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Akira Myoui

Publications and source records attributed to Akira Myoui.

At least 37 records · Page 2Linked to original sources
PubMed2005

The effect of simulated microgravity by three-dimensional clinostat on bone tissue engineering.

Evidence suggests that mechanical stress, including gravity, is associated with osteoblast differentiation and function. To examine effects of microgravity on bone tissue engineering, we used a three-dimensional (3D) clinostat manufactured by Mitsubishi Heavy Industries (Kobe, Japan). A 3D clinostat is a device that generates multidirectional G force. By controlled rotation on two axes, it cancels the cumulative gravity vector at the center of the device. We cultured rat marrow mesenchymal cells (MMCs) in the pores of interconnected porous calcium hydroxyapatite (IP-CHA) for 2 weeks in the presence of dexamethasone using the 3D clinostat (clinostat group). MMCs cultured using the 3D clinostat exhibited a 40% decrease in alkaline phosphatase activity (a marker of osteoblastic differentiation), compared with control static cultures (control group). SEM analysis revealed that although there was no difference between the two groups in number or distribution of cells in the pores, the clinostat group exhibited less extensive extracellular matrix formation than the control group. Cultured IP-CHA/MMC composites were then implanted into subcutaneous sites of syngeneic rats and harvested 8 weeks after implantation. All implants showed bone formation inside the pores, as indicated by decalcified histological sections and microfocus computed tomography. However, the volume of newly formed bone was significantly lower for the clinostat group than for the control group, especially in the superficial pores close to the implant surface. These results indicate that new bone formation in culture was inhibited by use of the 3D clinostat, and that this inhibition was mainly due to suppression of osteoblastic differentiation of MMCs.

Animals↗
PubMed2004

Crisscross CTL induction by SYT-SSX junction peptide and its HLA-A*2402 anchor substitute.

To investigate the effects of anchor substitutions in SYT-SSX junction peptide, an HLA-A24 anchor residue (position 9) of the SYT-SSX B peptide (GYDQIMPKK) was substituted to more favorable residues according to the HLA-A24-binding motif. Among four substitutes constructed, a substitute with isoleucine (termed K9I peptide) most apparently enhanced the affinity for HLA-A24 molecule. Subsequent in vitro CTL induction analysis using PBMCs of 15 HLA-A24(+) synovial sarcoma patients revealed that the original B peptide allowed to induce synovial sarcoma-specific CTLs from 7 patients (47%), whereas such CTLs were inducible from 12 patients (80%) with K9I peptide. Moreover, the extent of cytotoxicity against HLA-A24(+) synovial sarcoma cell lines was higher in K9I peptide-induced CTLs than B peptide-induced CTLs. Influence of anchor substitution on peptide/TCR interaction was evaluated by cytotoxicity assays against autologous cells and tetramer analysis. CTLs induced from a synovial sarcoma patient using K9I peptide did not lyse autologous PHA blasts or EBV-infected B cells. In vitro stimulations of PBMCs from 5 HLA-A24(+) synovial sarcoma patients with K9I peptide increased the frequency of T cells reacting with both HLA-A24/K9I peptide tetramer and HLA-A24/B peptide tetramer. In contrast, the frequency of T cells reacting with HLA/HIV-derived peptide tetramer remained low. These findings support the validity in design of anchor residue substitution in SYT-SSX fusion gene-derived peptide, and provide a potential clue to the current stagnation in vaccination trials of fusion gene-derived natural junction peptides.

Amino Acid Substitution↗
PubMed2004

Smad6/Smurf1 overexpression in cartilage delays chondrocyte hypertrophy and causes dwarfism with osteopenia.

Biochemical experiments have shown that Smad6 and Smad ubiquitin regulatory factor 1 (Smurf1) block the signal transduction of bone morphogenetic proteins (BMPs). However, their in vivo functions are largely unknown. Here, we generated transgenic mice overexpressing Smad6 in chondrocytes. Smad6 transgenic mice showed postnatal dwarfism with osteopenia and inhibition of Smad1/5/8 phosphorylation in chondrocytes. Endochondral ossification during development in these mice was associated with almost normal chondrocyte proliferation, significantly delayed chondrocyte hypertrophy, and thin trabecular bone. The reduced population of hypertrophic chondrocytes after birth seemed to be related to impaired bone growth and formation. Organ culture of cartilage rudiments showed that chondrocyte hypertrophy induced by BMP2 was inhibited in cartilage prepared from Smad6 transgenic mice. We then generated transgenic mice overexpressing Smurf1 in chondrocytes. Abnormalities were undetectable in Smurf1 transgenic mice. Mating Smad6 and Smurf1 transgenic mice produced double-transgenic pups with more delayed endochondral ossification than Smad6 transgenic mice. These results provided evidence that Smurf1 supports Smad6 function in vivo.

