Study On The Apoptosis By All-trans Retinoic Acid In Human Cholangiocarcinoma Cell

Objective Retinoids is the derivatives of Vitamin A, Which have the important effect on the process of embryo growth~ cell proliferation and differentiation. Retinoic acid was widely used to study the prevention and therapy of carcinoma in recent years. The experimental study and research show that all-trans retirioic acid (ATRA) could inhibit carcinoma cell proliferation~ promote cell differentiation to end and also induced apoptosis, but it is not reported in domestic and outside that all-trans retinoic acid was used on the study of cholangiocarcinoma cell line. Different experimental index were measured to study all-trans retinoie acid-induced apoptosis in human cholangio- carcinoma cell line QBC939 cultured in vivo, and find out the effect and anticancer mechanism of all-trans retinoic acid on the growth of cholangiocarcinoma cell QBC939. Methods Human cholangiocarcinoma cell line QBC939 was resuscitation and divide into control and experiment groups to culture. The experimental group cell was treated with different concentration of all-trans retinoic acid. (1) Benzo blue dyeing: QBC939 cell of control and experiment groups were dyed with benzo blue every day, and calculated cell number in the cell calculated counter to draw the growth curve of cell after a week. (2) MTT methods: Stop the experiment in different time, and add to MTT in cultured QBC939 cell about 4h before the end of the experiment, dissolve with DMSO after QBC939 cell was dyed, detect inhaling light degree (A numerical value) of 490nm in detection instrument, calculate proliferation inhibitory rate of QBC939 cell. (3) Scanning and transmission electronic microscopic technique: Control and experiment groups QBC939 cell were treated with lO5molfor 3d, digested by trypsin, washed with PBS, centrifugalized, fixed with 3% diamylic aldehye, and observed phenotype of cell line in optical and scanning and transmission electronic microscope. (4) Flow cytometric analysis: QBC939 cell was digested by trypsin, fixed with 70% alcohol, added to P1 to dye, and analysed cell cycle with flow cytometry. Results (1) Different concentration of all-trans retinoic acid could inhibit the proliferation of human cholangio- carcinoma cell line QBC939, which had obvious dose and time effect, but treatment with high dose would induce cell necrosis principally, the appropriate concentration of all-trans retinoic acid was I O5mobIY? (2) The MTT methods showed that the proliferation inhibitory rate were strengthened by the prolong of time, and the proliferation inhibitory rate of QBC939 cells was about 60.5% after treated with all-trans retinoic acid at lO5mol?for 7d. (3) Observing with optical microscope show that the shape of QBC939 cells were changed from long shuttle to round or oval, and apoptosis was observed through scanning and transmission electronic microscope. (4) Flow cytometric analysis demonstrated that cell cycle of QBC939 cells was prolonged after using lO5mol.L?all-trans retinoic acid. The G0/G1 proportion of cell cycle is significantly increased by 50.5%, and the S and G2 is reduced by 39.8% and 11.4%, and show up the summit of typical apoptosis. Conclusion All-trans retinoic acid could suppress the growth of cholangiocarcin...