Akira Yamaguchi

Publications (66)251.9 Total impact

• Article: CCN3 participates in bone regeneration as an inhibitory factor.

  Yuki Matsushita, Kei Sakamoto, Yoshihiro Tamamura, Yasuaki Shibata, Tokutaro Minamizato, Tasuku Kihara, Masako Ito, Ken-Ichi Katsube, Shuichi Hiraoka, Haruhiko Koseki, Kiyoshi Harada, Akira Yamaguchi
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  ABSTRACT: CCN3, a member of the CCN protein family, inhibits osteoblast differentiation in vitro. However, the role of CCN3 in bone regeneration has not been well elucidated. In this study, we investigated the role of CCN3 in bone regeneration. We identified Ccn3 gene by microarray analysis as a highly expressed gene at the early phase of bone regeneration in a mouse bone regeneration model. We confirmed the upregulation of Ccn3 at the early phase of bone regeneration by RT-PCR, western blot, and immunofluorescence analyses. Ccn3 transgenic mice, in which Ccn3 expression was driven by 2.3-kb Col1a1 promoter, showed osteopenia compared with wild-type mice, but Ccn3 knockout mice showed no skeletal changes compared with wild-type mice. We analyzed bone regeneration process in Ccn3 transgenic mice and Ccn3 knockout mice by microcomputed tomography and histological analyses. Bone regeneration in Ccn3 knockout mice was accelerated compared with that in wild-type mice. The mRNA expression levels of osteoblast-related genes (Runx2, Sp7, Col1a1, Alpl, Bglap) in Ccn3 knockout mice were upregulated earlier than those in wild-type mice, as demonstrated by RT-PCR. Bone regeneration in Ccn3 transgenic mice showed no significant changes compared with that in wild-type mice. Phosphorylation of Smad1/5 was highly upregulated at bone regeneration sites in Ccn3 KO mice compared with wild-type mice. These results indicate that CCN3 is upregulated in the early phase of bone regeneration, and acts as a negative regulator for bone regeneration. This study may contribute to the development of new strategies for bone regeneration therapy.
  Journal of Biological Chemistry 05/2013; · 4.77 Impact Factor
• Article: The expression profiles of acidic epithelial keratins in ameloblastoma.

  Samir Kumar Pal, Kei Sakamoto, Tadanobu Aragaki, Takumi Akashi, Akira Yamaguchi
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  ABSTRACT: To characterize the subtypes of ameloblastoma by differentiation markers. Expression of 9 major acidic epithelial keratins was immunohistochemically examined in 28 ameloblastomas. Keratin 15 (K15) expression patterns corresponded to histological variants: follicular, plexiform and acanthomatous. Tumor nests comprising K15-expressing basal cells mimicked oral epithelium or dental lamina, and tumor nests comprising K15-negative basal cells mimicked outer enamel epithelium. Keratin 19 (K19) was consistently expressed in solid/multicystic ameloblastoma and unicystic ameloblastoma, while peripheral ameloblastoma and desmoplastic ameloblastoma contained K19-negative cells. The 4 current subtypes had unvaried expression patterns within each group. However, they could be divided into 2 groups by K19 expression pattern: solid/multicystic and unicystic versus extraosseous/peripheral and desmoplastic. K15 expression pattern represented various types of differentiation for tumor nests mimicking tooth germ and oral epithelium. The results clarify the homogeneity and heterogeneity of ameloblastoma cell lineage and differentiation.
  Oral surgery, oral medicine, oral pathology and oral radiology. 04/2013; 115(4):523-31.
• Article: RANKL Synthesized by Both Stromal Cells and Cancer Cells Plays a Crucial Role in Osteoclastic Bone Resorption Induced by Oral Cancer.

