and Stat5a are essential for the PRL-induced RANKL expression in mammary gland.
  • S Srivastava, M Matsuda, ZY Hou, JP Bailey, R Kitazawa, MP Herbst, ND Horseman JOURNAL OF BIOLOGICAL CHEMISTRY 278 (46) 46171 -46178 2003年11月 [無し][無し]
     
    Prolactin (PRL) is the primary hormone that, in conjunction with local factors, leads to lobuloalveolar development during pregnancy. Recently, receptor activator of NF-kappaB ligand (RANKL) has been identified as one of the effector molecules essential for lobuloalveolar development. The molecular mechanisms by which PRL may induce RANKL expression have not been carefully examined. Here we report that RANKL expression in the mammary gland is developmentally regulated and dependent on PRL and progesterone, whereas its receptor RANK (receptor activator of NF-kappaB) and decoy receptor osteoprotegerin (OPG) are constitutively expressed at all stages in both normal (PRL+/-) and prolactin knockout (PRL-/-) mice. In vitro, PRL markedly increased RANKL expression in primary mammary epithelial cells and RANKL-luciferase reporter activity in CHOD6 cells, which constitutively express the PRL receptor. We identified a gamma-interferon activation sequence (GAS) in the region between residues -965 to -725 of the RANKL promoter, which conferred a PRL response. Using dominant negative mutants of recombinant Jak2 and Stat5 in CHOD6 cells, and by reconstituting the Jak2/Stat5 pathway in COS7 cells, we determined that Jak2 and Stat5a are essential for the PRL-induced RANKL expression in mammary gland.
  • 膵癌及び大腸癌細胞株におけるTrkAの発現調節機構
    2003年 [無し][無し]
  • 脂肪肉腫への分化を示す肺癌肉腫の一剖検例
    2003年 [無し][無し]
  • 胚細胞及び胚細胞由来の腫瘍を認識する単クローン抗体5G9の作製及びその抗原解析
    2003年 [無し][無し]
  • 肺梗塞で死亡した潰瘍性大腸炎の1例
    2003年 [無し][無し]
  • 日本人におけるMatrix Gla Protein T-138C遺伝子多型の剖検症例よりの解析
    2003年 [無し][無し]
  • 成長軟骨と関節軟骨における破骨細胞分化因子(RANKL)の発現
    2003年 [無し][無し]
  • 悪性腫瘍の骨破壊病変における遺伝子発現
    2003年 [無し][無し]
  • 心筋緻密化障害の1例
    2003年 [無し][無し]
  • ステロイドホルモンによるマウス破骨細胞抑制因子(OPG)発現調節機構の解析
    2003年 [無し][無し]
  • 膵癌及び大腸癌細胞株におけるTrkA発現の調節機構
    2003年 [無し][無し]
  • 胚細胞及び胚細胞由来腫瘍を認識する単クローン抗体5G9の作製とその抗原解析
    2003年 [無し][無し]
  • 悪性腫瘍の溶骨性骨転移におけるRANKL発現制御機構
    2003年 [無し][無し]
  • ラット睾丸の造精子過程におけるサイクリンD1発現、プロモータ領域のメチル化及びMeCP2発現の関連
    2003年 [無し][無し]
  • マウス破骨細胞抑制因子(OPG)遺伝子発現調節機構の解析
    2003年 [無し][無し]
  • 成長軟骨と関節軟骨における破骨細胞分化因子(RANKL)の発現
    2003年 [無し][無し]
  • 1, 25 (OH)2 vitamin D3によるマウス破骨細胞抑制因子(OPG)発現調節機構の解析
    2003年 [無し][無し]
  • 脳室上衣腫合併MEN I型の一例と副甲状腺におけるメニンの検討
    2003年 [無し][無し]
  • 転写早期のmRNAをターゲットとした電顕レベルでのin situ hybridization
    2003年 [無し][無し]
  • Structure and Functional Elements of Human and Mouse RANKL Gene Promoter
    2003年 [無し][無し]
  • Regulation of Mouse Osteoprotegerin Gene Expression by Steroid Hormones
    2003年 [無し][無し]
  • Role of TGF-β and Menin in the Proliferation and Secretion of Human Parathyroid Cells.
    2003年 [無し][無し]
  • Regulation of Mouse Osteoprotegerin Gene Expression by 1α,25 Dihydroxyvitamin D3.
    2003年 [無し][無し]
  • In Vivo Imaging of TGF-β-induced RANK Ligand Transcriptional Activation in Prostate Cancer.
    2003年 [無し][無し]
