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目的:观察羟基喜树碱(HCPT)对高危浅表性膀胱癌模型5637细胞的凋亡作用,研究HCPT不同作用时间和不同浓度对5637细胞的影响,进一步讨论HCPT在膀胱灌注化疗中的作用。方法:采用四甲基偶氮唑盐(MIT)检测不同浓度、不同作用时间下HCPT对5637细胞增殖的影响;采用流式细胞术测定低浓度(5μmol/L)、不同作用时间(12、24 h)HCPT诱导5637细胞的凋亡率;用Hochest 33258染色观察细胞凋亡情况;采用荧光定量PCR检测5μmol/L HCPT作用5637细胞12 h相较未加药5637细胞中凋亡相关基因的变化。结果:5μmol/L HCPT作用5637细胞12 h后,早期凋亡细胞占23.5%。HCPT随着作用时间的延长,凋亡率并不会随之增加,HCPT诱导凋亡的作用在肿瘤细胞与药物作用后的某一时间点达到高峰。Hochest 33258染色和荧光定量PCR检测凋亡相关基因也证明HCPT这一浓度在这一时间点有诱导癌细胞凋亡的作用。结论:羟基喜树碱具有诱导高危浅表性膀胱癌细胞凋亡的作用。适当延长药物作用时间,可以促进其药效的发挥。
OBJECTIVE: To observe the effect of HCPT on the apoptosis in 5637 cells of high-risk superficial bladder cancer model, and to study the effect of HCPT on 5637 cells at different time and different concentrations, and further discuss the role of HCPT in bladder infusion chemotherapy. Methods: The effects of HCPT on the proliferation of 5637 cells were detected by MTT assay at different concentrations and different time. Flow cytometry was used to determine the effect of HCPT at low concentration (5μmol / L) and different time (12, 24 h) The apoptotic rate of 5637 cells induced by HCPT; The apoptosis of 5637 cells was observed by Hochest 33258 staining; The apoptosis-related genes of 5637 cells treated by 5μmol / L HCPT for 56 h were compared with that of 5637 cells by fluorescence quantitative PCR. Results: After treated with 5μmol / L HCPT for 56h, the number of apoptotic cells in early stage was 23.5%. HCPT with the extension of time, apoptosis rate does not increase, HCPT induced apoptosis in tumor cells and drugs at a point in time after the peak. Hochest 33258 staining and fluorescence quantitative PCR detection of apoptosis-related genes also proved that the HCPT concentration at this time point to induce cancer cell apoptosis. CONCLUSION: Hydroxycamptothecin can induce the apoptosis of high-risk superficial bladder cancer cells. Appropriate to extend the role of drugs, can promote their efficacy.