索拉非尼对宫颈癌细胞的增殖抑制作用及对PCNA、Survivin表达的影响

目的 探讨索拉非尼对宫颈癌细胞株增殖和凋亡的影响。方法体外培养宫颈癌HeLa、SiHa细胞株，实验分为空白对照组、阴性对照组（DMSO）和索拉非尼（0、25、5、10、20 μmol／L）处理组。光学显微镜观察各组细胞形态学改变；Hoechst荧光染色观察凋亡细胞核的形态学变化；采用MTS法检测索拉非尼对HeLa和SiHa细胞的增殖抑制作用；免疫细胞化学法检测索拉非尼（10 μmol／L）处理HeLa、SiHa细胞48 h后Survivin蛋白的表达；Western blotting检测索拉非尼对宫颈癌HeLa、SiHa细胞PCNA、Survivin蛋白表达的影响。结果不同浓度索拉非尼作用48 h后，HeLa、SiHa细胞出现了凋亡形态学改变；Hoechst染色显示，HeLa、SiHa细胞的胞核变小、固缩，荧光明显增强。MTS检测显示，索拉非尼可抑制宫颈癌HeLa、SiHa细胞增殖，并呈浓度依赖性（P＜0.05）。免疫细胞化学法检测显示，阴性对照组Survivin在HeLa和SiHa细胞中的平均光密度值分别为0.19±0.05和0.17±0.07，与索拉非尼处理组的0.13±0.01和0.13±0.03比较，差异均有统计学意义（P＜0.05）。Western blotting检测显示，索拉非尼在蛋白水平上能够降低PCNA、Survivin表达，且这种下调作用具有浓度依赖性。结论索拉非尼能诱导HeLa、SiHa细胞凋亡，抑制其增殖且呈浓度依赖性，其机制可能与下调癌基因 PCNA、Survivin表达有关。

Effect of sorafenib on the proliferative inhibition of cervical cancer cells and the influence on the expression of PCNA,Survivin

Objective To explore the influence of sorafenib on the proliferation, apoptosis and expression of PCNA, Survivin of cervical cancer cells. Methods Cervical cancer cells HeLa and SiHa cultured in vitro were divided into blank control group, negative control group(DMSO) and sorafenib experimental group(2. 5, 5, 10, 20μmol/L). The light microscope was used to observe the morpho-logical changes. Hoechst 33258 was used to observe nucleus morphological changes in HeLa, SiHa cells treated with sorafenib(20μmol/L) for 48 h. The effect of sorafenib on the proliferation inhibition of HeLa, SiHa cells was detected by MTS. Immunocytochemical method was used to detect the expression of Survivin with sorafenib(10μmol/L) acting on HeLa and SiHa cells for 48 h. The influence of sor-afenib on the expressions of PCNA, Survivin protein of HeLa, SiHa cells was determined by Western blotting. Results After HeLa, Si-Ha cells exposed to sorafenib with different concentration for 48h, the typical apoptotic morphological changes were observed under the light microscope. Hoechst 33258 showed shrink and pyknosis of nucleus, and the fluorescence enhanced obviously. The results of MTS showed that sorafenib significantly inhibited the proliferation of cervical cancer cells in a dose-dependent manner( P<0. 05) . Immunocyto-chemistry showed that in negative control group, the average optical density of Survivin in HeLa, SiHa cells were 0. 19±0. 05 and 0. 17± 0. 07, higher than 0. 13±0. 01 and 0. 13±0. 03 in sorafenib experimental group(P<0. 05). Western blotting showed that the protein ex-pression of PCNA, Survivin of HeLa and SiHa cells treated by sorafenib was down-regulated compared with the negative control group in a dose-dependent manner. Conclusion Sorafenib induces apoptosis of HeLa and SiHa cells, and inhibits cell proliferation in a dose-de-pendent manner. The possible mechanism lays in the decrease of PCNA and Survivin in HeLa and SiHa cells.