MicroRNA-125b Inhibits Apoptosis By Down-regulating The Expression Of BAK1 In Pancreatic Ductal Adenocarcinoma

Background: Pancreatic cancer is one of the most malignant tumors, the overall 5-year survival rate is no more than 5%. And it can easily invasive and metastatic but has no relative symptoms or signs in early stage. Pancreatic cancer also has the poorest prognosis because of its late discovery and resistance to chemotherapy and radiation therapy. Micro RNA-125 b is a tumor-associated mi RNA which plays an important role in a variety of tumors development, but little is known in pancreatic cancer and the mechanism is not clear.Objective: To explore the function and possible mechanisms of micro RNA-125 b in pancreatic cancer.Methods: By bioinformatics Target Scan, we found the potential target BAK1 which had the most relationship with micro RNA-125 b, and Dual-Lusiferase reporter assay was used to confirm the relationship. Quantitative RT-PCR was used to measure the expression of micro RNA-125 b in pancreatic cancer cell lines, pancreatic cancer and adjacent normal tissues, as well as in normal pancreatic tissues. Western blotting was used to measure the expression of BAK1 in pancreatic cancer cell lines. Micro RNA-125 b expression and interference lentiviral construct were constructed to infect SW1990 and PANC-1, respectively. Then western blotting and immunofluorescence were used to analyze the expression of BAK1 protein. After the cells were treated with gemcitabine or serum-deprivation, the apoptosis assay was analyzed by flow cytometry, and Cytochrome C and active Caspase-3 were further detected by western blotting. To evaluate the role of mi R-125 b on apoptosis in vivo, we next examined its effects on growth of pancreatic cancer cell xenografts, andtested xenograft tumor tissues by Tunel and Caspase-3.Results: Micro RNA-125 b expression in the pancreatic cancer samples was significantly higher compared to the adjacent tissue samples(p<0.05), and the latter was slightly higher compared to the normal pancreatic tissue samples(p<0.05). In addition, both micro RNA-125 b and BAK1 were detected to express in all 8 pancreatic cancer cell lines. The stably cells, SW1990-mi R-125 b and PANC-1-anti-mi R-125 b, which expressing or interfering mi R-125 b were successfully constructed. The result of Dual-lusiferase reporter confirmed the relationship between micro RNA-125 b and BAK1 m RNA 3’-UTR fragment. Western blotting and immunofluorescence assays also showed that BAK1 was significantly reduced in SW1990-mi R-125 b while increased in PANC-1-anti-mi R-125 b compared with NC group. The results of the apoptosis assay showed that mi R-125 b obviously reduced the apoptotic rates after treated with gemcitabine or serum-deprivation. In vivo, the results demonstrated that micro RNA-125 b might promote pancreatic cancer cell tumorigenesis through playing anti-apoptotic effects.Conclusions: Micro RNA-125 b played anti-apoptotic effects on pancreatic cancer cells by down-regulating the expression of BAK1 through the mitochondrial pathway in vitro or vivo. In addition, micro RNA-125 b might decrease the gemcitabine sensitivity and increase serum-deprivation resistance of pancreatic cancer cells...