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目的:研究抗HPV16E6核酶(Ri-bozyme)对宫颈癌CaSki细胞顺铂敏感性的影响和机制。方法:将抗HPV16E6核酶、空载体质粒分别导入CaSki细胞,命名为CaSki-R、CaSki-P细胞。Northern blot、MTT比色法、Annexine/PI双标法、流式细胞术及RT-PCR扩增法分别检测E6基因、顺铂的敏感性、凋亡率、凋亡相关蛋白和基因。结果:CaSki-R表达E6较CaSki-P、CaSki明显降低;与CaSki细胞和CaSki-P细胞比较,CaSki-R对DDP敏感性分别增加了2.28倍和2.21倍。CaSki、CaSki-P和CaSki-R 3种细胞的凋亡率分别为(18.9±3.5)%、(19.7±4.8)%和(40.4±4.5)%,CaSki-R较CaSki细胞的凋亡率明显增加,P=0.003;DDP作用后CaSki-R细胞的p53蛋白(P=0.000)和Bax蛋白(P=0.002)表达较CaSK细胞明显增加,Bcl-2蛋白(P=0.005)、c-myc蛋白(P=0.005)表达则明显减小;与CaSK细胞比较,CaSki-R细胞表达p53、Bax mRNA明显增加,Bcl-2、c-myc mRNA表达明显减少。结论:转染抗HPV16E6-Ri-bozyme的CaSki-R细胞对顺铂的敏感性明显增加,p53、Bax蛋白表达增加,Bcl-2和c-myc蛋白表达减少可能与CaSki-R细胞对顺铂的敏感性增加有关。
Objective: To investigate the effect and mechanism of anti-HPV16E6 ribozyme on cisplatin sensitivity in cervical cancer CaSki cells. Methods: The anti-HPV16E6 ribozyme and empty plasmid were respectively introduced into CaSki cells and named as CaSki-R and CaSki-P cells. Northern blot, MTT assay, Annexin / PI double labeling, flow cytometry and RT-PCR were used to detect the expression of E6 gene, cisplatin sensitivity, apoptosis rate, apoptosis related proteins and genes respectively. Results: The E6 expression of CaSki-R was significantly lower than that of CaSki-P and CaSki. Compared with CaSki cells and CaSki-P cells, the sensitivity of CaSki-R to DDP increased 2.28-fold and 2.21-fold, respectively. The apoptotic rates of CaSki, CaSki-P and CaSki-R were (18.9 ± 3.5)%, (19.7 ± 4.8)% and (40.4 ± 4.5)%, respectively. (P = 0.002), the expression of Bcl-2 protein (P = 0.006), c-myc protein (P = 0.005). Compared with CaSK cells, CaSki-R cells expressed p53, Bax mRNA and Bcl-2, c-myc mRNA significantly decreased. CONCLUSIONS: The sensitivity of cisplatin-transfected CaSki-R cells transfected with HPV16E6-Ri-bozyme is significantly increased, the expressions of p53 and Bax are increased, and the decreased expression of Bcl-2 and c-myc proteins may be related to the inhibitory effect of CaSki-R cells on cisplatin Increased sensitivity of the relevant.