Animals↗
PubMed2004

New cell lines with chondrocytic phenotypes from human chondrosarcoma.

In the present study, we investigated chondrocytic characterization for newly established human chondrosarcoma cell lines. A chondrosarcoma cell line, HCS-TG, was established by the implantation of grade-2 human chondrosarcoma into athymic mice. Cloning of HCS-TG cells from passage 17 was performed. After cell cloning, two clonal-cell lines (HCS-TG C3 and E2) with good proliferative activities were obtained. These cell lines in monolayer culture retained a polygonal morphology. Their doubling times were 68 h and 45 h, respectively. mRNA expressions of type-I, -II, -X, and -XI collagens and aggrecan core protein were detected on reverse-transcription polymerase chain reaction. Protein expression of type-II collagen was confirmed in each cell line using Western blotting. However, there was no expression of type-I collagen. Moreover, gelatin zymography revealed that both cell lines produced extracellular matrices with matrix metalloproteinases 2 and 9. The parental HCS-TG cells had tumorigenicity in athymic mice; however, C3 and E2 were not tumorigenic. New clonal-cell lines HCS-TG C3 and E2 derived from human chondrosarcoma are morphologically chondrocytic in serial monolayer cultures and express chondrocytic phenotypes.

Adult↗
PubMed2004

Localization of RANKL in osteolytic tissue around a loosened joint prosthesis.

Osteoclastogenesis is a key event of the cellular reaction in prosthetic loosening. Immunohistochemistry and reverse transcription-polymerase chain reaction (RT-PCR) were used to study the localization and expression of receptor activator of nuclear factor kappa B ligand (RANKL), a potent factor for osteoclastogenesis in the membranous tissue formed around loosened prosthetic joint implants. RANKL was identified in a wide variety of cells appearing in this membranous tissue. At least three types of RANKL-positive cells were identified, including prolyl 4-hydroxylase (PH)-positive fibroblast lineage cells, CD68 cells, and tartrate-resistant acid phosphatase (TRAP)-positive mononuclear and multi-nucleated macrophage lineage cells. Tumor necrosis factor (TNF)-alpha-converting enzyme (TACE) was colocalized with RANKL in these cells, suggesting the in-situ release of this factor. RT-PCR confirmed the actual expression of the RANKL and TACE genes in the tissues around the loosened implant. These observational findings indicate the possible synthesis of RANKL by fibroblast and macrophage lineage cells, and suggest the in-situ involvement of RANKL in both osteoclastogenesis and osteoclastic bone resorptive events occurring in prosthetic joint loosening.

ADAM Proteins↗
PubMed2004

Bone morphogenetic proteins in bone tumors.

Bone morphogenetic proteins (BMPs), inducers of ectopic bone formation in vivo, are present in a number of osteosarcomas. BMPs are responsible for reactive bone formation, including periosteal reactions by normal osteoblasts, rather than production of tumorous osteoid by tumor cells. Osteosarcomas producing BMPs contain less-differentiated mesenchymal cells, resulting in a poorer prognosis for those patients. BMPs are also expressed in malignant fibrous histiocytomas (MFHs) of bone and dedifferentiated chondrosarcomas exhibiting undifferentiated features. However, BMPs in MFH do not show any osteoinductive activity in vivo, suggesting that those BMPs may be inactive forms and have additional functions unrelated to bone formation. Among benign bone tumors, BMPs are expressed in osteoid osteomas or osteoblastomas and effect reactive bone formation such as a surrounding sclerosis. BMPs and a BMP receptor (BMPRIB) are also detected in the cartilage cap in osteochondroma, suggesting that BMP signaling via BMPRIB might be involved in the pathogenesis of osteochondroma. Clinically, BMPs have utility as diagnostic and prognostic markers for characterizing the stage of differentiation of mesenchymal cells and mesenchymal tumors, and they may be of value in predicting the prognosis of sarcoma patients. This article reviews the accumulated information on BMPs in bone tumors, including the most recent findings, and discusses the biological and clinical significance of BMPs in bone tumors.