  • Functional Elements for Transactivation by PTHrP Are Preserved in Both Human and Mouse RANKL Gene Promoters.
    2003年 [無し][無し]
  • Expression of Mouse Receptor of NF-kappaB Ligand(RANKL) in Growth Plate and Articular Cartilage.
    2003年 [無し][無し]
  • Epigenetic Regulation of Mouse RANKL Gene Expression.
    2003年 [無し][無し]
  • Antiestrogens and Mechanical Loading both Stimulate Transcriptional Activity of the BMP-6 Promoter in Osteoblasts in an ERα-Dependent Manner.
    2003年 [無し][無し]
  • Role of TGF-beta and menin in the proliferation and secretion of human parathyroid cells.
    2003年 [無し][無し]
  • Role of TGF-β and menin in the proliferation and secretion of human parathyroid cells.
    2003年 [無し][無し]
  • 糸球体内転移を伴った膵悪性腫瘍の3例
    診断病理 20巻, pp. 144-145 2003年 [無し][無し]
  • 感染性大動脈瘤破綻による心タンポナーデの1例
    診断病理 20巻, 3号, pp. 236-238 2003年 [無し][無し]
  • アミオダロン加療中に間質性肺炎,薬剤性肝炎及び膵炎を伴った陳旧性心筋梗塞の1例
    診断病理 20巻, 4号, pp. 328-330 2003年 [無し][無し]
  • Becker型筋ジストロフィーの1剖検例
    診断病理 20巻, 3号, pp. 283-285 2003年 [無し][無し]
  • 悪性腫瘍の骨破壊病変における遺伝子発現
    病理と臨床 22巻, 3号, pp. 285-289 2003年 [無し][無し]
  • 日本臨床外科学会雑誌 65巻, 2号, pp. 410-413 2003年 [無し][無し]
  • 脂肪肉腫への分化を示す肺癌肉腫の一剖検例
    日本病理学会会誌 92巻, 1号, pp. 279-279 2003年 [無し][無し]
  • 肺梗塞で死亡した潰瘍性大腸炎の1例
    日本病理学会会誌 92巻, 1号, pp. 393-393 2003年 [無し][無し]
  • 医学物理 23巻, 2号, pp. 157-159 2003年 [無し][無し]
  • Imaging of fine structure of bone sample with high coherent X-ray beam and high spatial resolution detector
    Radiation Medicine Vol. 22, No. 1, pp. 56-59 2003年 [無し][無し]
  • Journal Of Cellular Biochemistry Vol. 89, No. 4, pp. 771-777 2003年 [無し][無し]
  • Igaku Butsuri Vol. 23, No. 2, pp. 157-159 2003年 [無し][無し]
  • European Journal Of Endocrinology / European Federation Of Endocrine Societies Vol. 148, No. 4, pp. 403-411 2003年 [無し][無し]
  • Endocrine Journal Vol. 50, No. 2, pp. 215-219 2003年 [無し][無し]
  • Structure and Functional Elements of Human and Mouse RANKL Gene Promoter
    2003年 [無し][無し]
  • Regulation of Mouse Osteoprotegerin Gene Expression by Steroid Hormones
    2003年 [無し][無し]
  • Functional Elements for Transactivation by PTHrP Are Preserved in Both Human and Mouse RANKL Gene Promoters.
    2003年 [無し][無し]
  • Expression of Mouse Receptor of NF-kappaB Ligand(RANKL) in Growth Plate and Articular Cartilage.
    2003年 [無し][無し]
  • Epigenetic Regulation of Mouse RANKL Gene Expression.
    2003年 [無し][無し]
  • Role of TGF-beta and menin in the proliferation and secretion of human parathyroid cells.
    2003年 [無し][無し]
  • Imaging of fine structure of bone sample with high coherent X-ray beam and high spatial resolution detector
    Radiation Medicine Vol. 22, No. 1, pp. 56-59 2003年 [無し][無し]
  • Journal Of Cellular Biochemistry Vol. 89, No. 4, pp. 771-777 2003年 [無し][無し]
  • Igaku Butsuri Vol. 23, No. 2, pp. 157-159 2003年 [無し][無し]
  • European Journal Of Endocrinology / European Federation Of Endocrine Societies Vol. 148, No. 4, pp. 403-411 2003年 [無し][無し]
  • Endocrine Journal Vol. 50, No. 2, pp. 215-219 2003年 [無し][無し]
  • Parathyroid apoplexyと術後血中PTH値低下の遅延をきたした原発性副甲状腺機能亢進症の一例
    Clinical Calcium 13巻, 4号, pp. 496-499 2002年 [無し][無し]

