Masahiko Miura

Tokyo University of Science, Tokyo, Tokyo-to, Japan

Are you Masahiko Miura?

Claim your profile

Publications (57)163.86 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Chk1 inhibitor acts as a potent radiosensitizer in p53-deficient tumor cells by abrogating the G2/M checkpoint. However, the effects of Chk1 inhibitor on the duration of G2 arrest have not been precisely analyzed. To address this issue, we utilized a cell-cycle visualization system, fluorescent ubiquitination-based cell-cycle indicator (Fucci), to analyze the change in the first green phase duration (FGPD) after irradiation. In the Fucci system, G1 and S/G2/M cells emit red and green fluorescence, respectively; therefore, G2 arrest is reflected by an elongated FGPD. The system also allowed us to differentially analyze cells that received irradiation in the red or green phase. Cells irradiated in the green phase exhibited a significantly elongated FGPD relative to cells irradiated in the red phase. In cells irradiated in either phase, Chk1 inhibitor reduced FGPD almost to control levels. The results of this study provide the first clear information regarding the effects of Chk1 inhibition on radiation-induced G2 arrest, with special focus on the time dimension.
    Journal of radiation research. 06/2014;
  • Atsushi Kaida, Masahiko Miura
    [Show abstract] [Hide abstract]
    ABSTRACT: In this study, we visualized the effect of tumor microenvironments on radiation-induced tumor cell kinetics. For this purpose, we utilized a multicellular spheroid model, with a diameter of ∼500 m, consisting of HeLa cells expressing the fluorescent ubiquitination-based cell-cycle indicator (Fucci). In live spheroids, a confocal laser scanning microscope allowed us to clearly monitor cell kinetics at depths of up to 60 μm. Surprisingly, a remarkable prolongation of G2 arrest was observed in the outer region of the spheroid relative to monolayer-cultured cells. Scale, an aqueous reagent that renders tissues optically transparent, allowed visualization deeper inside spheroids. About 16 h after irradiation, a red fluorescent cell fraction, presumably a quiescent G0 cell fraction, became distinct from the outer fraction consisting of proliferating cells, most of which exhibited green fluorescence indicative of G2 arrest. Thereafter, the red cell fraction began to emit green fluorescence and remained in prolonged G2 arrest. Thus, for the first time, we visualized the prolongation of radiation-induced G2 arrest in spheroids and the differences in cell kinetics between the outer and inner fractions.
    Biochemical and Biophysical Research Communications 09/2013; · 2.28 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Abstract Conclusions: The findings of this study demonstrated that the wait-and-watch strategy for neck metastasis from squamous cell carcinoma (SCC) of oral tongue is a reliable option and that salvage by surgical treatment is effective. However, younger patients should be closely monitored for recurrence. Adjuvant therapy may be recommended for patients with pathologically advanced disease. Objectives: Metastatic involvement of cervical lymph nodes is the most important prognostic indicator in patients with oral tongue SCC. With the objective of determining the most appropriate treatment strategy for regional recurrence, we conducted a retrospective review of clinicopathologic factors. Methods: The clinicopathologic features of 103 patients with oral tongue SCC, in whom the local lesions were treated successfully by low-dose interstitial brachytherapy (LD-IBT), but who subsequently developed cervical lymph node metastases and were treated by salvage surgery, were reviewed. Results: In the patients who underwent surgical treatment at our hospital, 5-year disease-free survival and regional control rates were 69.3% and 85.3%, respectively. The clinicopathologic factors significantly associated with unfavorable disease-free survival were the presence of extracapsular spread (hazard ratio (HR) = 3.005, p = 0.045), multiple and large lymph nodes (HR = 2.850, p = 0.010 and HR = 3.112, p = 0.007, respectively), younger age (HR = 2.429, p = 0.048), and shorter interval from the LD-IBT to detection of neck metastasis (HR = 1.749, p = 0.013).