Animals↗
PubMed2004

Interleukin-6/soluble interleukin-6 receptor signaling attenuates proliferation and invasion, and induces morphological changes of a newly established pleomorphic malignant fibrous histiocytoma cell line.

Pleomorphic malignant fibrous histiocytoma (MFH) is occasionally associated with inflammatory paraneoplastic syndrome (PNS). Recently, we reported that interleukin (IL)-6, one of the candidate cytokines, which induces such systemic inflammatory reaction, may be a tumor-associated factor involved in the pathogenesis and its clinical manifestations of MFH. In the local microenvironment, tumor-induced inflammatory reaction may play a role favoring tumor progression. To clarify the biological relevance of IL-6 in MFH, we established a human MFH cell line, named MIPS-2, derived from a resected specimen of a patient presenting with PNS. In this patient, the serum IL-6 level ran parallel to the disease course: elevated serum IL-6 concentration normalized immediately after radical surgery, and re-elevation occurred on tumor recurrence. MIPS-2 presented pleomorphic appearance, severe nuclear abnormalities with prominent nucleoli, and tumorigenesis in nude mice. MIPS-2 expressed IL-6, IL-6 receptor (IL-6R), and glycoprotein 130 (gp130) but lacked the soluble form of IL-6R (sIL-6R), as determined by flow cytometry and reverse transcriptase-polymerase chain reaction analyses. Stimulation of MIPS-2 with IL-6 combined with exogenous sIL-6R induced phosphorylation of both signal transducer and activator of transcription 3 (STAT3) and mitogen-activated protein kinase (MAPK), decreased cell proliferation, attenuated invasion, and induced morphological changes. Collectively, these data suggested that the IL-6/sIL-6R signaling pathway plays a pivotal role for proliferation, invasion, and morphology of MFH via STAT3 and MAPK pathway as autocrine and/or paracrine manner, and proposed the therapeutic potential for the use of both anti-growth factor and proinflammatory cytokine-targeting strategies to combat devastating MFH.

Adult↗
PubMed2004

Synovial sarcoma after chemotherapy for osteosarcoma: a case report.

A 23-year-old man had eight cycles of adjuvant chemotherapy including doxorubicin, cisplatin, methotrexate, and ifosfamide and radical surgery for biopsy-proved osteosarcoma of the right fibular shaft. Two years after the initial diagnosis, he noticed a mass in the medial aspect of his right knee. Magnetic resonance imaging scans revealed a soft tissue tumor measuring 2 x 2 cm in the pericapsular region of the right knee. Histologically, this soft tissue tumor was composed of spindle cells with occasional atypical mitoses and without matrix formation. Immunohistochemically, the tumor cells were positive for vimentin, cytokeratin, and epithelial membrane antigen, and negative for alpha smooth muscle actin. A fusion gene, SYT-SSX was detected with reverse transcription-polymerase chain reaction. From the results, the secondary tumor was diagnosed as a synovial sarcoma. The current case of double sarcomas is rare. Both sarcomas were diagnosed accurately using immunohistochemical and molecular procedures. This case suggests a positive association between a second tumor and chemotherapy including intraarterial perfusion of doxorubicin.

Adult↗
PubMed2004

Bone tissue engineering using novel interconnected porous hydroxyapatite ceramics combined with marrow mesenchymal cells: quantitative and three-dimensional image analysis.