共同研究・競争的資金等の研究課題

  • 非定型的DNAメチル化修飾を指標とする腫瘍初期病変の同定
    愛媛大学:基盤研究(B)
    研究期間 : 2016年 -2019年 
    代表者 : 北澤 荘平, 医学系研究科
  • 成長軟骨板に依存する長管骨発生プロセスの理解 : 組織系譜解析によるアプローチ
    愛媛大学:基盤研究(C)
    研究期間 : 2015年 -2018年 
    代表者 : 原口 竜摩, 医学系研究科
  • 破骨細胞分化因子受容体(RANK)発現制御機構の解析
    愛媛大学:基盤研究(C)
    研究期間 : 2015年 -2018年 
    代表者 : 北澤 理子, 医学部附属病院
  • 塩基配列特異的なメチル化シトシンin situ検出法の開発
    愛媛大学:挑戦的萌芽研究
    研究期間 : 2014年 -2017年 
    代表者 : 北澤 荘平, 医学系研究科
  • 破骨細胞前駆細胞における破骨細胞分化因子受容体(RANK)発現制御機構の解析
    愛媛大学:基盤研究(C)
    研究期間 : 2012年 -2015年 
    代表者 : 北澤 理子, 医学部附属病院
  • 胸膜中皮腫発生・進展に関わるエピジェネティクス変化の病理病態解析
    神戸大学:基盤研究(C)
    研究期間 : 2010年 -2015年 
    代表者 : 出射 由香, 医学(系, 研究科, 究
  • 常温遺伝子増幅によるメチル化シトシンin situ検出法の開発
    愛媛大学:挑戦的萌芽研究
    研究期間 : 2011年 -2013年 
    代表者 : 北澤 荘平, 医学(系, 研究科
     
    染色体上で標的DNAと塩基特異的なメチル化シトシンに結合するICONプローブとを結合させ、強固な化学結合により固定化する。その後、このICONプローブをもとに染色体上でPCR反応を進展させ(いわゆるPRINS法)、標式核酸をこのDNA合性、進展過程でとりこませ、組織化学的に検出することにより、染色体や細胞、組織構築を保ったままin situで特異的な塩基配列のメチル化シトシンの有無を検出する方法の基礎的データを収集できた。マウス精巣や培養細胞を用いた系で条件設定を行い、単一遺伝子での塩基配列特異的なメチル化シトシンの検出ができるようになった。現在、安定した増幅方法について追加検討中である。
  • 転写因子CREBの遺伝子変異による新規胎児致死症候群の解析
    神戸大学:挑戦的萌芽研究
    研究期間 : 2010年 -2012年 
    代表者 : 藤本 昌代, 医学(系, 研究科, 究
     
    本研究課題は、子宮内発育遅延による死産・新生児死亡の病理解剖症例を対象に、シグナル伝達に関連する転写調節因子や形態形成に関連した遺伝子の異常を解析し、病態との関わりを検討し、新規の胎児致死症候群を同定することをめざす。 これまでに、子宮内胎児死亡剖検症例のパラフィン包埋病理組織からCREBのexon 5の遺伝子変異を検出している。116番目アミノ酸がアスパラギンからグリシンに変異(CREE116)するものであり、この変異が機能的ドメインのどの位置に存在するか、アミノ酸の3次元解析から想定される部位について、詳細を検討中である。 症例と同一のCREB変異体の発現ベクターを構築して培養細胞に導入し、CRE-luciferaseを用いたレポーターアッセイを行った結果、CREB116の転写促進活性は野生型の1/6と著しい減弱を示した。培養細胞に、野生型とCREB116、既報のCERB変異体を導入して免疫沈降Western blotting法にて比較検討すると、CREB116では、CREB蛋白のリン酸化は野生型と同等であったが、その後の転写共役因子CBP/P300との結合が見られず、dominant negativeとして作用し、野生型と競合することで、転写効率を下げる可能性が示唆された。アミノ酸変異が共役因子との結合に及ぼす影響について、詳細を検討中である。 発端となった症例では、諸臓器の異常の中でも、肺の拡張障害が高度で治療抵抗性であった。CREB116が、特定臓器の形態異常や機能不全を引き起こす機序は明らかでない。本年度は、肺の病理組織を用いて、肺胞上皮の蛋白発現を検索した。新生児/胎児死亡の剖検例で、CREB野生型の症例を対象に比較検討すると、一連のサーファクタント関連蛋白のうち、著しい低下を示すものがあり、呼吸不全に寄与した可能性が示唆された。 以上の知見に関しては、今後内外の学会にて報告するとともに、論文投稿準備中である。
  • 骨髄単球細胞の分化過程における破骨細胞分化因子受容体(RANK)の発現制御
    神戸大学:基盤研究(C)
    研究期間 : 2009年 -2011年 
    代表者 : 北澤 理子, 医学研究科, 特命准教授
     