  Kiyoshi Sato, Ji-Won Lee, Kei Sakamoto, Tadahiro Iimura, Kou Kayamori, Hisataka Yasuda, Masanobu Shindoh, Masako Ito, Ken Omura, Akira Yamaguchi
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  ABSTRACT: The molecular mechanisms underlying bone destruction by invading oral cancer are not well understood. Using IHC, we demonstrated that receptor activator of nuclear factor-κB ligand (RANKL)-positive fibroblasts and cancer cells were located at sites of bone invasion in human oral cancers. HSC3 and HO-1-N-1, human oral cancer cell lines, expressed RANKL and stimulated Rankl expression in the UAMS-32 murine osteoblastic cell line. We discriminated the roles of RANKL synthesized by stromal cells and cancer cells in cancer-associated bone resorption by using species-specific RANKL antibodies against murine RANKL and human RANKL, respectively. Osteoclastogenesis induced by the conditioned medium of HSC3 and HO-1-N-1 cells in a co-culture of murine bone marrow cells and UAMS-32 cells was inhibited by the addition of antibodies against either mouse or human RANKL. HSC3-induced bone destruction was greatly inhibited by the administration of anti-mouse RANKL antibody in a xenograft model. HO-1-N-1-induced bone destruction was inhibited by the administration of either anti-mouse or anti-human RANKL antibody. Bone destruction induced by the transplantation of human RANKL-overexpressing cells (HSC3-R2) was greatly inhibited by the injection of anti-human RANKL antibody. The present study revealed that RANKL produced by both stromal and cancer cells is involved in oral cancer-induced osteoclastic bone resorption. These results provide important information for understanding the cellular and molecular basis of cancer-associated bone destruction and the mechanism of action underlying RANKL antibody (denosumab) therapy.
  American Journal Of Pathology 03/2013; · 4.89 Impact Factor
• Article: Roles of chemokine receptor CX3CR1 in maintaining murine bone homeostasis through the regulation of both osteoblasts and osteoclasts.

  Akiyoshi Hoshino, Satoshi Ueha, Sanshiro Hanada, Toshio Imai, Masako Ito, Kenji Yamamoto, Kouji Matsushima, Akira Yamaguchi, Tadahiro Iimura
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  ABSTRACT: Chemokines have recently been reported to be involved in pathological bone destruction. However, the physiological roles of chemokines in bone metabolism in vivo have not been well documented. We analyzed the bone phenotypes in Cx3cr1-deficient mice. The mice exhibited slight but significant increases in trabecular and cortical thickness, reduced numbers of osteoclasts and increased rates of osteoid formation. Although the morphometric parameters showed marginal differences, the Cx3cr1-deficient bones showed an elevated expression of Osterix/SP7, an essential transcriptional factor for osteoblasts, while a late marker, Osteocalcin/Bglap, was downregulated. The levels of various osteoclastic markers, such as receptor activator of NF-κB (Rank)/Tnfrsf11a, receptor activator of NF-κB ligand (RANKL)/Tnfsf11, tartrate-resistant acid phosphatase 5b (Trap5b)/Acp5b, Cathepsin K/Ctsk, Mmp3 and Mmp13, were significantly decreased in the Cx3cr1-deficient bones. Cultured Cx3cr1-deficient osteoblastic cells showed inverse temporal patterns of osteoblastic marker expression and reduced calcium deposition. Further in vitro studies and immunofluorescence staining against CX3CR1 and CX3CL1 suggested a role for the CX3CR1-CX3CL1axis in an early stage of osteoblast differentiation, possibly through their trans- and cis-interactions. Cultured Cx3cr1-deficient pre-osteoclasts showed impaired differentiation, mainly due to a deficiency of the CD115(+)CD11b(lo) osteoclastogenic population of myeloid-lineage precursors. The treatment of bone marrow-derived osteoclastic cultures with recombinant CX3CL1 at different time points suggested that the CX3CR1-CX3CL1 axis favors the maintenance of osteoclastic precursors, but not differentiated osteoclasts. The current observations uncovered novel roles of the CX3CR1-CX3CL1 axis in the differentiation of both osteoblasts and osteoclasts.
  Journal of Cell Science 12/2012; · 6.11 Impact Factor
• Article: Spatiotemporal disorder in the axial skeleton development of the Mesp2-null mouse: A model of spondylocostal dysostosis and spondylothoracic dysostosis.