We developed fully opened interconnected porous calcium hydroxyapatite ceramics having two different pore sizes. One has pores with an average size of 150 microm in diameter, an average 40-microm interconnecting pore diameter, and 75% porosity (HA150). The other has pores with an average size of 300 microm in diameter, an average 60-100-microm interconnecting pore diameter, and 75% porosity (HA300). Because of its smaller pore diameter, HA150 has greater mechanical strength than that of HA300. These ceramics were combined with rat marrow mesenchymal cells and cultured for 2 weeks in the presence of dexamethasone. The cultured ceramics were then implanted into subcutaneous sites in syngeneic rats and harvested 2-8 weeks after implantation. All the implants showed bone formation inside the pore areas as evidenced by decalcified histological sections and microcomputed tomography images, which enabled three-dimensional analysis of the newly formed bone and calculation of the bone volume in the implants. The bone volume increased over time. At 8 weeks after implantation, extensive bone volume was detected not only in the surface pore areas but also in the center pore areas of the implants. A high degree of alkaline phosphatase activity with a peak at 2 weeks and a high level of osteocalcin with a gradual increase over time were detected in the implants. The levels of these biochemical parameters were higher in HA150 than in HA300. The results indicate that a combination of HA150 and mesenchymal cells could be used as an excellent bone graft substitute because of its mechanical properties and capability of inducing bone formation.

Animals↗
PubMed2004

Prediction of response to neoadjuvant chemotherapy for osteosarcoma by gene-expression profiles.

To establish a method for predicting the response to chemotherapy for osteosarcoma (OS), we performed expression profile analysis using cDNA microarray consisting of 23,040 genes. Hierarchical clustering based on the expression profiles of 19 biopsy samples of OS demonstrated two major clusters, one of which consisted exclusively of typical OS, i.e. conventional central OS in long bone of patients in the second decade. A set of genes was identified to characterize this subgroup, some of which have previously indicated some relation to carcinogenesis. Thirteen of the 19 patients were treated with an identical protocol of chemotherapy containing doxorubicin, cis-platinum and ifosfamide, and histological examination of resected specimens after operation classified 6 cases as responder and 7 as non-responder. A comparison of expression profiles of these two groups identified 60 genes whose expression levels were likely to be correlated with the response to chemotherapy (P<0.008). A drug response scoring (DRS) system was developed based on the expression levels of these genes, which proved to be applicable to predict the response to chemotherapy irrespective for the subclassification of OS. The reliability of the DRS system was further confirmed by testing additional 5 OS cases. These results indicated that scoring system based on gene-expression profiles might be useful to predict the response to chemotherapy for OS.

Adolescent↗
PubMed2003

Monoclonal antibody to parathyroid hormone-related protein induces differentiation and apoptosis of chondrosarcoma cells.

We investigated the effects of treatment with anti-parathyroid hormone-related protein (1-34) monoclonal murine antibody (anti-PTHrP MoAb) on apoptosis and the differentiation of chondrosarcoma HTB-94 cells. Treatment with anti-PTHrP MoAb accelerated apoptosis of HTB-94 cells in a dose-dependent manner, and anti-PTHrP MoAb also promoted the chondrogenic differentiation of HTB-94 cells. The induction of apoptosis by anti-PTHrP MoAb via imbalance of Bcl-2/Bax ratio and activation of caspase-3 may provide a mechanistic explanation for its potential antitumor effects. Our results suggest the possibility that anti-PTHrP MoAb may be beneficial as a new treatment for chondrosarcoma.

Antibodies, Monoclonal↗
PubMed2003

C-SRC tyrosine kinase activity is associated with tumor colonization in bone and lung in an animal model of human breast cancer metastasis.

The proto-oncogene, c-src, has been implicated in the tumorigenesis in breast cancer. However, the relationship of c-src with distant metastasis is unclear. Moreover, the role of c-src in organ-preferential metastasis of breast cancer is unknown. Because breast cancer has a strong predilection for metastasizing to bone, we examined the role of c-src in bone metastases using an animal model in which inoculation of the MDA-231 human breast cancer cells into the left cardiac ventricle preferentially developed osteolytic bone metastases in female nude mice. A clone of the MDA-231 with the increased capacity of bone metastasis exhibited elevated c-src tyrosine kinase (TK) activity compared with parental cells. MDAsrc527 cells caused significantly increased size of the osteolytic bone metastases with increased number of osteoclasts and mitotic cancer cells compared with MDA-231EV or MDAsrcWT. In contrast, MDAsrc295 cells caused impaired metastases to bone. Of note, mice inoculated with MDAsrc295 cells via tail vein developed reduced lung metastases and prolonged survival compared with mice with MDA-231EV cells, suggesting that c-src TK is unlikely to play a specific role in bone metastases. The growth in vitro and in vivo and production of parathyroid hormone-related protein, a key cytokine in the pathogenesis of osteolytic bone metastases in breast cancer, were promoted in MDAsrc527 and diminished in MDAsrc295. These results suggest that c-src TK is associated with the capacity of breast cancer to metastasize to bone through regulating cell growth and parathyroid hormone-related protein production. Our results together with the fact that c-src is an essential molecule for bone resorption by osteoclasts, which are central players in osteolytic bone metastases, support the notion that c-src TK is a potential target molecule for designing novel therapeutic interventions, especially for bone metastases in breast cancer.