    Acta oto-laryngologica 01/2013; · 0.98 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Plinabulin (NPI-2358) is a novel microtubule-depolymerizing agent. In HeLa cells, plinabulin arrests the cell-cycle at M phase and subsequently induces mitotic catastrophe. To better understand the effects on this compound on the cell-cycle, we used the fluorescent ubiquitination-based cell cycle indicator (Fucci), which normally enables G1 and S/G2/M cells to emit red and green fluorescence, respectively. When HeLa-Fucci cells were treated with 50nM plinabulin, cells began to fluorescence both green and red in an unusual pattern; most cells exhibited the new pattern after 24h of treatment. X-irradiation efficiently induced G2 arrest in plinabulin-treated cells and significantly retarded the emergence of the unusual pattern, suggesting that entering M phase is essential for induction of the pattern. By simultaneously visualizing chromosomes with GFP-histone H2B, we established that the pattern emerges after nuclear envelope breakdown but before metaphase. Pedigree assay revealed a significant relationship between the unusual expression and mitotic catastrophe. Nocodazole, KPU-133 (a more potent derivative of plinabulin), and paclitaxel also exerted similar effects. From these data, we conclude that the unusual pattern may be associated with dysregulation of late M phase-specific E3 ligase activity and mitotic catastrophe following treatment with anti-microtubule agents.
    Biochemical and Biophysical Research Communications 10/2012; · 2.28 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Recent reports have described that NCSCs (neural crest-derived stem cells) are not only present in the embryonic neural crest but also in the adult tissues. Dental pulp is one of mesenchymal soft tissues origin from cranial neural crest cells, and thought to be a source of adult stem cells. Here, we investigated the existence of NCSC-like cells in apical pulp of human developing tooth. Human impacted third molars with immature apex freshly extracted were obtained. The cells derived from the apical pulp tissue not framed by dentin or the coronal pulp tissues were cultured by primary explant culture. APDCs (apical pulp-derived cells) and CPCs (coronal pulp cells) formed spheres under neurosphere culture condition. The number of spheres from APDCs was larger than that from CPCs. The sphere-forming cells derived from APDCs had self-renewal capacity, and expressed neural crest-associated markers (p75, Snail and Slug) and NSC (neural stem cell) markers (Nestin and Musashi1). The expression pattern of mesenchymal stem cell markers, CD105 and CD166, on the surface of sphere-forming cells derived APDCs was different from that of APDCs. These sphere-forming cells could differentiate into multiple mesenchymal lineages (osteoblasts, adipocytes, chondrocytes and smooth muscle cells) and neural lineage (neurons) in vitro, and generated ectopic bone tissues on the border of HA (hydroxyapatite) scaffold in vivo. The results of this study suggest that APDCs contain cells with characteristics of NCSCs reported previously in mice. Humans developing tooth with immature apex is an effective source of cells for neural crest lineage tissue regeneration.
    Cell Biology International 06/2012; 36(10):927-36. · 1.64 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Perfluorooctane sulfonate (PFOS) is a pollutant widely found throughout nature and is toxic to animals. We created a PFOS analogue on a polyethylene glycol polyacrylamide copolymer and isolated peptides that preferentially bound the PFOS analogue using a T7 phage display system. Bioinformatic analysis using the FASTAskan program on the RELIC bioinformatics server showed several human proteins that likely bound PFOS. Among them, we confirmed binding between PFOS and a recombinant soluble form of monocyte differentiation antigen CD14 (sCD14) by a surface plasmon biosensor. Furthermore, PFOS inhibited TNF-α production induced by the sCD14 in mouse macrophage RAW264.7 cells.
    Bioorganic & medicinal chemistry 05/2012; 20(13):3985-90. · 2.82 Impact Factor
  • Atsushi Kaida, Masahiko Miura
    [Show abstract] [Hide abstract]
    ABSTRACT: Although oxygen is required for functional chromophore formation during the maturation process of fluorescent proteins, the effects of hypoxia on their fluorescence have rarely been studied in mammalian cells. We recently reported that severe hypoxia (pO(2)<0.1%) abrogates fluorescence from the fluorescent ubiquitination-based cell cycle indicator (Fucci) expressed in HeLa cells. Fucci is a system for visualizing cell cycle progression in live cells using red (monomeric Kusabira Orange 2, mKO(2)) and green (monomeric Azami Green, mAG) fluorescent proteins. In this study, taking advantage of the system, we attempted to determine the dependence on oxygen tension (pO(2)) of these two fluorescent proteins during the maturation process. The oxygen tension at which the number of fluorescence-positive cells was reduced by 50% (pO(2)·50) was 0.9% and 0.3% for mKO2 and mAG, respectively. Furthermore, we measured fluorescence recovery kinetics after reoxygenation in cells treated at two different pO(2) levels, and observed that mKO2 exhibits slower kinetics of oxidation than mAG. Thus, we demonstrate that mKO2 exhibits a stronger dependence on oxygen tension than mAG, as well as the usefulness of this novel method to produce varying levels of hypoxic conditions.