  Yuji Makino, Yu Takahashi, Rieko Tanabe, Yoshihiro Tamamura, Takashi Watanabe, Mayu Haraikawa, Miwako Hamagaki, Kenji Hata, Jun Kanno, Toshiyuki Yoneda, Yumiko Saga, Masae Goseki-Sone, Kazuo Kaneko, Akira Yamaguchi, Tadahiro Iimura
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  ABSTRACT: Spondylocostal dysostosis (SCDO) is a genetic disorder characterized by severe malformation of the axial skeleton. Mesp2 encodes a basic helix-loop-helix type transcription factor that is required for somite formation. Its human homologue, MESP2, is a gene affected in patients with SCDO and a related vertebral disorder, spondylothoracic dysostosis (STDO). This work investigated how the loss of Mesp2 affects axial skeleton development and causes the clinical features of SCDO and STDO. We first confirmed, by three-dimensional computed tomography scanning, that Mesp2-null mice exhibited mineralized tissue patterning resembling the radiological features of SCDO and STDO. Histological observations and in situ hybridization probing for extracellular matrix molecules demonstrated that the developing vertebral bodies in Mesp2-null mice were extensively fused with rare insertions of intervertebral tissue. Unexpectedly, the intervertebral tissues were mostly fused longitudinally in the vertebral column, instead of exhibiting extended formation, as was expected based on the caudalized properties of Mesp2-null somite derivatives. Furthermore, the differentiation of vertebral body chondrocytes in Mesp2-null mice was spatially disordered and largely delayed, with an increased cell proliferation rate. The quantitative three-dimensional immunofluorescence image analyses of phospho-Smad2 and -Smad1/5/8 revealed that these chondrogenic phenotypes were associated with spatially disordered inputs of TGF-β and BMP signaling in the Mesp2-null chondrocytes, and also demonstrated an amorphous arrangement of cells with distinct properties. Furthermore, a significant delay in ossification in Mesp2-null vertebrae was observed by peripheral quantitative computed tomography. The current observations of the spatiotemporal disorder of vertebral organogenesis in the Mesp2-null mice provides further insight into the pathogenesis of SCDO and STDO, and the physiological development of the axial skeleton.
  Bone 12/2012; · 4.02 Impact Factor
• Article: Osterix Regulates Calcification and Degradation of Chondrogenic Matrices through Matrix Metalloproteinase 13 (MMP13) Expression in Association with Transcription Factor Runx2 during Endochondral Ossification.

  Riko Nishimura, Makoto Wakabayashi, Kenji Hata, Takuma Matsubara, Shiho Honma, Satoshi Wakisaka, Hiroshi Kiyonari, Go Shioi, Akira Yamaguchi, Noriyuki Tsumaki, Haruhiko Akiyama, Toshiyuki Yoneda
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  ABSTRACT: Endochondral ossification is temporally and spatially regulated by several critical transcription factors, including Sox9, Runx2, and Runx3. Although the molecular mechanisms that control the late stages of endochondral ossification (e.g. calcification) are physiologically and pathologically important, these precise regulatory mechanisms remain unclear. Here, we demonstrate that Osterix is an essential transcription factor for endochondral ossification that functions downstream of Runx2. The global and conditional Osterix-deficient mice studied here exhibited a defect of cartilage-matrix ossification and matrix vesicle formation. Importantly, Osterix deficiencies caused the arrest of endochondral ossification at the hypertrophic stage. Microarray analysis revealed that matrix metallopeptidase 13 (MMP13) is an important target of Osterix. We also showed that there exists a physical interaction between Osterix and Runx2 and that these proteins function cooperatively to induce MMP13 during chondrocyte differentiation. Most interestingly, the introduction of MMP13 stimulated the calcification of matrices in Osterix-deficient mouse limb bud cells. Our results demonstrated that Osterix was essential to endochondral ossification and revealed that the physical and functional interaction between Osterix and Runx2 were necessary for the induction of MMP13 during endochondral ossification.
  Journal of Biological Chemistry 08/2012; 287(40):33179-90. · 4.77 Impact Factor
• Article: CXCL2 synthesized by oral squamous cell carcinoma is involved in cancer-associated bone destruction.

  Erika Oue, Ji-Won Lee, Kei Sakamoto, Tadahiro Iimura, Kazuhiro Aoki, Kou Kayamori, Yasuyuki Michi, Masashi Yamashiro, Kiyoshi Harada, Teruo Amagasa, Akira Yamaguchi
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  ABSTRACT: To explore the mechanism of bone destruction associated with oral cancer, we identified factors that stimulate osteoclastic bone resorption in oral squamous cell carcinoma. Two clonal cell lines, HSC3-C13 and HSC3-C17, were isolated from the maternal oral cancer cell line, HSC3. The conditioned medium from HSC3-C13 cells showed the highest induction of Rankl expression in the mouse stromal cell lines ST2 and UAMS-32 as compared to that in maternal HSC3 cells and HSC3-C17 cells, which showed similar activity. The conditioned medium from HSC3-C13 cells significantly increased the number of osteoclasts in a co-culture with mouse bone marrow cells and UAMS-32 cells. Xenograft tumors generated from these clonal cell lines into the periosteal region of the parietal bone in athymic mice showed that HSC3-C13 cells caused extensive bone destruction and a significant increase in osteoclast numbers as compared to HSC3-C17 cells. Gene expression was compared between HSC3-C13 and HSC3-C17 cells by using microarray analysis, which showed that CXCL2 gene was highly expressed in HSC3-C13 cells as compared to HSC3-C17 cells. Immunohistochemical staining revealed the localization of CXCL2 in human oral squamous cell carcinomas. The increase in osteoclast numbers induced by the HSC3-C13-conditioned medium was dose-dependently inhibited by addition of anti-human CXCL2-neutralizing antibody in a co-culture system. Recombinant CXCL2 increased the expression of Rankl in UAMS-32 cells. These results indicate that CXCL2 is involved in bone destruction induced by oral cancer. This is the first report showing the role of CXCL2 in cancer-associated bone destruction.
  Biochemical and Biophysical Research Communications 07/2012; 424(3):456-61. · 2.48 Impact Factor
• Article: Increasing participation of sclerostin in postnatal bone development, revealed by three-dimensional immunofluorescence morphometry.