Animals↗
PubMed2003

Extraskeletal myxoid chondrosarcoma: a Multi-Institutional Study of 42 Cases in Japan.

BACKGROUND: Extraskeletal myxoid chondrosarcoma (EMC) is a rare malignant neoplasm. Despite a consensus for the distinct clinicopathologic entity of EMC, its clinical features remain controversial. In addition, most studies have contained a small number of patients who underwent definitive surgical treatment. METHODS: Forty-two cases of EMC, which had been identified from files of eight affiliated hospitals and confirmed for histologic diagnosis at the Pathology Center, were analyzed for histologic grade, demographics, treatments, outcomes, and prognostic factors. The average follow-up period was 7.4 years. RESULTS: Included in the study were 20 men and 22 women with a mean age at diagnosis of 52.1 years. The tumors were located mainly in the lower extremities (69%). Thirty-three tumors (79%) were classified as Grade 1 and nine as Grade 2 according to the modified French System. Overall survival was 100% at 5 years and 88% at 10 years. Disease-free survival was 45% at 5 years and 36% at 10 years. Inadequate initial surgery was defined as a significant risk factor for local recurrence by univariate analysis of all 42 patients but not by the analysis of those 30 patients who had undergone wide tumor excision or amputation. Wide excision led to the recurrence rate of 14%. CONCLUSIONS: These findings supported the role of wide excision in the local control of EMC, irrespective of the previous excision procedure or recurrence. The protracted clinical course of the tumors and the presence of patients who had distant metastasis develop after definitive surgery of the primary tumor represented EMC as intermediate malignancy.

Adult↗
PubMed2003

COL11A2 collagen gene transcription is differentially regulated by EWS/ERG sarcoma fusion protein and wild-type ERG.

A specific t(21;22) chromosomal translocation creates the chimeric EWS/ERG gene in some cases of Ewing's sarcoma. In the resultant EWS/ERG fusion protein, the N-terminal part of the ETS family protein ERG is replaced by the N terminus of the RNA-binding protein EWS. We found that both the EWS/ERG and COL11A2 genes are expressed in the Ewing's sarcoma cell line, CADO-ES1. To investigate a potential role for EWS/ERG in COL11A2 gene expression, we characterized the COL11A2 promoter and tested the ability of wild-type ERG and EWS/ERG sarcoma fusion protein to transactivate COL11A2 promoter using a luciferase assay. We found that expression of EWS/ERG, but not wild-type ERG, transactivated the COL11A2 promoter and that this transactivation required not only the N-terminal region of EWS but also an intact DNA-binding domain from ERG. Electrophoretic mobility shift assay using COL11A2 promoter sequence showed involvement of EWS/ERG in the formation of DNA-protein complexes, and chromatin immunoprecipitation assay revealed direct interaction between COL11A2 promoter and EWS/ERG fusion protein in vivo. EWS/ERG, but not wild-type ERG, bound to RNA polymerase II. Treatment of cells with the histone deacetylase inhibitor trichostatin A enabled ERG to transactivate the COL11A2 promoter, therefore abolishing the differential effects of EWS/ERG and ERG. Taken together, these findings indicate that the COL11A2 gene is regulated both by potential ERG association with a histone deacetylase complex and by direct EWS/ERG recruitment of RNA polymerase II.

Animals↗
PubMed2003

Prevention of the onset and progression of collagen-induced arthritis in rats by the potent p38 mitogen-activated protein kinase inhibitor FR167653.

