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Role of mammalian target of rapamycin signalling in bone destruction by gingival squamous cell carcinoma
428 Free Papers—Oral Presentations
O1.9 Co-expression of laminin-5 gamma2 and p53 in oral squamous cell carcinoma H. Suzuki ∗ , K. Hashimoto Department of Oral and Maxillofacial Surgery, Hamamatsu University School of Medicine, Hamamatsu, Japan
Background and Objectives: The identification of a specific factor for predicting the clinical outcome of an oral cancer would be helpful for selecting more effective treatments. In this research, we evaluated the relationship between the prognosis and the co-expression of laminin-5 gamma2/p53. Laminin is the most abundant glycoprotein in basement membrane. It is an 820-kD cross-shaped heterotrimer that connects cells to underlining extracellular matrix components that plays a key role of cell survival, proliferation, differentiation and motility. p53 gene is one of the most mutated genes in human cancers. It can exert antiproliferative effects, but equally important, it regulates apoptosis. Methods: The study group comprised 48 patients (28 men, 20 women) with oral squamous cell carcinoma (OSCC) who underwent chemotherapy and surgery at our hospital. Biopsy specimens were used for all experiments. The immunohistochemical studies were evaluated as follows: 1) tissues were scored at 20 magnification under light microscopy. Five available areas in the section were evaluated; 2) measuring the positive areas using Image-Pro Plus (Media Cybernetics, Ink.) soft ware program; and 3) localisation: we divided into 2 types. Results: We can observe the tumour cells at the cancer-stromal interface frequently expressed immunopositivity for the laminin-5 gamma2 and p53 in OSCC. The analysis of the patient’s survival showed that the prognosis of diffuse type of laminin-5 gamma2 and p53 cases are significantly poorer than marginal type ones (P < 0.05). Conclusion: The laminin-5 gamma2 and p53 localisation pattern may be a marker for prognosis of oral squamous cell carcinomas. doi:10.1016/j.ijom.2009.03.113
O1.10 S-1 mediates the inhibition of lymph node metastasis in oral cancer cells H. Sato ∗ , M. Hatori, Y. Kurihara, T. Shirota, S. Shintani
Department of Oral and Maxillofacial Surgery, School of Dentistry, Showa University, Ohta-ku, Tokyo, Japan
Background and Objectives: We examined the effect of S-1 on regional lymph node metastasis using the green fluorescence protein (GFP)-SAS-L1 lymph node metastasis model. We also performed in vitro experiments to determine the effects of S-1 on oral squamous cell carcinoma (OSCC) invasion and metastasis. Methods: First, we injected GFP-SASL1 into the tongues of nude mice. Three days after the injection, S-1 (20 mg/kg) was orally administered 5 times per week for 2 weeks. Autopsy was performed 2 weeks after injection. The status of the tumour and the lymph node metastases were examined by GFP fluorescence. To explore the mechanism of down-regulation of OSCC metastases, we performed cell adhesion assay and western blotting analysis for the expression of integrins. We also examined the phosphorylation of FAK. Results: S-1 significantly reduced tumour volume and inhibited lymph node metastasis when compared with non-treated mice. GFP-SAS-L1 cells treated with 5-FU and CDHP showed significantly lower invasiveness than control cells. Additionally, GFP-SAS-L1 cells treated with 5-FU and CDHP showed the reduction of adhesion to laminin and downregulation of integrin alpha-3, alpha-6, alpha-v, beta-1, beta-3, beta-4, and FAK phosphorylation. Conclusions: The results of this study confirm that systemic administration of S-1 inhibits metastasis of human OSCC cells grafted in the tongues of athymic nude mice. Furthermore, the selective downregulation of integrin expression and FAK phosphorylation by S-1 might lead to reduce cell adhesion to the extracellular matrix. These results suggested that the S-1 inhibited the metastases of OSCC by regulation of cell adhesion molecules. doi:10.1016/j.ijom.2009.03.114
O1.11 Role of mammalian target of rapamycin signalling in bone destruction by gingival squamous cell carcinoma T. Okui ∗ , T. Shimo, N. Kurio, M. Takaoka, N. Yoshioka, N.M.M. Hassan, Y. Naomoto, A. Sasaki Department of Oral and Maxillofacial Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan
