维生素c诱导人胃癌细胞株mkn45凋亡机制的分析-analysis of the mechanism of vitamin c inducing apoptosis of human gastric cancer cell line mkn 45.docxVIP

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维生素c诱导人胃癌细胞株mkn45凋亡机制的分析-analysis of the mechanism of vitamin c inducing apoptosis of human gastric cancer cell line mkn 45.docx

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维生素c诱导人胃癌细胞株mkn45凋亡机制的分析-analysis of the mechanism of vitamin c inducing apoptosis of human gastric cancer cell line mkn 45

Study of gastric cancer cell MKN45 apoptois and related mechanism induced by vitamin CPostgraduate: Tutor :AbstractObjective:(1) Investigate the capability of Vitamin C to induce the apoptosis of poorly differentiated gastric cancer cell line MKN45 in vitro. (2) To determine the role of oxidative stress in Vitamin C-induced apoptosis in human gastric cance cells MKN45. (3)Investigate correlated possible mechanism of Vitamin C- induced apoptosis in gastric cance cells MKN45.Method:1、Add different concentrations of Vitamin C（0.05mg/ml? 0.1mg/ml? 0.5mg/ml? 1mg/ml? 1.5mg/ml） into gastric cancer cell line MKN45 which was cultured in vitro, Moreover add the specific H2O2-degrading enzyme(CAT) alone into MKN45 cell line and add CAT into MKN45 cell line with the concentrations of Vitamin C which can induce apoptosis in MKN45 cells , the effect of Vitamin C and CAT on growth was measured by MTT assay. Apoptosis was detected by fluorescence microscope, flow cytometry (FCM) and DNA Ladder analysis method. The activity of SOD and the levels of MDA in gastric cancer cells after 24h treatment were assayed by chemical staining method. Level of H2O2 in medium supernatant was detected by special kits.2、After the stimulation by Vitamin C and CAT,the change of Caspase-3 activity wastested by spectrophotometry . Reverse transcriptase polymerase chain reaction (RT- PCR) was used to detect Caspase-3 expression levels and the mRNA level of bcl-2 gene family was semi-quantified by a reverse transcriptase-polymerase chain reaction technique.Results:1、After treatment with 0.05-0.5mg/ml concentration of vitamin C for 12-36 hours, the growth inhibition ratio of MKN45 cells has no time dependent manner but concentration dependent manner. the proliferation of MKN45 cells was significantly inhibited . 1mg/mlconcentration of vitamin C can induce apoptosis of gastric cancer cells MKN45 after vitamin C was added into MKN45 cells for 24 hours, the apoptotic rate of 1mg/ml vitamin C treatment gro