    本研究計画は、骨髄単核細胞から破骨細胞への分化のメカニズムを解明する目的で、破骨細胞分化因子の受容体RANKの発現制御について解析を行った。破骨細胞分化には、RANKL-RANK結合が必須であるが、adaptor分子TRAFのリン酸化を経て転写因子PU. 1, MITF, NFATc1, AP-1はカテプシンKやTRACPなどの破骨細胞形質遺伝子の発現を誘導する。本研究計画ではPU. 1, MITFに加えて、NFATc1やc-Fosが受容体RANK自体の発現に必要であることを明らかにした。さらに、RANKLはNFATc1を介して受容体RANKを正に制御するのみならず、前破骨細胞RAWへのRANKL投与により、RANKのsplicing variant(vRANK)が生じることを示した。vRANKは破骨細胞の分化や生存(抗アポトーシスシグナル)に対して拮抗的な作用を有することから、破骨細胞を抑制する治療戦略に繋がる可能性が示唆された。
  • 非CpGアイランド領域シトシンメチル化の包括的病態解析
    神戸大学:基盤研究(B)
    研究期間 : 2007年 -2010年 
    代表者 : 北澤 荘平, 医学研究科, 客員教授
     
    種々の遺伝子の転写調節領域に存在するCpG islandのシトシンメチル化は遺伝子発現を抑制的に制御することが知られている。一方で、非CpG island領域のシトシンメチル化についても、転写制御に重要な影響を示すものがあり、私どもは、膀胱癌におけるTGF-βシグナル経路の偽受容体BAMBIと、糖尿病ラットの腎腫瘍発生とP16の発現制御を対象として、メチル化の進展が腫瘍の形態変化に関与することを示した。
  • 組織標本上での遺伝子特定部位のメチル化シトシンin situ検出法の開発
    神戸大学:挑戦的萌芽研究
    研究期間 : 2007年 -2009年 
    代表者 : 北澤 荘平, 医学研究科, 特命教授
     
    単一の受精卵に由来しながら、神経細胞から皮膚に至る多種多様な組織細胞分化が起こるメカニズムには。エピジェネティクス制御が重要とされている。メチル化シトシンは、エピジェネティクス制御機構の主体をなすDNA修飾であり、組織細胞分化や再生医療のみならず、腫瘍発生・進展においても重要である。特に、癌の個性に応じた薬物療法として、近年ではDNAメチル化阻害剤の適応について検索する手法が重要となる。私どもは、対象遺伝子の調節領域に存在するCpG-islandのシトシンメチル化を、細胞や組織の形態を維持したまま、in situで検出する組織化学的方法の開発についての検討を行った。 本年度は、部位特異的DNAメチル化に対して、メチル化シトシンとオスミウム酸により錯体形成するICONプローブ(bipyridine-adenine標識プローブ)に着目し、染色体上で標的DNAとICONプローブとを結合させて、強固な化学結合で固定し、このICONプローブをもとに染色体上でPRINS法を行う実験系を構築することを目指して予検討を行った。染色体上でICONプローブをもとにPCR反応を行う際に、標識化合物を取り込ませた後、標識化合物に対する免疫組織化学にてシグナルを検出する。特異的で高感度のプローブ結合やPCR反応、メチル化シトシン-標識グアニン複合体に対する単クローン抗体の作成が課題となる。本年度は、単クローン抗体作成法の予検討と、染色体標本上でのPRINS法施行についての検討を行った。特に、標本の固定やheat denatureの条件設定や、標本上でのDNA合成進展に最適な酵素試薬の検定などを施行した。効率的なマウス免疫方法や、脾臓リンパ球回収、ミエローマ細胞との細胞融合、HAT培地による選択培養については、最適な実験系を整備することが出来た。
  • 腫瘍発生・進展における非定型的メチル化シトシンの病態学的意義についての網羅的解析
    神戸大学:基盤研究(B)
    研究期間 : 2006年 -2009年 
    代表者 : 前田 盛, 医学研究科, 戦略的客員
     