OBJECTIVE: FR167653 is a potent inhibitor of p38 mitogen-activated protein kinase (MAPK) and inhibits tumor necrosis factor alpha (TNFalpha) and interleukin-1 beta (IL-1 beta) production in inflammatory cells. In this study we investigated the effect of FR167653 on collagen-induced arthritis (CIA). METHODS: Rats with CIA were subcutaneously injected with FR167653 (32 mg/kg/day) starting on the day of the booster injection (day 7) in the prophylactic treatment group and after the onset of arthritis (day 21) in the therapeutic treatment group. Hind-paw swelling, body weight, radiographic and histologic scores, and osteoclast number were evaluated. Cytokine levels in the serum and tissue were assessed by enzyme-linked immunosorbent assays. Flow cytometric analysis of T lymphocytes from bone marrow was performed. The effect of FR167653 on in vitro osteoclast formation induced by soluble receptor activator of nuclear factor kappa B ligand (sRANKL) and TNFalpha was examined. RESULTS: Swelling of hind paws and loss of weight occurred in the CIA rats, but this was not evident in the prophylactic treatment group. Therapeutic treatment also significantly reduced paw swelling. The mean radiographic and histologic scores as well as the osteoclast numbers were significantly lower in the treatment group than in the CIA rats without treatment. FR167653 treatment reduced the serum levels of TNFalpha and IL-1 beta, lowered the IL-1 beta concentration in the ankle joints, and decreased the CD4-,CD8a+ T cell population in bone marrow. Furthermore, FR167653 inhibited the osteoclast-like cell differentiation induced by both sRANKL and TNFalpha in vitro. CONCLUSION: FR167653 prevents the onset of arthritis in a prophylactic treatment model and suppresses the progression of joint destruction in a therapeutic treatment model, suggesting that p38 MAPK is a potential therapeutic target for rheumatoid arthritis.

Animals↗
PubMed2003

Unusual skeletal metastases from myxoid liposarcoma only detectable by MR imaging.

We present two cases of skeletal metastases from myxoid liposarcoma, occurring several years after treatment of the primary tumors in the lower limb. The present two case reports have unusual radiological features only detectable by MR imaging and not by plain radiographs or bone scans. From the present two cases, we found that a negative plain radiograph of the spine or a negative bone scan could not exclude skeletal metastases from myxoid liposarcoma, and MRI was a more sensitive screening procedure for their detection, especially in T1-weighted images. Unusual radiological features of skeletal metastases from myxoid liposarcoma are not well documented and only a few cases have been previously reported. Our aim is to document two more patients exhibiting the unusual radiological features of skeletal metastases from myxoid liposarcoma to improve their early detection and management.

Adult↗
PubMed2003

Overexpression of the Wilms' tumor gene WT1 in human bone and soft-tissue sarcomas.

The expression levels of the Wilms' tumor gene WT1 were examined in 36 cases of various types of human bone and soft-tissue sarcomas using quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR). They included 12 malignant fibrous histiocytomas (MFH), 3 malignant peripheral nerve sheath tumors (MPNST), 6 synovial sarcomas (SyS), 4 myxoid liposarcomas (MyLS), one angiosarcoma (AGS), one clear cell sarcoma (CCS), and 9 osteosarcomas (OS). Eleven (92%) of 12 MFH, 2 (67%) of 3 MPNST, all (100%) of 6 SyS, 2 (50%) of 4 MyLS, one AGS, one CCS, and 5 (56%) of 9 OS cases overexpressed WT1 in the range of 1.4 x 10(-3)-3.9 x 10(-1) levels (WT1 expression level in K562 leukemic cells was defined as 1.0). Thus, 28 (78%) out of 36 various types of human bone and soft-tissue sarcomas overexpressed the WT1 gene. Immunohistochemical analysis showed positive staining for WT1 protein in all of 4 cases (one case each of MFH, MyLS, AGS and OS) with WT1 gene overexpression detected by RT-PCR analysis, demonstrating clearly that WT1 was expressed at the protein level in various types of human bone and soft-tissue sarcomas. The direct sequencing analysis of the WT1 genomic DNA showed no mutations in any of 10 exons of the WT1 gene in 8 different sarcoma samples (3 MFH, one SyS, one MyLS, one AGS, and 2 OS). The present study demonstrates that various types of human bone and soft-tissue sarcomas frequently overexpress the wild-type WT1 gene, suggesting an important role of the wild-type WT1 gene in tumorigenesis of various types of human bone and soft-tissue sarcomas.

Adult↗