    Biochemical and Biophysical Research Communications 04/2012; 421(4):855-9. · 2.28 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: To analyze data from patients receiving repeat brachytherapy (re-BT) for the treatment of residual or recurrent tumor in the oral cavity. Between January 2003 and December 2007, 62 patients who had undergone definitive BT as an initial treatment of oral cancer subsequently underwent re-BT for the treatment of residual or recurrent tumors at the diagnostic radiology and oncology department (Tokyo Medical and Dental University Hospital). Re-BT was performed 0.9-73 months (median, 5.7) after the initial BT. Au-198 grains were used as the re-BT source in all 62 patients, and an area of 0.8-6.3 cm(2) (median, 3.1) was permanently irradiated with 60-110 Gy (median, 83) according to the system of Paterson-Parker. The 2-year local control and overall survival rate was 53% and 66%, respectively, and local control significantly affected overall survival. Both local control and overall survival were affected by the initial tumor characteristics and the macroscopic appearance of the residual or recurrent tumor. Grade 3 or 4 complications were seen in 5 patients. The incidence of mandibular and mucosal complications was significantly related to a biologic effective dose of α/β of 3 Gy to the surface of the gingiva and mucosa, respectively. Re-BT using Au-198 grains for the treatment of residual or recurrent tumor after definitive BT in the oral cavity is effective and well tolerated.
    International journal of radiation oncology, biology, physics 11/2011; 83(4):1198-204. · 4.59 Impact Factor
  • Atsushi Kaida, Masahiko Miura
    [Show abstract] [Hide abstract]
    ABSTRACT: Fluorescent proteins are widely used for the direct visualization of events such as gene expression and subcellular localization in mammalian cells. It is well established that oxygen is required for formation of functional chromophore; however, the effect of hypoxia on fluorescence emission has rarely been studied. For this purpose, under hypoxic conditions, we investigated the kinetics of red and green fluorescence in HeLa cells from two fluorescent proteins, monomeric Kusabira Orange 2 (mKO2) and monomeric Azami Green (mAG), respectively, using the fluorescent ubiquitination-based cell cycle indicator (Fucci). In this system, cells in G1 or other phases emit red or green fluorescence, respectively. We found that hypoxia abrogated both red and green fluorescence about ~10h after the treatment, although their protein levels were almost maintained. The treatment did not significantly affect fluorescence in cells constitutively expressing the same fluorescent proteins lacking the ubiquitin ligase-binding domains. The abrogation of fluorescence resulted from a combination of ubiquitination-dependent degradation of pre-existing functional proteins during specific cell cycle phases, and the expression of newly synthesized non-fluorescent proteins containing non-oxidized chromophore during hypoxic treatment. Indeed, non-fluorescent cells after hypoxic treatment gradually developed fluorescence after reoxygenation in the presence of cycloheximide; kinetics of recovery were much faster for mAG than for mKO2. Using the Fucci system, we could clearly visualize for the first time the effect of hypoxia on the fluorescence kinetics of proteins expressed in living mammalian cells.