    種々の遺伝子の転写調節領域に存在するCpG-islandのメチル化は、遺伝子発現を抑制的に非CpG-islandの意義は不明であった。私どもは、病理組織学的観察ににおける遺伝子発現制御の解析を、遺伝子プロモータ領域の非定型的なメチル化シトシンの解析を中心に行い、非CpG-islandのシトシンメチル化による転写制御が、悪性腫瘍の進展に関わっていることを明らかにした。
  • 骨髄微小環境での破骨細胞分化因子受容体(RANK)発現調節と破骨細胞分化の解析
    神戸大学:基盤研究(C)
    研究期間 : 2006年 -2008年 
    代表者 : 北澤 理子, 医学研究科
     
    マウス破骨細胞分化因子受容体RANKの遺伝子プロモータ領域をクローニングして発現制御機構を検討した。RANK遺伝子の転写開始部位より1kb以内には、血球分化に重要なPU.1(-480)、MITF(-100)結合配列が存在した。ゲルシフトアッセイにて、結合配列の蛋白DNA結合を証明し、変異を導入したプロモータを用いた解析にて、PU.1、MITFは、これらの結合配列に作用して、RANK転写を促進することを明らかにした。
  • 溶骨性骨転移における破骨細胞分化因子(RANKL)の役割
    神戸大学:基盤研究(C)
    研究期間 : 2004年 -2005年 
    代表者 : 北澤 理子, 医学系研究科
     
    大多数の癌の骨転移は溶骨性であるが、骨の脆弱性により病的骨折を来たし患者のQOLを損なうため、臨床的に重要である。骨組織の微小環境に腫瘍細胞が介入することにより、破骨細胞形成・骨破壊が促進するが、副甲状腺ホルモン関連蛋白(PTHrP)などの腫瘍由来因子が破骨細胞分化因子(RANKL)の発現を亢進させるメカニズムについて検討した。 形態学的には、ヒトの癌の溶骨性骨転移巣の病理組織標本や、ヌードマウス頭蓋にPTHrP産生癌細胞を移植した溶骨性骨病変の実験モデルの組織標本を用いて、RANKL mRNA発現を解析した。骨転移巣では、腫瘍細胞に近接する部位の骨芽細胞・骨髄間質細胞にRANL発現を認め、引き続いて破骨細胞形成が観察された。 従来より解析してきたマウスRANKL遺伝子プロモータ領域に存在するRunx2結合配列の機能について検討した。Runx2欠損マウスには破骨細胞が形成されないことが知られてきたが、Runx2欠損マウス由来の骨芽細胞ではRunx2を持つST2細胞よりも、定常状態でのRANKL発現が高いこと、Runx2欠損細胞へのRunx2強制発現でRANKL発現は減少し、ST2細胞へのsiRNAを用いたRunx2ノックダウンによりRANKL発現が増加することから、Runx2がRANKL発現を抑制することを示した。RANKL遺伝子プロモータのさらに上流を解析すると、Runx2がクロマチン凝集を促進した。PTH/PTHrPのシグナル伝達系としては、PKAがクロマチン凝集を解除することにより、RANKL転写を促進することを示した。 RANKL発現がプロモータ領域のCpGメチル化により制御される機構に関しても解析した。プロモータDNAをin vitroでメチル化すると転写活性は1/3に減少し、高メチル化状態の培養細胞を脱メチル化剤で処理すると、RANKL発現と破骨細胞形成支持能が回復した。マウス諸臓器のgemomic DNAの解析から、TATA-box直前のCpG locusのメチル化がRANKL発現低下と相関し、ChIP assayやEMSAにより、同部位のメチル化がMeCP2結合に与りTATA-boxでのDNA蛋白結合を阻害することにより、RANKL転写を抑制する機構が存在することを示した。 以上の成果は、第22回、23回日本骨代謝学会、第93回、94回日本病理学会にて報告し、これらの成果はActa Histochem Cytochem、Histochem Cell Biol、J Cell Biochemに報告し、J Bone Miner Resに投稿中である。
  • 破骨細胞分化因子(RANKL)遺伝子発現調節機構の解析
    神戸大学:基盤研究(C)
    研究期間 : 2002年 -2003年 
    代表者 : 北澤 理子, 医学系研究科
     
