hepaCAM基因协同吡柔比星对膀胱癌细胞增殖影响的研究

发布时间：2018-03-20 03:37

本文选题：hepaCAM　切入点：吡柔比星　出处：《重庆医科大学》2014年硕士论文　论文类型：学位论文

【摘要】：第一部分hepaCAM基因联合吡柔比星增强对膀胱癌细胞BIU-87增殖的研究目的：研究肝细胞粘附分子(hepatocyte cell adhesion molecule，hepaCAM)联合吡柔比星（pirarubicin, THP）对膀胱癌细胞BIU-87增殖的影响。方法：用腺病毒向膀胱癌BIU-87导入hepaCAM基因，加吡柔比星处理后，采用实时荧光定量PCR检测pAdH5-hepaCAM处理组，吡柔比星处理组，吡柔比星联合pAdH5-hepaCAM处理组和未感染组的膀胱癌BIU-87细胞中相关基因cyclinD1、CDK2的mRNA表达水平，同时MTT测各处理组细胞抑制率及FCM测各处理组细胞周期变化。结果：实时荧光定量PCR结果显示：吡柔比星联合pAdH5-hepaCAM处理组，较单独处理组，cyclinD1、CDK2的mRNA水平明显降低（P0.05）； MTT检测的各处理组中联合处理组细胞增殖明显受到抑制；FCM检测证实，hepaCAM基因和吡柔比星联合处理组细胞G0/G1期较空白组和hepaCAM基因、吡柔比星单独处理组相比，明显升高（P0.05）。结论： hepaCAM基因感染膀胱癌细胞BIU-87，再给予吡柔比星药物处理后，细胞增殖受到抑制和细胞周期出现了明显的G0/G1阻滞，因此，hepaCAM基因联合吡柔比星可以增强对膀胱癌细胞增殖作用。第二部分hepaCAM基因对膀胱癌肿瘤生长体内实验研究目的：研究hepaCAM基因对膀胱癌BIU-87细胞株裸鼠移植肿瘤生长情况的影响。方法：将9只裸鼠随机为空载组和hepaCAM组，于裸鼠右下肢背外侧皮下接种BIU-87细胞l×107/只，接种完毕后放SPF环境培养，，每天定时观察成瘤情况并记录瘤体体积和裸鼠重量。三周后处死老鼠，取瘤体比较两组裸鼠的成瘤大小，同时取瘤组织做免疫组化检测hepaCAM蛋白的表达。结果：空白组与感染组相比较，瘤体积明显较大（p0.05），免疫组化显示空白组hepaCAM蛋白表达比感染组明显减弱。结论：以腺病毒为载体，体内实验进一步证实hepaCAM基因能明显抑制膀胱肿瘤的生长。因此，以腺病毒为载体的hepaCAM基因有可能成为一种有效的基因治疗方法。
[Abstract]:A study on the effect of hepaCAM gene combined with pirarubicin on the proliferation of bladder cancer cell line BIU-87. Aim: to study the effect of hepatocyte cell adhesion molecule (hepatocyte cell adhesion molecule) combined with pirarubicin (THP1) on BIU-87 proliferation of bladder cancer cells. Methods: hepaCAM gene was introduced into bladder cancer BIU-87 by adenovirus. After treated with pirarubicin, real-time fluorescence quantitative PCR was used to detect pAdH5-hepaCAM treatment group and pirarubicin treatment group. The mRNA expression of cyclin D1 CDK2 was detected in bladder cancer BIU-87 cells treated with pirarubicin combined with pAdH5-hepaCAM, and the cell cycle changes of each treatment group were measured by MTT and FCM. Results: the results of real-time fluorescence quantitative PCR showed that Pirarubicin combined with pAdH5-hepaCAM treatment group, The level of mRNA in cyclin D1 + CDK2 group was significantly lower than that in the control group (P 0.05), and the cell proliferation in the combined treatment group was significantly inhibited by MTT. The results of FCM showed that the G0 / P G 1 phase of the treated group was significantly lower than that of the control group and the hepaCAM gene of the combined treatment group was higher than that of the control group, and that of the combined treatment group was significantly lower than that of the control group and the hepaCAM gene group. Compared with the group treated with pirarubicin alone, it was significantly higher than that of the control group (P 0.05). Conclusion: after hepaCAM gene was infected with BIU-87 and then treated with pirarubicin, cell proliferation was inhibited and cell cycle was significantly blocked by G _ 0 / G _ 1. Therefore, hepaCAM gene combined with pirarubicin could enhance the proliferation of bladder cancer cells. The second part: an in vivo study on the effect of hepaCAM gene on the growth of bladder cancer. Aim: to study the effect of hepaCAM gene on the growth of bladder cancer BIU-87 cell line in nude mice. Methods: nine nude mice were randomly divided into no-load group and hepaCAM group. BIU-87 cells were inoculated subcutaneously on the dorsolateral side of right lower extremity of nude mice. After inoculation, BIU-87 cells were cultured in SPF environment. The tumor size and weight of nude mice were recorded three weeks later. The tumor size of the two groups was compared and the expression of hepaCAM protein was detected by immunohistochemistry. Results: compared with the infected group, the tumor volume of the blank group was significantly larger than that of the infected group, and the expression of hepaCAM protein in the blank group was significantly lower than that in the infected group. Conclusion: using adenovirus as vector, it is further confirmed that hepaCAM gene can inhibit the growth of bladder tumor in vivo. Therefore, hepaCAM gene with adenovirus as vector may become an effective gene therapy method.
【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R737.14

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