| Cholangiocarcinoma is the most frequently occurred malignancy of the biliary tract, it has a dismal prognosis due to low surgery resection rate resulting from delayed symptoms and complicated anatomy of the tumor location, as well as the refractoriness to conventional chemotherapy and radiotherapy. To improve the therapy outcome of cholangiocarcinoma, lots of research work is going on in different ways. It shows that some peptide hormones , such as EGF, insulin IGF, gastrin, bombesin and somatostatin(SST) which inhibits proliferation of gastrointestinal cancer cells, could regulate the proliferation of gastrointestinal cancer cells. The comprehensive biology functions of somatostatin and its analogues are mediated by the five somatostatin receptor subtypes expressing on the cell membrane, in which somatostatin receptor 2 (SSTR2) plays an important role in the anti-proliferation effects. SSTR2 inhibits proliferation of cancer cells directly by inducing G1 cell cycle arrest or apoptosis . As a result of the inhibition of secretion of growth promoting hormones and growth factors, angiogenesis or blood supply to the tumor, indirect anti-proliferation can also be mediated by SSTR2 . Currently available data have demonstrated expression of messenger RNA for human SST-receptor subtype 2 (SSTR2) in biliary tract endothelial cell. In this study, we would provide a few certain proofs for clinic therapy with somatostatin through evaluting the growth-inhibiting and apoptosis-inducing effects of somatostatin and its probable mechanisms on human cholangiocarcinoma cell lineQBC939 cultured in vitro.Objective The purpose of this study is to examine the effect of somatostain on cholangiocarcinoma cell line QBC939 in vitro and its probable mechanisms.Methods QBC939 cells were cultured in vitro and seeded into culture plates. Suseptibility of Octreotide on QBC939 human cholangiocarcinoma cell lines in vitro were determineted by MTT assay, cell quantification and colony-forming test at different concentrations of Octreotide , and the growth rate were calculated, the content of cell DNA were determined by FCM (flowcytometry) /electronic microscope/DNA ladder determinations for evaluation of the change of every-phase fraction of cell cycle, proliferous index and apoptosis for cholangiocarcinoma cell at different concentrations of drug. Expression of wild P53 mRNA of QBC939 human cholangiocarcinoma cell lines was investigates by RT-PCR. The effect of somatostatin analogue Octreotide on the expression of cell inhibition protein P53 families in QBC939 and the expression of P73 protein was detected with Western-blotting. All the data were analyze with the statistical software SPSS 11.0. Categorical variables were analyzed with Chi-square test and continous variables with One-way ANOVA.Results The results of experiment showed that Octreotide can inhibit the growth of QBC939 human cholangiocarcinoma cell line in vitro. The inhibitory effects was dose-dependent and time-dependent: the more affective when the concentration of Octreotide was higher and the time of contacting was longer. According to tumor cell counting, the growth rate of cells of every experiment groups were: 96.65%, 96.38%, 95.29%, 81.80% and 81.30% (0.01mg/L group), 93.12%, 92.60%, 91.32%, 72.19% and 70.46% (0.1mg/L group), 93.03%, 91.70%, 72.29%, 51.80% and 50.85% (1.0mg/L group) respectively after 241h, 36h, 48h, 72h, 96h of Octreotide exposure. Thecolony-forming rates were 70.73% (control group) and 8.08% (P<0.001) respetively. After 48h exposure to 1.0mg/L Octreotide, flow cytometric analysis demonstrated an increased number of cells in G0/G1 phase associated with a decreased number of cells in G2/M and S phase. Apoptosis was observed in QBC939 cells after 66h exposure to 1.0mg/L Octreotide. Apoptosis was detected by eletronic microscope/flow cytometry/DNA ladder determinations. The mRNA expression of wild p53 was not detected and semi-quantified by RT-PCR technique. The protein level of p73 had not statistic significance using Western-blots.Conclusions:... |
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