    骨芽細胞は、1)骨髄造血系細胞の増殖に必要なM-CSFなど液性因子の供給、2)破骨細胞への最終分化に不可欠な破骨細胞分化因子(RANKL)の発現を介して破骨細胞形成を支持する。マウスならびにヒトRANKL遺伝子5'上流プロモータ領域を解析して、RANKL発現を介するvitamin D3の破骨細胞形成促進作用を明らかにした。マウスプロモータでは1箇所のVDRE(-940)が機能的に有効であるが、ヒトプロモータについては3箇所の候補配列(-1584)、(-1418)、(-1383)のうち、最上流の(-1584)が機能的VDREであることを明らかにした。マウスとヒトのプロモータ構造は類似性が高く、VDREを介する制御機構は種を超えてよく保存されていることが示された。 悪性腫瘍の溶骨性転移の機構をRANKL発現制御の観点から検討した、マウスおよびヒトRANKLに対する特異的なプローブを作成してin situ hybridization法を行い、病理解剖症例のPTHrP産生腫瘍の転移巣の組織標本にて骨芽細胞・骨髄間質細胞にRANKL mRNAを検出した。 PTHrPによるRANKL発現制御の検討も継続して行った。マウス及びヒトのRANKL遺伝子プロモータコンストラクトを用いたtransfection系に、PTHrP処理を行うとともにPKAのagonist, antagonist投与を行い転写活性に及ぼす効果を検討した。マウス及びヒトRANKL遺伝子プロモータにおけるcAMP応答配列(CRE)の存在に関しては、マウス、ヒト各々数力所の候補配列に着目してEMSA assayや変異導入実験を行い、機能的CREの存在を明らかにした。プロモータ活性の検討に加えて、H3,H4ヒストンに対する抗体を用いてのCHIP assayを行い、PTHrPもしくはPKA agonistの作用でクロマチン活性化が起こることを確認した。 以上の成果は、英文誌に報告した(BBRC2002,J Pathol 2002,J Cell Biochem 2003)。また、成果の一部は第91回92回日本病理学会総会、第20回21回日本骨代謝学会、第61回62回日本癌学会総会、第24回25回米国骨代謝学会議にて報告した。
  • 自己免疫性関節炎における骨破壊の分子機構:破骨細胞分化因子(RANKL)の関与
    神戸大学:萌芽研究
    研究期間 : 2001年 -2002年 
    代表者 : 前田 盛, 医学系研究科
     
    慢性関節リウマチ(RA)等の自己免疫性関節炎においては、滑膜増生・パンヌス形成に引き続いて軟骨・骨組織の破壊が起こる。さらに関節炎の進行に伴い、関節近傍の骨量減少や全身性の骨粗鬆症が生じる。消炎鎮痛剤や免疫抑制剤投与により炎症を沈静化しえても、骨破壊による関節の機能障害が残るため、患者のQOL低下を招く。 炎症早期には、サイトカインの作用が重要であるが、進行期の骨破壊病変には破骨細胞が中心的な役割を果たすと考えられている。破骨細胞の最終分化・活性化には骨芽細胞/間質細胞との相互作用が必須とされてきたが、破骨細胞分化因子(RANKL)が破骨細胞分化誘導に不可欠な因子として分離同定され、骨吸収性サイトカインによりその発現が増強することが想定されている。さらに、RANKL受容体であるRANK発現状態、破骨細胞抑制因子Ostcoprotegerinが破骨細胞数や活動性を制御すると考えられている。 自己免疫関節炎における骨破壊の分子機構を解明する目的で、マウスのII型コラーゲン誘導関節炎(Collagen Induced Arthritis、CIA)におけるRANKL、RANK、OPG各々の遺伝子発現をin situ hybridization (ISH)法にて解析した。関節炎局所では、炎症性滑膜やパンヌス内に単核〜多核の破骨細胞(TRAP陽性・RANK陽性)が存在し、炎症性滑膜組織が破骨細胞の増殖と蓄積を支持している事が示唆された。同部位にRANKL陽性滑膜線維芽細胞の増加を認めたことから、滑膜細胞が破骨細胞形成に直接関与する可能性、即ち滑膜線維芽細胞とマクロファージ/単核細胞とのRANKL-RANKを介する相互作用により破骨細胞前駆細胞が形成される機構を明らかにした。以上の成果は内外の学会にて報告するとともに、英文誌Journal of Histochemistry and Cell Biologyに報告した。
  • マウス破骨細胞分化因子(RANKL)遺伝子プロモータ領域の解析
    神戸大学:基盤研究(C)
    研究期間 : 2000年 -2001年 
    代表者 : 北澤 理子, 医学系研究科
     