  Takashi Watanabe, Yoshihiro Tamamura, Akiyoshi Hoshino, Yuji Makino, Hiroshi Kamioka, Teruo Amagasa, Akira Yamaguchi, Tadahiro Iimura
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  ABSTRACT: Confocal immunofluorescence tiling imaging revealed the spatio-temporal distributions of osterix and sclerostin in femurs from 3-day-old, 2-week-old and 4-week-old rats to be reciprocally exclusive at the tissue level. Further quantitative three-dimensional immuno fluorescence morphometry demonstrated the increasing distribution of sclerostin in the osteocytic lacuno-canalicular system specifically in diaphysis, which paralleled the cooperative participation and depletion of osterix and β-catenin in adjacent periosteum cells. Treating MC3T3-E1 cells with BIO (a GSK3 inhibitor) induced the stabilization of β-catenin and nuclear translocation of osterix, and negatively regulated osteocalcin/BGLAP and Dmp1. These results collectively demonstrate that the increasing distribution of sclerostin in diaphyseal cortical bone appears to be involved in the attenuation of osterix and β-catenin in adjacent periosteum cells, thus possibly contributing to osteoblast maturation and reducing the osteoblast formation at this bone site. Our confocal microscopy-based imaging analyses provide a comprehensive and detailed view of the spatio-temporal distribution of sclerostin, β-catenin and osterix at the tissue to subcellular level in a coherent manner, and uncovered their spatio-temporal cooperation in postnatal bone development, thus providing evidence that they link skeletogenic growth and functional bone development.
  Bone 07/2012; 51(3):447-58. · 4.02 Impact Factor
• Article: CCN3 and bone marrow cells

  Ken-ichi Katsube, Saki Ichikawa, Yuko Katsuki, Tasuku Kihara, Masanori Terai, Lester F. Lau, Yoshihiro Tamamura, Shin’ichi Takeda, Akihiro Umezawa, Kei Sakamoto, Akira Yamaguchi
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  ABSTRACT: CCN3 expression was observed in a broad variety of tissues from the early stage of development. However, a kind of loss of function in mice (CCN3 del VWC domain -/-) demonstrated mild abnormality, which indicates that CCN3 may not be critical for the normal embryogenesis as a single gene. The importance of CCN3 in bone marrow environment becomes to be recognized by the studies of hematopoietic stem cells and Chronic Myeloid Leukemia cells. CCN3 expression in bone marrow has been denied by several investigations, but we found CCN3 positive stromal and hematopoietic cells at bone extremities with a new antibody although they are a very few populations. We investigated the expression pattern of CCN3 in the cultured bone marrow derived mesenchymal stem cells and found its preference for osteogenic differentiation. From the analyses of in vitro experiment using an osteogenic mesenchymal stem cell line, Kusa-A1, we found that CCN3 downregulates osteogenesis by two different pathways; suppression of BMP and stimulation of Notch. Secreted CCN3 from Kusa cells inhibited the differentiation of osteoblasts in separate culture, which indicates the paracrine manner of CCN3 activity. CCN3 may also affect the extracellular environment of the niche for hematopoietic stem cells.
  Journal of Cell Communication and Signaling 04/2012; 3(2):135-145.
• Article: Effects of veraguensin and galgravin on osteoclast differentiation and function.