    Experimental Cell Research 11/2011; 318(3):288-97. · 3.56 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: The mitotic shake-off method revealed the remarkable variation of radiosensitivity of HeLa cells during the cell cycle: M phase shows the greatest radiosensitivity and late S phase the greatest radioresistance. This method harvests all M-phase cells with a round shape, making it impossible to further subdivide M-phase cells. Recently, the fluorescent ubiquitination-based cell cycle indicator (Fucci) was developed; this system basically causes cells in G(1) to emit red fluorescence and other cells to emit green fluorescence. Because the green fluorescence rapidly disappears at late M phase, two-dimensional flow cytometry analysis can usually detect a green(high)/red(low) fraction including S-, G(2)- and early M-phase cells but not a transitional fraction between green(high)/red(low) and green(low)/red(low) including late M-phase cells. However, combining the shake-off method concentrated the transitional fraction, which enabled us to separate early and late M-phase cells without using any drugs. Here we demonstrate for the first time that cells in early M phase are more radiosensitive than those in late M phase, implying that early M phase is the most radiosensitive sub-phase during the cell cycle.
    Radiation Research 06/2011; 176(3):407-11. · 2.70 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The purpose of this study was to characterize the radiobiological properties of stem/progenitor cells derived from apical papilla-derived cells (APDCs) compared to bulk APDCs. APDCs were isolated from freshly extracted human third molars with immature apices. Multipotent spheres, which are thought to contain an enriched population of stem/progenitor cells, were formed from the APDCs, using a neurosphere culture technique. After γ-irradiation, papillary sphere-forming cells (PSFCs) and bulk APDCs were subjected to radiosensitivity and hard tissue-forming assays. Compared to bulk APDCs, the PSFCs exhibited a radioresistant phenotype and a higher capacity for DNA double strand break repair. Irradiation induced a significant increase in a senescence-like phenotype in both cell types. Neither type of cells exhibited a significant induction of apoptotic changes after 8 Gy of irradiation. Ability to form hard tissue in vivo was significantly decreased in PSFCs, but not in APDCs following 4 Gy of irradiation. We demonstrated for the first time that stem/progenitor cells derived from APDCs exhibit a radioresistant phenotype; however, the hard tissue forming ability in vivo, but not bulk APDCs, was significantly reduced after irradiation.
    Stem Cell Research & Therapy 01/2011; 2(1):2. · 3.65 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Fucci (fluorescent ubiquitination-based cell cycle indicator) is able to visualize dynamics of cell cycle progression in live cells; G1- and S-/G2-/M-phase cells expressing Fucci emit red and green fluorescence, respectively. This system could be applied to cell kinetic analysis of tumour cells in the field of cancer therapy; however, it is still unclear how fluorescence kinetics change after various treatments, including exposure to anticancer agents. To explore this, we arrested live HeLa cells expressing the Fucci probes at various cell cycle stages and observed the fluorescence, in conjunction with flow cytometric analysis. X-irradiation, HU (hydroxyurea) and nocodazole arrest cells at G2/M boundary, early S-phase and early M-phase, respectively. Although X-irradiation and HU treatment induced similar accumulation kinetics of green fluorescent cells, nocodazole treatment induced an abnormal red fluorescence at M phase, followed by accumulation of both red and green fluorescent cells with 4N DNA content. We conclude that certain agents that disrupt normal cell cycle regulation could cause unexpected fluorescence kinetics in the Fucci system.
    Cell Biology International 01/2011; 35(4):359-63. · 1.64 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The aim of this study was to evaluate the outcome and complications of low-dose-rate brachytherapy (LDR-BT) for oral cancer according to comorbidity. The records of a total of 180 patients who received LDR-BT for T1-2N0M0 oral cancers between January 2005 and December 2007 were analysed. The comorbidities of the patients were retrospectively graded according to the Adult Comorbidity Evaluation-27, and the relationships between the comorbidity grades and survival, disease control and the incidence of complications were analysed. The 2 year overall survival rates of patients with no comorbidity, Grade 1, Grade 2 and Grade 3 comorbidity were 87%, 85%, 76% and 65%, respectively, and the reduction in the survival rate according to comorbid severity was significant in a univariate analysis (p = 0.032) but not in a multivariate analysis including other clinical factors. Cause-specific survival, locoregional control and local control were not related to the comorbidity grade, or any other clinical factors. Grade 2 or 3 complications developed in 27% of the patients. The incidence of complications was unrelated to the comorbidity grade. The disease control of oral cancer and the incidence of complications after LDR-BT were not related to comorbid severity. LDR-BT is a useful and safe treatment for patients regardless of the presence of severe comorbidity.