    骨芽細胞は、1)骨髄造血系細胞の増殖に必要なM-CSFの供給、2)破骨細胞分化に不可欠な破骨細胞分化因子(RANKL)の発現を介して破骨細胞形成を支持する。私どもはマウスRANKL遺伝子5'上流プロモータ領域を解析した。putative VDRE(-937/-922)は典型的なDR3構造ではないが、vitD存在下でEMSA法にてVDR-RXRと特異的に結合した。同部位にmutationを導入したconstructを培養細胞に導入して転写活性を評価し、-937/-922がvitD作用に不可欠であることを示した。マウスRANKL遺伝子プロモータ領域のputativeなグルココルチコイド応答配列(-642/-628)についても、mutationを導入して転写活性を検討した。さらにプロモータ領域に存在するCpG islandのメチル化がRANKL遺伝子発現を制御することを示した。マウス培養ストローマ細胞ST2では継代数に依存して破骨細胞形成支持能とRANKL発現が低下する。RANKL遺伝子高発現のPassage9(P9)に比して低発現のP16では、外因性に導入したpromoterの転写活性には差がないが、内因性のpromoterの遺伝子転写開始部位周辺のCpGにメチル化修飾を受けていることを示した。 悪性腫瘍骨転移巣におけるRANKL発現を検索するために、マウスおよびヒトRANKL遺伝子に特異的なプローブを作成してin situ hybridization法を行いRANKL mRNAを検出した。 以上の成果は、英文誌に投稿した(BBRC, Histochem Cell Biol, J Pathol)。また、成果の一部は第89回、第90回日本病理学会総会、第18回、第19回日本骨代謝学会、第22回、第23回米国骨代謝学会議、第23回IAP総会にて発表した。
  • 前立腺癌造骨性骨転移に関わる形態形成遺伝子群の解析
    神戸大学:萌芽的研究
    研究期間 : 1999年 -2000年 
    代表者 : 前田 盛, 神戸, 学, 医学部
     
    前立腺癌におけるBMP-6の遺伝子発現調節機構について検討した。まず前立腺癌培養細胞株DU-145,LNCap,PC-3,PC-3Mにおいて、Norhtern blotによりBMP-6mRNA発現の差を見い出した。これらの培養細胞にBMP-6遺伝子プロモータを導入しても転写活性は同レベルであり、プロモータ領域のCpGメチル化によってBMP-6遺伝子発現が制御されることを明らかにした。さらに11例の前立腺癌症例についてin situ hybridizationを行い、原発巣と転移巣とにおけるBMP-6,BMP-2mRNA発現を解析した。原発巣において分化度の高い症例ではBMP-2mRNAの発現レベルが高いのに対して、BMP-6mRNAは低分化型腺癌の症例に高率に発現し、他臓器の転移浸潤部位ではBMP-6の発現レベルが高くなる傾向を認めた。BMP-6mRNA発現レベルと対応させて、原発巣および転移巣の標本からmicro dissectionにて腫瘍組織を切り出してBMP-6遺伝子5'側上流領域のCpGメチル化を解析した。原発巣の腫瘍において検出されるBMP-6遺伝子プロモータ領域のSp1結合配列周辺のCpGメチル化が、浸潤転移病巣においては脱メチル化を示していることを明らかにした。このように、腫瘍の増殖・進展の過程で生じるCpGメチル化の変化により付加的な遺伝子発現あるいは遺伝子発現の抑制を来たし、転移浸潤に関わる腫瘍の生物学的特性をもたらす可能性が示唆された。 以上の成果は、英文誌Journal of Bone and Mineral Researchに報告し印刷中である。また、成果の一部は第89回日本病理学会総会ワークショップ、第23回IAP総会シンポジウムにて発表した。
  • ヒトα6インテグリン遺伝子プロモータ領域の解析
    神戸大学:基盤研究(C)
    研究期間 : 1998年 -1999年 
    代表者 : 北澤 理子, 医学部
     