  Midori Asai, Ji-Won Lee, Yasunori Itakura, Bong-Keun Choi, Takayuki Yonezawa, Toshiaki Teruya, Byung-Yoon Cha, Motoko Ohnishi, Akira Yamaguchi, Je-Tae Woo
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  ABSTRACT: The dried flower buds of Magnolia sp. are widely used as herbal medicines because of their anti-inflammatory, anti-malarial and anti-platelet activities. Here, we found that veraguensin and galgravin, lignan compounds derived from Magnolia sp., dose-dependently inhibited osteoclast formation in co-cultures of bone marrow cells and osteoblastic cells. These compounds also inhibited receptor activator of nuclear factor κB ligand (RANKL)-induced osteoclast differentiation in RAW264.7 cells and bone marrow macrophages. In the RANKL-induced signaling pathway, veraguensin and galgravin reduced p38 phosphorylation and suppressed the expression of c-Fos, a key transcription factor for osteoclastogenesis. Veraguensin and galgravin also inhibited osteoclastic pit formation, which was accompanied by decreased mature osteoclast viability. In conclusion, these results indicate that veraguensin and galgravin can inhibit bone resorption and may offer novel compounds for the development of drugs to treat bone-destructive diseases such as osteoporosis.
  Cytotechnology 04/2012; 64(3):315-22. · 1.21 Impact Factor
• Article: Reduction of NOTCH1 expression pertains to maturation abnormalities of keratinocytes in squamous neoplasms.

  Kei Sakamoto, Takuma Fujii, Hiroshi Kawachi, Yoshio Miki, Ken Omura, Kei-ichi Morita, Kou Kayamori, Ken-ichi Katsube, Akira Yamaguchi
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  ABSTRACT: Notch is a transmembrane receptor functioning in the determination of cell fate. Abnormal Notch signaling promotes tumor development, showing either oncogenic or tumor suppressive activity. The uncertainty about the exact role of Notch signaling, partially, stems from inconsistencies in descriptions of Notch expression in human cancers. Here, we clarified basal-cell dominant expression of NOTCH1 in squamous epithelium. NOTCH1 was downregulated in squamous neoplasms of oral mucosa, esophagus and uterine cervix, compared with the normal basal cells, although the expression tended to be retained in cervical lesions. NOTCH1 downregulation was observed even in precancers, and there was little difference between cancers and high-grade precancerous lesions, suggesting its minor contribution to cancer-specific events such as invasion. In culture experiments, reduction of NOTCH1 expression resulted in downregulation of keratin 13 and keratin 15, and upregulation of keratin 17, and NOTCH1 knockdown cells formed a dysplastic stratified epithelium mimicking a precancerous lesion. The NOTCH1 downregulation and the concomitant alterations of those keratin expressions were confirmed in the squamous neoplasms both by immunohistochemical and cDNA microarray analyses. Our data indicate that reduction of NOTCH1 expression directs the basal cells to cease terminal differentiation and to form an immature epithelium, thereby playing a major role in the histopathogenesis of epithelial dysplasia. Furthermore, downregulation of NOTCH1 expression seems to be an inherent mechanism for switching the epithelium from a normal and mature state to an activated and immature state, suggesting its essential role in maintaining the epithelial integrity.
  Laboratory Investigation 02/2012; 92(5):688-702. · 3.64 Impact Factor
• Article: [Bone and tooth in calcium and phosphate metabolism].

  Yoshihiro Tamamura, Akira Yamaguchi
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  ABSTRACT: Tight regulation of serum concentrations of calcium and phosphate is indispensable for maintaining normal physiological condition. Imbalance of this regulation leads to pathophysiological disorders including heart disease, chronic kidney disease, and ectopic calcification. Formation and mineralization of bone and tooth are greatly influenced by calcium and phosphate metabolism since both organs are mainly consist of calcium-phosphate. Calcium and phosphate homeostasis is under hormonal control on its target organs such as kidney, bone and intestine. Calcium and phosphate are absorbed in intestine and reabsorbed and excreted in kidney. Bone store and release them in response to changing physiological demand by osteoblastic bone formation and osteoclastic bone resorption. Bone is also important as an endocrine organ that releases FGF23 from osteocytes, a novel hormone that targets the kidney to inhibit phosphate reabsorption and 1α, 25 (OH) (2)D(3) production.
  Clinical calcium 01/2012; 22(1):11-7.
• Article: Clinical significance of lymphatic and blood vessel invasion in oral tongue squamous cell carcinomas.