    The British journal of radiology 01/2011; 84(1006):930-8. · 2.11 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: The purpose of this study was to examine the effects of combined treatment with a specific type of sulfoglycolipid, α-sulfoquinovosylmonoacylglycerol (α-SQMG), and X-irradiation (XRT) on the remodeling of tumor microenvironments. A human colon cancer cell line, SW480, was used in this study. The cells were injected subcutaneously into nude mice and the resulting tumors were treated with α-SQMG that was injected intravenously and/or X-irradiation (XRT). The tumor volumes were monitored and the microenvironments of the treated tumors were immunohistochemically analyzed for angiogenesis, pericyte recruitment, and hypoxic fractions using markers for CD31, collagen IV, α-Sma, and pimonidazole. The combined treatment with α-SQMG (five daily injections from days zero to four) and X-irradiation (two fractions on days zero and three) synergistically enhanced the radioresponse of tumor growth in vivo, whereas α-SQMG treatment alone had no effect. The tumor vessel density was significantly decreased at days 10 and 20 after initiating the combined treatment. On day 20, areas with overlapping CD31 and collagen IV expression were rarely observed, suggesting that the normal structures of most tumor vessels had collapsed. α-Sma staining was significantly increased and pimonidazole staining was significantly reduced at 24 and 72 h, but not 6 h, after the first combined treatment. The combined treatment induced remodeling of the microenvironments in SW480 tumors, which might contribute to the radiosensitization to the second irradiation.
    Anticancer research 11/2010; 30(11):4397-404. · 1.71 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: The effects of ionizing radiation (IR) on tumor neovascularization are still unclear. We previously reported that vascular endothelial cells (ECs) expressing the IR-induced senescence-like (IRSL) phenotype exhibit a significant decrease in angiogenic activity in vitro. In this study, we examined the effects of the IRSL phenotype on adhesion to early endothelial progenitor cells (early EPCs). Adhesion of human peripheral blood-derived early EPCs to human umbilical vein endothelial cells (HUVECs) expressing the IRSL phenotype was evaluated by an adhesion assay under static conditions. It was revealed that the IRSL HUVECs supported significantly more adhesion of early EPCs than normal HUVECs. Expressions of ICAM-1, VCAM-1 and E-selectin were up-regulated in IRSL HUVECs. Pre-treatment of IRSL HUVECs with adhesion-blocking monoclonal antibodies against E-selectin and VCAM-1 significantly reduced early EPC adhesion to IRSL HUVECs, suggesting a potential role for the E-selectin and VCAM-1 in the adhesion between IRSL ECs and early EPCs. Therefore, the IRSL phenotype expressed in ECs may enhance neovascularization via increased homing of early EPCs. Our findings are first to implicate the complex effects of this phenotype on tumor neovascularization following irradiation.
    Journal of Radiation Research 08/2009; 50(5):469-75. · 1.45 Impact Factor
  • Fuel and Energy Abstracts 01/2009; 75(3).
  • Kaori Igarashi, Masahiko Miura
    [Show abstract] [Hide abstract]
    ABSTRACT: The well-established process of potentially lethal damage (PLD) repair enhances plateau-phase cell survival after exposure to ionizing radiation. PLD repair requires that confluent cells be incubated prior to plating for a colony-forming assay rather than being plated immediately. Enhanced double-strand break (DSB) repair during this incubation period has been implicated in the enhanced survival, but the precise molecular mechanism and its biological significance remain largely unclear. Radiation has been recently reported to induce premature senescence, and increasing evidence suggests that DSBs commonly mediate cellular senescence. Here we successfully related these two biological phenomena using bovine aortic endothelial cells, and propose that enhanced DSB repair during the plateau-phase incubation prevents expression of the radiation-induced senescence-like phenotype, eventually leading to an enhanced colony-forming ability. This could be a novel biological interpretation of PLD repair.
