The Growth-inhibiting And Apoptosis-inducing Effects Of Arsenic Trioxide On Human Cholangilcarinoma Cell Line QBC939

Background: Cholangiocarcinoma is a kind of digestive malignant tumor which grow slowly and metastasize at the later stage. Cholangiocarcinoma have a high degree of malignancy and a poor prognosis. Cholangiocarcinoma is resistant to current surgica. chemotherapy and radiotherapy.therefor Cholangiocarcinoma is a virtually incurable tumor. Patients who suffer from Cholangiocarcinoma have bad prognosis and the death rate is higher than many other tumors.Arsenic trioxide is a kind of extensive toxic material in nature. In ancient.lt was used by Chinese medicine to theropy psoriasis and syphilis. In recent years, arsenic trioxide has been applied for the treatment of Leukemia successfully. The experimental studies and the clinical trail have show that arsenic trioxide could inhibit carcinoma cell proliferation and induce apoptosis of tumor cells, but ,It is not reported in domestic and outside that arsenic trioxide was applied on the study of human Cholangiocarcinoma cell line.ABM:The purpose of this study is to evaluate the growth-inhibiting and apoptosis-inducing effects of arsenic trioxide on human cholangilcarinoma cell line QBC939 cultured in vitro.fmd out the effect and anticancer mechanism ofarsenic trioxide on human cholangiocarcinoma cell line QBC939.Method: human cholangiocarcinoma cell line QBC939 was recovered and divided into control and experiment groups to culture.The experiment groups cell was treated with different concentration of arsenic trioxide and control groups without it. (1) Benzo blue dyeing: QBC939cell 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 20 u 1 MTT in cultured QBC939 cell about 4h before the end of experiment, dissolved with 150 n 1DMSO 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) Morphology observation of the cell: at the state of culturing, observe the morphology of cell of experiment groups and control in culture flask by invert microscope. Collecting cell differently that has been cultured with different concentration of arsenic trioxide and the control groups cell after 72h of being cultured , washed with PBS, centrifugalized, fixed with 40 u mol.L"1 diamylic aldehye, and observed phenotype of cell line in transmission electronic microscope.(4) Row cytometric: collecting the cell that had been treated by different concentration of arsenic trioxide and the cell without it, washed with PBS, centrifugalized, fixed with 700 mL ?L"1 alcohol dyed with 50 u g.iriT1 PI, and analysed cell cycle and DAN content with flow cytometry. (5) DNA Ladder determinations: routinessly, collecting the cell of experiment groups and the control groups that has been digested by trypsin, washed with PBS. According to the explanation steps of Test Kit, to burst the cell treated with Rnase, toprecipitate protein and get out the DNA, then, the DNA be dissolved in TE Buffer, 15mL -L"1 agarose gel electrophoresis, abservation and photography for it.Results: (1) Different concentration of arsenic trioxide could inhibit the proliferation of human cholangiocarcinoma cell line QBC939, which had obvious dose and time effect, but treatment with high dose would induce necrosis principally, the appropriate concentration of arsenic trioxide was 4n mol.L '.(2) The MTT methods showed that the proliferation inhibitory rate was strengthened by the prolong of time, and the proliferation inhibitory rate of QBC939 cells about 76.5% after treated with arsenic trioxide at 4u mol ?L"1 for 96h. (3) Observing with optical microscope show that the shape of QBC939 cells were changed from long shuttle to round or oval, and apoptosis corpuscle was observed through scanning and transmission electronic microscope. (4) Flow cytometric analysis demonstrated that cell cycle of QBC939 cells was pro...