  Chieko Michikawa, Narikazu Uzawa, Kou Kayamori, Itaru Sonoda, Yoshio Ohyama, Norihiko Okada, Akira Yamaguchi, Teruo Amagasa
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  ABSTRACT: Although vascular invasion (VI) is recognized as an important predictor of lymph node metastasis and a significant prognostic factor in head and neck squamous cell carcinoma (HNSCC), there is currently no common definition for the pathological evaluation of VI status. We reviewed the medical records of 63 consecutive resected primary oral tongue SCCs (OTSCCs) without preoperative treatment between June 1999 and April 2008, and evaluated VI status by investigating lymphatic vessel invasion (LVI) and blood vessel invasion (BVI) by using immunohistochemistry (IHC) with monoclonal antibody D2-40 (D2-40) and Elastica van Gieson (EVG) staining, respectively. Subsequently, we analyzed their correlations with cervical lymph node metastasis and prognosis. LVI was found in 16 of the 63 tumors (25.4%) and BVI was in 32 tumors (50.8%). Univariate analysis revealed that the presence of LVI is statistically correlated with lymph node metastasis. Moreover, multivariate logistic regression analysis revealed that LVI is an independent risk factor of nodal metastasis (odds ratio=4.262, 95% confidence interval=1.262-14.397, p=0.020). In contrast, Kaplan-Meier survival analysis revealed that patients with BVI had a significantly shorter disease-free survival (DFS) and overall survival (OS) rates than those without BVI (68.6% versus 90.3%, p=0.028 and 68.6% versus 93.5%, p=0.013, respectively). The present study clearly demonstrated that LVI at primary OTSCC had significant correlation with lymph node metastasis, and that BVI was significantly associated with recurrence and poor prognosis. Evaluation of VI status, as LVI and BVI status separately, using IHC with D2-40 and EVG staining may be useful in predicting lymph node metastasis and poor prognosis in OTSCCs.
  Oral Oncology 12/2011; 48(4):320-4. · 2.86 Impact Factor
• Article: Structural differences in the osteocyte network between the calvaria and long bone revealed by three-dimensional fluorescence morphometry, possibly reflecting distinct mechano-adaptations and sensitivities.

  Akiko Himeno-Ando, Yuichi Izumi, Akira Yamaguchi, Tadahiro Iimura
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  ABSTRACT: The structural features of osteocytes and their cellular process network are thought to allow for mechanotransduction from the bone tissue to these cells. This study applied three-dimensional fluorescence microscopy to fixed and decalcified bone specimens to quantitatively compare the osteocytes and their networks between mouse parietal bone and tibia that are physiologically enforced by distinct mechanical loads. The subsequent morphometric analysis by the surface rendering of osteocyte cell bodies revealed the tibia to have relatively enriched cytoplasm in the osteocyte cell body in comparison to the parietal bone. Furthermore, quantitative tracing of the cellular processes in silico demonstrated that the numbers of the cellular processes and their bifurcation points per osteocyte in the tibia were significantly higher than those in the parietal bone. Though the total length of the processes per osteocyte in the tibia was two times longer, its total surface area and total volume were smaller than those in the parietal bone, due to its thinner diameter. These architectural differences in the osteocytes and their networks are thus implicated in the adaptation to physiologically different loading, and may also induce distinct mechanosensitivities.
  Biochemical and Biophysical Research Communications 12/2011; 417(2):765-70. · 2.48 Impact Factor
• Article: A fluorescence spotlight on the clockwork development and metabolism of bone.

  Tadahiro Iimura, Ayako Nakane, Mayu Sugiyama, Hiroki Sato, Yuji Makino, Takashi Watanabe, Yuzo Takagi, Rika Numano, Akira Yamaguchi
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  ABSTRACT: Biological phenomena that exhibit periodic activity are often referred as biorhythms or biological clocks. Among these, circadian rhythms, cyclic patterns reflecting a 24-h cycle, are the most obvious in many physiological activities including bone growth and metabolism. In the late 1990s, several clock genes were isolated and their primary structures and functions were identified. The feedback loop model of transcriptional factors was proposed to work as a circadian core oscillator not only in the suprachiasmatic nuclei of the anterior hypothalamus, which is recognized as the mammalian central clock, but also in various peripheral tissues including cartilage and bone. Looking back to embryonic development, the fundamental architecture of skeletal patterning is regulated by ultradian clocks that are defined as biorhythms that cycle more than once every 24 h. As post-genomic approaches, transcriptome analysis by micro-array and bioimaging assays to detect luminescent and fluorescent signals have been exploited to uncover a more comprehensive set of genes and spatio-temporal regulation of the clockwork machinery in animal models. In this review paper, we provide an overview of topics related to these molecular clocks in skeletal biology and medicine, and discuss how fluorescence imaging approaches can contribute to widening our views of this realm of biomedical science.
  Journal of Bone and Mineral Metabolism 07/2011; 30(3):254-69. · 2.27 Impact Factor
• Article: Comparative morphology of the osteocyte lacunocanalicular system in various vertebrates.