    Radiation Research 11/2008; 170(4):534-9. · 2.70 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: To assess the quality of life (QOL) of oral cancer patients treated with low-dose-rate interstitial brachytherapy (LDR-BT) alone. Between June 2005 and July 2006, a total of 56 patients with oral cancer were enrolled in this prospective study. QOL was assessed by means of the core questionnaire and head and neck questionnaire module of the European Organization for Research and Treatment of Cancer (EORTC Quality of Life Questionnaire-Core 30 [QLQ-C30] and QLQ Head and Neck 35 [H&N35]). The questionnaires were distributed to the patients before the start of treatment and 3 months, 6 months, and 12 months after the start of LDR-BT. It was possible to analyze the results for 20 of the initial 56 patients because they did not experience metastasis or recurrence during this study. No functions or symptoms asked about in the QLQ-C30 deteriorated during the first year. The emotional function score steadily and significantly increased. No symptoms in the QLQ-H&N35 significantly deteriorated. The scores for pain, trouble with social eating, and weight loss on the QLQ-H&N35 steadily and significantly decreased. Age, gender, and LDR-BT source had no effect on the change in QOL during the first year, but T-stage significantly affected the change in global health status, tumor site affected the changes in swallowing, sensory problems, sticky saliva, and complications affected the changes in pain, swallowing, and mouth opening. QOL of oral cancer patients treated with LDR-BT is high. However, tumor stage, tumor site, and complications affected the changes in a few functions and symptoms during the first year.
    International journal of radiation oncology, biology, physics 08/2008; 73(3):772-8. · 4.59 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: We previously reported that 3'-sulfoquinovosyl-1'-monoacylglycerol (SQMG) was effective in suppressing the growth of solid tumors due to hemorrhagic necrosis in vivo. In the present study, we investigated the antiangiogenic effect of SQMG. In vivo assessment of antitumor assays showed that some tumor cell lines, but not others, were sensitive to SQMG. Microscopic study suggested that in SQMG-sensitive tumors, but not SQMG-resistant tumors, angiogenesis was reduced. We next investigated gene expression relating to angiogenesis in tumor tissues by quantitative real-time polymerase chain reaction. Consequently, although vascular endothelial growth factor gene expression was not detected with significant differences among the cases, significant downregulation of Tie2 gene expression was observed in all SQMG-sensitive tumors as compared with controls, but not in SQMG-resistant tumors. These data suggested that the antitumor effects of SQMG could be attributed to antiangiogenic effects, possibly via the downregulation of Tie2 gene expression in SQMG-sensitive tumors.
    Cancer Science 06/2008; 99(5):1063-70. · 3.48 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The management of lymphatic metastasis of early-stage oral tongue carcinoma patients is crucial for its prognosis. The purpose of this study was to evaluate the predictive ability of lymphatic metastasis after brachytherapy (BRT) for early-stage tongue carcinoma based on gene expression profiling. Pre-therapeutic biopsies from 39 patients with T1 or T2 tongue cancer were analyzed for gene expression signatures using Codelink Uniset Human 20K Bioarray. All patients were treated with low dose-rate BRT for their primary lesions and underwent strict follow-up under a wait-and-see policy for cervical lymphatic metastasis. Candidate genes were selected for predicting lymph-node status in the reference group by the permutation test. Predictive accuracy was further evaluated by the prediction strength (PS) scoring system using an independent validation group. We selected a set of 19 genes whose expression differed significantly between classes with or without lymphatic metastasis in the reference group. The lymph-node status in the validation group was predicted by the PS scoring system with an accuracy of 76%. Gene expression profiling using 19 genes in primary tumor tissues may allow prediction of lymphatic metastasis after BRT for early-stage oral tongue carcinoma.
    Radiotherapy and Oncology 06/2008; 87(2):237-42. · 4.52 Impact Factor

Publication Stats

488 Citations
163.86 Total Impact Points

Institutions

  • 2006–2012
    • Tokyo University of Science
      • Department of Applied Biological Science
      Tokyo, Tokyo-to, Japan
    • Asahi General Hospital
      Asahi, Chiba, Japan
  • 1992–2012
    • Tokyo Medical and Dental University
      • • Department of Oral Restitution
      • • Department of Diagnostic Radiology and Oncology
      • • Department of Radiology
      • • Department of Dental Radiology and Radiation Research
      Edo, Tōkyō, Japan
  • 2001
    • National Institute of Radiological Sciences
      Tiba, Chiba, Japan