Involvement Of BCL2L10 Protein In Regulation Of Apoptosis And Proliferation In Human Gastric Carcinoma And Implicated Mechanisms

IntroductionThough it is no longer the second most common cancer worldwide, gastric cancer is still the second most common cause of death from cancer. The geographical distribution of gastric cancer is characterized by wide international variations. High-risk areas (age-standardized rate in men> 20 per 100,000) include East Asia (China, Japan), Eastern Europe, and parts of Central and South America. It is a relatively uncommon neoplasm in North America, but gastric cancer is the third most lethal neoplasm overall in North America, contributing substantially to the burden of cancer deaths.BCL2L10 gene is located on human chromesome 15q12.2 and expresses 204 amino acids protein, about 23kDa. BDL2L10 protein is strongly expressed in all normal plasma cells but found in only 18% of multiple myelomas. BCL2L10 immunostaining is also present in approximately half of diffuse large B-cell lymphoma specimens, whereas follicular lymphomas do not contain BCL2L10. In normal adult human stomach tissues, BCL2L10 mRNA is expressed at high levels (three-plus). However, no significant association with survival was observed in gastric cancer patients.BCL2L10 is the last protein member found in Bcl-2 family. There are at least 20 Bcl-2-related proteins in the life or death decision of mammalian cells. Intriguingly, in addition to its central role in regulating apoptosis, the Bcl-2 family influences the cell cycle or more specifically, the transit between quiescence and proliferation, which, together with apoptosis regulation, makes its important role in oncogenesis. However, no such research on BCL2L10 regulation on gastric cancer cells.Using three experimental models - retrospective clinical specimens of human gastric carcinoma, gastric cancer cell lines, and mouse xenograft model - our study investigated impact of BCL2L10 on gastric cancer cells.PartⅠLoss of BCL2L10 Protein Expression as Prognostic Predictor for Poor Clinical Outcome in Gastric CarcinomaAims: BCL2L10 protein is an apoptosis-related member of the Bcl-2 protein family. The clinical significance of its expression in gastric carcinoma is poorly understood. Our aim was to investigate BCL2L10 expression and its clinical and prognostic significance in gastric carcinoma patients.Methods and results: Immunohistochemistry revealed extensive loss of BCL2L10 expression in gastric cancer cells. The scaled BCL2L10 expression data was categorized into 3 groups (groups 0-2) to facilitate statistical analysis. We observed a significant correlation between the lower BCL2L10 expression group and shorter disease-free survival (P= 1.956×10-18). Multivariate regression analysis showed that loss of BCL2L10 protein expression (P= 4.883×10-8, HR= 0.252) is an independent prognostic predictor of gastric carcinoma. The receiver operator characteristic (ROC) curve showed that the area for BCL2L10 protein was 0.817 (P= 8.331×10-14), indicating that loss of BCL2L10 protein expression is an excellent prognostic predictor of gastric carcinoma. We re-investigated BCL2L10 expression by quantitative real time PCR, immunoblotting and examined methylation status of BCL2L10 gene promoter by bisulfite sequencing in fresh gastric normal and carcinoma tissues. We observed a significant correlation of lower BCL2L10 expression with hypermethylated CpG island of gene promoter in gastric carcinoma.Conclusions: To our knowledge, it is the first report that loss of BCL2L10 protein expression predicts poor clinical outcome in gastric carcinoma. Hypermethylated gene promoter is responsible for loss of BCL2L10 in gastric carcinoma. Part II BCL2L10 Protein Regulates Apoptosis/Proliferation through Differential Pathways in Gastric Cancer CellsAims: The reason and consequence of BCL2L10 downregulation in gastric carcinoma is poorly understood. Our aim was to investigate the function of BCL2L10 protein in gastric carcinoma.Methods and results: We studied its apoptosis and proliferation regulation in gastric cancer cell lines by flow cytometry, fluorescence staining, murine xenograft model and immunoblotting. Pathway inhibitors were also applied to confirm the major pathway involved in apoptosis or proliferation regulation. We observed apoptosis induced by upregualted BCL2L10 through the mitochondrial pathway and proliferation accelerated by BCL2L10 siRNA via PI3K/Akt signaling pathway in gastric cancer cell lines.Conclusions: The pro-apoptotic effect of BCL2L10 and growth promotion by BCL2L10 siRNA in gastric cancer cells might suggest a tumor suppressor of BCL2L10.