  Lei Cao, Takeshi Moriishi, Toshihiro Miyazaki, Tadahiro Iimura, Miwako Hamagaki, Ayako Nakane, Yoshihiro Tamamura, Toshihisa Komori, Akira Yamaguchi
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  ABSTRACT: Osteocytes are embedded in the bone matrix, and they communicate with adjacent osteocytes, osteoblasts, and osteoclasts through the osteocyte lacunocanalicular system. Osteocytes are believed to be essential for the maintenance of bone homeostasis because they regulate mechanical sensing and mineral metabolism in mammalian bones; however, osteocyte morphology in other vertebrates has not been well documented. We conducted a comparative study on the morphology of osteocytes and the lacunocanalicular system of the following vertebrates: two teleost fishes [medaka (Oryzias latipes), and zebrafish (Danio rerio)], three amphibians [African clawed frog (Xenopus laevis), black-spotted pond frog (Rana nigromaculata), and Japanese fire-bellied newt (Cynops pyrrhogaster)], two reptiles [four-toed tortoise (Testudo horsfieldii) and green iguana (Iguana iguana)], and two mammals (laboratory mouse C57BL6 and human). The distribution of the osteocyte lacunocanalicular system in all these animals was investigated using the modified silver staining and the fluorescein-conjugated phalloidin staining methods. Bones of medaka had few osteocytes (acellular bone). Bones of zebrafish contained osteocytes (cellular bone) but had a poorly developed osteocyte lacunocanalicular system. Bones of Xenopus laevis, a freshwater species, and of other amphibians, reptiles, and mammals contained numerous osteocytes and a well-developed lacunocanalicular system. The present study indicates that development of the osteocyte lacunocanalicular system differs between teleost fishes and land vertebrates, but this pattern is not directly related to aquatic habitat.
  Journal of Bone and Mineral Metabolism 04/2011; 29(6):662-70. · 2.27 Impact Factor
• Article: Down-regulation of keratin 4 and keratin 13 expression in oral squamous cell carcinoma and epithelial dysplasia: a clue for histopathogenesis.

  Kei Sakamoto, Tadanobu Aragaki, Kei-ichi Morita, Hiroshi Kawachi, Kou Kayamori, Shoichi Nakanishi, Ken Omura, Yoshio Miki, Norihiko Okada, Ken-Ichi Katsube, Toichiro Takizawa, Akira Yamaguchi
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  ABSTRACT: This study aimed to identify relevant keratin subtypes that may associate with the pathogenesis of oral epithelial neoplasms. Expression of all the keratin subtypes was examined by cDNA microarray analysis of 43 oral squamous cell carcinoma (OSCC) cases. Immunohistochemical expression of the major keratins was examined in 100 OSCC and oral epithelial dysplasia (OED) cases. Many changes in keratin expression were observed and, significantly, consistent down-regulation of keratin 4 (K4) and K13 expression was observed. Aberrant expression of K4 and K13 was associated with morphological changes in the affected oral epithelium. Experiments with cell cultures transfected with various keratin subtypes suggested that alterations in keratin subtype expression can cause changes in cell shape and movement. Aberrant expression of K4 and K13, which are the dominant pair of differentiation-related keratins in oral keratinocytes, indicates dysregulation of epithelial differentiation in OSCC and OED. These keratins, especially K4, may be useful for pathological diagnosis. We propose that the aberrant expression of K4 and K13 and concomitant up-regulation of the other keratins may be one of the causative factors for morphological alterations in the affected epithelium.
  Histopathology 03/2011; 58(4):531-42. · 3.08 Impact Factor
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  Available from: PubMed Central

  Article: Küttner's tumor of the sub-mandibular gland associated with fibrosclerosis and follicular hyperplasia of regional lymph nodes: a case report.

