| Objective The early detection of pancreatic cancer is very difficult. It has the high malignant degree and its prognosis is also extremely poor. mi R-21 is an important member of the miRNAs and plays a biological function by regulating the expression of multiple genes. In pancreatic cancer tissue samples, miR-21 also has a relatively high level of expression, but how to control the development of tumor, as well as the detailed molecular mechanisms are not well understood. This topic focuses on the biological function of miR-21 and the molecular mechanism of miR-21 in angiogenesis of pancreatic carcinoma by HGF/c-MET signal pathway which may provide theoretical basis for gene therapy of pancreatic cancer.Methods First of all, pancreatic cancer cells were cultivated, such as PANC- 1, MiaPaca – 2, CFPAC-1ã€BxPc-3;Then miR- 21 mimic were transfected into these cells. The effects of miR- 21 expression in pancreatic cancer cells on cell viability were detected by MTT. Growth curve and plate clone formation assay were used to analyze the growth and proliferation ability of tumor cells; The changes of tumor cell migration and invasion were detected by Scratch Test, transwell migration and invasion assay.Using PCR technique to detect the expression of miR-21 level. Cells separately treated with miR-21 inhibitors(AC1MMYR2) and miR-21 mimic, then quantitative PCR was used to detect the expression level of miR- 21, VHL, C- MET in each group. The Western Blot was used to analyze some molecules related to mi R-21 levels in cells, and the possible mechanisms of regulating miR-21 expression. We use ELISA technology to detect the secretion of HGF, and further analyze the effect of miR-21 in angiogenesis of pancreatic carcinoma by vasculogenic mimicry experiment.Results In human pancreatic cancer cell lines, the expression level of mi R-21 was relatively higher. The expression of miR-21 in MiaPaca-2 cells was the lowest in the four pancreatic cancer cells. And the expression level of miR-21 in PANC-1 cells was about 4 times of MiaPaca-2, in CFPAC-1 cell about 5 ~ 6 times which was the highest one, In BxPc-3 only about 1.2 times. After the transfection of miR-21 mimic, the expression levels of miR-21 in MiaPaca-2 cell increased obviously, about 25 to 28 times. Elevated miR- 21 expression level can increase the activity of pancreatic cancer cells, improve the activity of proliferation and strengthen the capacity of migration and invasion.PCR confirmed that miR-21 inhibitor could reduce the expression levels of miR-21〠c-MET and increase the relative expression levels of VHL in PANC-1 cell. miR-21 mimic could increase the expression levels of mi R-21, bring down expression levels of VHL, but the relative expression levels of c-MET had not seen obvious rise. Western Blot indicated that the decreased expression of miR-21 can relatively increase the level of VHL and downgrade the expression levels of HGF, c-MET, AKT. The expression of VHL was decreased when miR-21 expression level was elevated and the protein expression levels of HGF, c-MET,and AKT were raised at the same time. ELISA detected that the overexpressed of miR-21 increased the content of HGF in the medium. Vasculogenic mimicry found that the formation of blood vessels by reducing the expression of mi R-21 is less than the control group. Meanwhile, the increased expression of miR-21, rose the number of blood vessels formed by mimicry compared with the control group.Conclusions In pancreatic cancer cell lines, mi R-21 may play cancer-promoting role and enhance the activity of cancer cells. miR-21 may also improve the activity of proliferation and strengthen the capacity of migration and invasion. miR-21 through HGF/c-MET signaling pathway may play a role in promoting angiogenesis in pancreatic carcinoma. This study mainly discussed the influence of miR- 21 on malignant phenotype of carcinoma and its function in pancreatic cancer angiogenesis. This may provide a new method for treatment of pancreatic cancer. |