Background and Objectives: Gingival squamous cell carcinomas (SCC) frequently invade the mandible or maxilla, and this invasion is associated with a worse prognosis. Transforming growth factor beta (TGF-beta) and receptor activator of nuclear factor (NF)-kappaB ligand (RANKL) are critical factors of osteoclast formation and function. In this study, we focused on mammalian target of rapamycin (mTOR) downstream of TGF-beta and RANKL, and investigated the role of mTOR signalling on SCC induced bone destruction. Methods: Bone-resorbing activity was measured by calcium release from the calvaria contact with human oral SCC cell line HSC-2 cells in type I collagen gel. To evaluate the effects of temsirolimus on the HSC-2 cells function, [3H] thymidine incorporation assay and migration assay were done. Osteoclast formation and bone resorption were evaluated by tartrateresistant acid phosphatase (TRAP) staining and pit formation assay. Results: Temsirolimus suppressed the proliferation and migration of HSC-2 cells. Temsirolimus also decreased HSC-2 cells induced calcium release from the calvaria. HSC-2 culture medium stimulated osteoclast formation, whereas temsirolimus suppressed the osteoclast formation in mouse bone marrow cells. Next we confirmed that TGF-beta and RANKL activated Akt, IkB and mTOR signalling in murine pre-osteoclastic RAW264.7 cells, and this activation was inhibited by temsirolimus. Treatment of temsirolimus inhibited both TGF-beta and RANKL induced osteoclasts formation and activity. Conclusion: mTOR signalling may be a new molecular target for anti-osteolytic therapy to shut off the TGF-beta and RANKL signalling pathway. doi:10.1016/j.ijom.2009.03.115
O1.12 Analysis of p53, c-Myc and c-Erb B2 gene in histopathologically tumour-free surgical margins in patients with oral squamous cell carcinoma D. Jelovac ∗ , V. Konstantinovic, B. Ilic, B. Nesic, M. Manasijevic, B. Popovic, J. Milasin Clinic of Maxillofacial Surgery, Faculty of Dentisty, University of Belgrade, Gandijeva, Belgrade, Serbia
Background and Objectives: In total, 10–30% of patients with oral squamous cell carcinoma (OSCC) develop local
O1.9 Co-expression of laminin-5 gamma2 and p53 in oral squamous cell carcinoma H. Suzuki ∗ , K. Hashimoto Department of Oral and Maxillofacial Surgery, Hamamatsu University School of Medicine, Hamamatsu, Japan
Background and Objectives: The identification of a specific factor for predicting the clinical outcome of an oral cancer would be helpful for selecting more effective treatments. In this research, we evaluated the relationship between the prognosis and the co-expression of laminin-5 gamma2/p53. Laminin is the most abundant glycoprotein in basement membrane. It is an 820-kD cross-shaped heterotrimer that connects cells to underlining extracellular matrix components that plays a key role of cell survival, proliferation, differentiation and motility. p53 gene is one of the most mutated genes in human cancers. It can exert antiproliferative effects, but equally important, it regulates apoptosis. Methods: The study group comprised 48 patients (28 men, 20 women) with oral squamous cell carcinoma (OSCC) who underwent chemotherapy and surgery at our hospital. Biopsy specimens were used for all experiments. The immunohistochemical studies were evaluated as follows: 1) tissues were scored at 20 magnification under light microscopy. Five available areas in the section were evaluated; 2) measuring the positive areas using Image-Pro Plus (Media Cybernetics, Ink.) soft ware program; and 3) localisation: we divided into 2 types. Results: We can observe the tumour cells at the cancer-stromal interface frequently expressed immunopositivity for the laminin-5 gamma2 and p53 in OSCC. The analysis of the patient’s survival showed that the prognosis of diffuse type of laminin-5 gamma2 and p53 cases are significantly poorer than marginal type ones (P < 0.05). Conclusion: The laminin-5 gamma2 and p53 localisation pattern may be a marker for prognosis of oral squamous cell carcinomas. doi:10.1016/j.ijom.2009.03.113
O1.10 S-1 mediates the inhibition of lymph node metastasis in oral cancer cells H. Sato ∗ , M. Hatori, Y. Kurihara, T. Shirota, S. Shintani