    腫瘍細胞の浸潤初期過程には、基底膜の構成成分ラミニンに対する受容体の発現が必須である。腫瘍転移浸潤機序を解明するために、ラミニン受容体を構成するα6インテグリンの遺伝子5'側上流領域を解析した。α6遺伝子の基本転写調節領域の上流にはAP-1、c-Mycのpositive regulatory elementがあり、その直下にはプロゲステロンレセプター結合配列の存在が想定された。今回α6遺伝子5'側上流、3個のSp1 siteを含むGC rich領域におけるメチル化について検索した。種々の乳癌培養細胞株ならびに前立腺癌培養株から抽出したDNAを、メチル化耐性、不耐性の制限酵素であるMsp IとHap IIにて処理後にα6遺伝子5'側上流領域をプローブとするSouthern hybridizationを行った。前立腺癌培養株では殆どメチル化がなく、乳癌培養細胞株については高率にメチル化を示唆するバンドを検出したので、特異的プライマーを設定してPCRで該当部位を増幅しシークエンス解析を行ってメチル化を確認した。α6インテグリン発現調節にプロモータ領域のSp1 siteを含むGC rich領域のメチル化修飾が関与する可能性を示した。 さらに、病理組織標本における遺伝子プロモータ領域のメチル化を評価するために、パラフィン包埋組織よりのDNA抽出とメチル化検出法に関する検討を行い多くの知見を集積した。加えて、細胞接着因子・細胞外マトリックスをはじめとする遺伝子の発現を形態学的に評価するために、組織分子雑種法の基礎検討を行ってきた。特に組織上でのmRNAの保存に適した固定法やPCRを利用した高感度・特異的なプローブ作製方法について多くの知見を集積した。以上の成果の一部は第87回・88回日本病理学会総会、第16回・17回日本骨代謝学会、第20回・21回米国骨代謝学会議にて報告した。
  • 骨髄ストローマ細胞の機能分化と破骨細胞形成支持に関わる因子の解析
    神戸大学:基盤研究(B)
    研究期間 : 1997年 -1998年 
    代表者 : 北澤 理子, 医学部
     
    破骨細胞形成において骨芽細胞系の果たす役割は、1)骨髄造血系細胞〜破骨細胞系の増殖に必要な液性因子の供給、2)破骨細胞への最終分化に不可欠な直接接触を介する相互作用、の2段階が想定される。本計画において、2)の観点より、マウス骨髄ストローマ細胞ST2と骨髄細胞の共存培養において、ST2細胞の破骨細胞形成支持能の推移(ST2細胞の継代数依存的)に着目し、Differential DispIay法にて破骨細胞形成能と連動して発現の推移する分子の分離を目指した。 一方、1998年破骨細胞分化因子が分離同定されたが(RANKL/TRANCE/OPGL/ODF:H.Yasuda et al.,PNAS,D.Lacy et al.,Cell,1998)、私どもは破骨細胞分化因子(RANKL.)遺伝子5'上流プロモータ領域をクローニングし、RANKL遺伝子発現調節機序について検討した。特にST2細胞の継代数依存的な破骨細胞形成支持能がRANKL遺伝子発現レベルによって既定されること示し、遺伝子プロモータ領域のメチル化修飾が関与する可能性を見い出した。成果の一部は既に投稿し論文印刷中である。今後RANKL.遺伝子発現調節機序の詳細について研究を進めていく。さらに、サイトカインや上記遺伝子の発現を形態学的に評価するために、硬組織を用いた組織分子雑種法の基礎的検討を行ってきた。特に組織上でのmRNAの保存に適した固定法・脱灰法の条件設定やPCRを利用した高感度・特異的なプローブ作製方法について多くの知見を集積し、脱灰骨標本を用いた組織分子雑種法にて骨形成因子(BMP)や血小板由来増殖因子(PDGF)の局在を示すことが可能となった。以上の成果の一部は第86回・87回日本病理学会総会、第15回・16回日本骨代謝学会、第19回・20回米国骨代謝学会議にて報告した。
  • 癌の骨転移とRANKL-RANKシグナルによる破骨細胞形成機構
  • Osteoclastogenesis via RANKL-RANK signaling in bone metastasis

愛媛大学教員活動実績

教育活動(B)

担当授業科目(B01)

  • 2012, 通年, 学部, 病理学
  • 2012, 前期, 学部, こころと健康
  • 2013, 通年, 学部, 病理学各論
  • 2013, 前期, 学部, 病理学総論
  • 2013, 前期, 学部, こころと健康
  • 2014, 前期, 学部, こころと健康
  • 2014, 通年, 学部, 病理学各論
  • 2014, 前期, 学部, 病理学総論
  • 2014, 前期, 学部, こころと健康
  • 2015, 通年, 学部, 病理学総論各論
  • 2015, 前期, 学部, こころと健康
  • 2015, 前期, 学部, こころと健康
  • 2015, 前期, 学部, こころと健康


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