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目的观察黄荆子乙酸乙酯提取物(Evn-50)和顺铂(DDP)在体外对人卵巢癌耐顺铂细胞株COC1/DDP凋亡的影响,探讨Evn-50提高人卵巢癌耐顺铂细胞株COC1/DDP对DDP敏感性的机制。方法运用MTT、FCM、Hoechst33258染色法、Westernblot等方法,观察Evn-50增强DDP抑制COC1/DDP细胞增殖作用和此过程中COC1/DDP细胞凋亡,caspase-3蛋白、增殖细胞核抗原(PCNA)及其泛素化修饰蛋白(Ubi-PCNA)表达的改变。结果 Evn-50不仅增强DDP对COC1/DDP细胞的抑制增殖作用,也增强DDP对COC1/DDP细胞的凋亡作用,同时caspase-3蛋白表达增加(P<0.05);而Evn-50(20μg/ml)组与空白对照组相比PCNA蛋白表达下调(P<0.05),Evn-50与DDP合用组与DDP单药组均出现了Ubi-PCNA蛋白表达,Evn-50与DDP合用组与DDP组比较,Ubi-PCNA蛋白表达下调(P<0.05)。结论 Evn-50可能通过减少PC-NA表达,使DDP诱导的PCNA泛素化水平下降,从而逆转DDP耐药,最终通过激活caspase-3蛋白高表达,启动caspase级联反应,增强DDP对人卵巢癌DDP耐药细胞株COC1/DDP的凋亡作用。
Objective To observe the effect of Evn-50 and DDP on the apoptosis of human ovarian cancer cell line cisplatin-resistant COC1 / DDP in vitro and to explore the effect of Evn-50 on the proliferation of cisplatin-resistant human ovarian cancer cell line Mechanism of strain COC1 / DDP sensitivity to DDP. Methods MTT, FCM, Hoechst33258 staining and Western blot were used to observe the effects of Evn-50 on proliferation of COC1 / DDP cells induced by DDP and the changes of apoptosis, expression of caspase-3 protein, proliferating cell nuclear antigen (PCNA) Its ubiquitylation modified protein (Ubi-PCNA) expression changes. Results Evn-50 not only enhanced the inhibitory and proliferative effects of DDP on COC1 / DDP cells but also enhanced the apoptosis of COC1 / DDP cells induced by DDP and the expression of caspase-3 protein increased (P <0.05) (P <0.05). The expressions of Ubi-PCNA protein in Evn-50 and DDP combined groups and DDP alone group were significantly lower than those in control group Ubi-PCNA protein expression was down-regulated (P <0.05). Conclusions Evn-50 may reduce the PCNA expression and decrease the DDP-induced PCNA ubiquitination, thus reversing the drug resistance of DDP. Finally, Evn-50 may activate Caspase-3 cascade and activate caspase cascade, Apoptosis of DDP Cell Line COC1 / DDP.