Department of Oral and Maxillofacial Surgery, School of Dentistry, Showa University, Ohta-ku, Tokyo, Japan
Background and Objectives: We examined the effect of S-1 on regional lymph node metastasis using the green fluorescence protein (GFP)-SAS-L1 lymph node metastasis model. We also performed in vitro experiments to determine the effects of S-1 on oral squamous cell carcinoma (OSCC) invasion and metastasis. Methods: First, we injected GFP-SASL1 into the tongues of nude mice. Three days after the injection, S-1 (20 mg/kg) was orally administered 5 times per week for 2 weeks. Autopsy was performed 2 weeks after injection. The status of the tumour and the lymph node metastases were examined by GFP fluorescence. To explore the mechanism of down-regulation of OSCC metastases, we performed cell adhesion assay and western blotting analysis for the expression of integrins. We also examined the phosphorylation of FAK. Results: S-1 significantly reduced tumour volume and inhibited lymph node metastasis when compared with non-treated mice. GFP-SAS-L1 cells treated with 5-FU and CDHP showed significantly lower invasiveness than control cells. Additionally, GFP-SAS-L1 cells treated with 5-FU and CDHP showed the reduction of adhesion to laminin and downregulation of integrin alpha-3, alpha-6, alpha-v, beta-1, beta-3, beta-4, and FAK phosphorylation. Conclusions: The results of this study confirm that systemic administration of S-1 inhibits metastasis of human OSCC cells grafted in the tongues of athymic nude mice. Furthermore, the selective downregulation of integrin expression and FAK phosphorylation by S-1 might lead to reduce cell adhesion to the extracellular matrix. These results suggested that the S-1 inhibited the metastases of OSCC by regulation of cell adhesion molecules. doi:10.1016/j.ijom.2009.03.114
O1.11 Role of mammalian target of rapamycin signalling in bone destruction by gingival squamous cell carcinoma T. Okui ∗ , T. Shimo, N. Kurio, M. Takaoka, N. Yoshioka, N.M.M. Hassan, Y. Naomoto, A. Sasaki Department of Oral and Maxillofacial Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan
Background and Objectives: Gingival squamous cell carcinomas (SCC) frequently invade the mandible or maxilla, and this invasion is associated with a worse prognosis. Transforming growth factor beta (TGF-beta) and receptor activator of nuclear factor (NF)-kappaB ligand (RANKL) are critical factors of osteoclast formation and function. In this study, we focused on mammalian target of rapamycin (mTOR) downstream of TGF-beta and RANKL, and investigated the role of mTOR signalling on SCC induced bone destruction. Methods: Bone-resorbing activity was measured by calcium release from the calvaria contact with human oral SCC cell line HSC-2 cells in type I collagen gel. To evaluate the effects of temsirolimus on the HSC-2 cells function, [3H] thymidine incorporation assay and migration assay were done. Osteoclast formation and bone resorption were evaluated by tartrateresistant acid phosphatase (TRAP) staining and pit formation assay. Results: Temsirolimus suppressed the proliferation and migration of HSC-2 cells. Temsirolimus also decreased HSC-2 cells induced calcium release from the calvaria. HSC-2 culture medium stimulated osteoclast formation, whereas temsirolimus suppressed the osteoclast formation in mouse bone marrow cells. Next we confirmed that TGF-beta and RANKL activated Akt, IkB and mTOR signalling in murine pre-osteoclastic RAW264.7 cells, and this activation was inhibited by temsirolimus. Treatment of temsirolimus inhibited both TGF-beta and RANKL induced osteoclasts formation and activity. Conclusion: mTOR signalling may be a new molecular target for anti-osteolytic therapy to shut off the TGF-beta and RANKL signalling pathway. doi:10.1016/j.ijom.2009.03.115
O1.12 Analysis of p53, c-Myc and c-Erb B2 gene in histopathologically tumour-free surgical margins in patients with oral squamous cell carcinoma D. Jelovac ∗ , V. Konstantinovic, B. Ilic, B. Nesic, M. Manasijevic, B. Popovic, J. Milasin Clinic of Maxillofacial Surgery, Faculty of Dentisty, University of Belgrade, Gandijeva, Belgrade, Serbia
Background and Objectives: In total, 10–30% of patients with oral squamous cell carcinoma (OSCC) develop local