  Yumi Mochizuki, Ken Omura, Kou Kayamori, Kei Sakamoto, Hiroaki Shimamoto, Akira Yamaguchi
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  ABSTRACT: Küttner's tumor is characterized through histology by peri-ductal fibrosis, dense lymphocytic infiltration with lymphoid follicles, loss of acini, and occasional marked sclerosis of the salivary gland. On occasion, Küttner's tumor can be difficult to distinguish from malignant neoplasm. A 58-year-old Japanese man was referred to our hospital with a three-month history of a painless swollen mass in the right sub-mandibular region. Histological findings revealed both lymphoid follicles with reactive germinal centers and variously sized lymphoid follicle-like nodules without definitive germinal centers or mantle zones. B-cells of similar size and shape occupied the lymphoid follicle-like nodules and stained positive for B-cell lymphoma. These cells were detected in the polyclonal B-cells by flow cytometric analysis and tested negative for CD10. Unusual B-cell proliferation was observed, but as there was no definitive evidence of B-cell lymphoma, the lesion was diagnosed as Küttner's tumor. We report on a rare case of Küttner's tumor associated with fibrosclerosis and atypical lymphoid hyperplasia in both the sub-mandibular gland and regional lymph nodes. Although more cases need to be investigated, our findings might be helpful to further studies seeking to clarify the etiology of idiopathic sclerosing lesions arising in the organs and regional lymph nodes.
  Journal of Medical Case Reports 03/2011; 5:121.
• Article: Acerogenin A, a natural compound isolated from Acer nikoense Maxim, stimulates osteoblast differentiation through bone morphogenetic protein action.

  Tasuku Kihara, Saki Ichikawa, Takayuki Yonezawa, Ji-Won Lee, Toshihiro Akihisa, Je Tae Woo, Yasuyuki Michi, Teruo Amagasa, Akira Yamaguchi
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  ABSTRACT: We investigated the effects of acerogenin A, a natural compound isolated from Acer nikoense Maxim, on osteoblast differentiation by using osteoblastic cells. Acerogenin A stimulated the cell proliferation of MC3T3-E1 osteoblastic cells and RD-C6 osteoblastic cells (Runx2-deficient cell line). It also increased alkaline phosphatase activity in MC3T3-E1 and RD-C6 cells and calvarial osteoblastic cells isolated from the calvariae of newborn mice. Acerogenin A also increased the expression of mRNAs related to osteoblast differentiation, including Osteocalcin, Osterix and Runx2 in MC3T3-E1 cells and primary osteoblasts: it also stimulated Osteocalcin and Osterix mRNA expression in RD-C6 cells. The acerogenin A treatment for 3days increased Bmp-2, Bmp-4, and Bmp-7 mRNA expression levels in MC3T3-E1 cells. Adding noggin, a BMP specific-antagonist, inhibited the acerogenin A-induced increase in the Osteocalcin, Osterix and Runx2 mRNA expression levels. These results indicated that acerogenin A stimulates osteoblast differentiation through BMP action, which is mediated by Runx2-dependent and Runx2-independent pathways.
  Biochemical and Biophysical Research Communications 02/2011; 406(2):211-7. · 2.48 Impact Factor
• Article: Comprehensive keratin profiling reveals different histopathogenesis of keratocystic odontogenic tumor and orthokeratinized odontogenic cyst.

  Tadanobu Aragaki, Yasuyuki Michi, Ken-ichi Katsube, Narikazu Uzawa, Norihiko Okada, Takumi Akashi, Teruo Amagasa, Akira Yamaguchi, Kei Sakamoto
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  ABSTRACT: Keratocystic odontogenic tumor is a cystic lesion that behaves more aggressively than other jaw cysts. One of its characteristic histologic features is a parakeratinized uniform layer of lining epithelium. A jaw cyst lined with orthokeratinized epithelium is called an orthokeratinized odontogenic cyst. These keratinized jaw cysts are thought to be separate entities, although their histopathogenesis has not been fully assessed. To better understand these lesions, we performed comprehensive immunohistochemical profiling of the keratin expression of each. Orthokeratinized odontogenic cysts expressed keratin 1, keratin 2, keratin 10, and loricrin, suggesting differentiation toward normal epidermis. Keratocystic odontogenic tumors expressed keratin 4, keratin 13, keratin 17, and keratin 19, which is a unique expression pattern reminiscent of a mucosal squamous epithelium and an epithelial appendage. In neonatal rat tooth germ, cells strongly positive for keratin 17 and keratin 19 were observed, specifically in the dental lamina, implying the origin of keratocystic odontogenic tumor. GLI2, a downstream effector of hedgehog signaling, was significantly expressed in keratocystic odontogenic tumor and basal cell carcinoma, accompanied with robust expression of keratin 17, mammalian target of rapamycin, and BCL2. The expression of these GLI2- or keratin 17-related factors was not significantly observed in orthokeratinized odontogenic cysts. These findings provide evidence to support the viewpoint that keratocystic odontogenic tumor and orthokeratinized odontogenic cyst are separate entities, and furthermore suggest their characteristic histology, pathogenesis, and biological behaviors.
  Human pathology 12/2010; 41(12):1718-25. · 3.03 Impact Factor