曲古抑菌素A和紫杉醇对子宫内膜癌细胞凋亡和微管稳定性的影响

目的 探讨曲古抑菌素A(TSA)和紫杉醇(PTX)体外对人子宫内膜癌细胞凋亡和微管稳定性的影响及其机制.方法 将浆液性乳头状子宫内膜癌Ark2细胞、低分化子宫内膜样腺癌KLE和AN3细胞各分为TSA、PTX单独作用组(TSA组、PTX组)和联合作用组(TSA+PTX组),并设空白对照组,以锥虫蓝拒染法观察药物对细胞增殖的影响,膜联蛋白V、Hoechst染色法检测细胞凋亡率,流式细胞仪测定线粒体膜电位(MMP),蛋白质印迹法检测半胱天冬酶9(caspase-9)、多聚ADP核糖聚合酶(PARP)和乙酰化微管蛋白的表达.结果 TSA、PTX对Ark2、KLE和AN3细胞均有抑制作用,二药联用后抑制作用更强.药物作用后4 d,TSA、PTX、TSA+PTX和对照组Ark2细胞凋亡率膜联蛋白V染色法检测分别为4.25%±0.25%、12.12%±0.62%、16.56%±0.74%和46.78%±2.68%,Hoechst染色法检测分别为3.39%±0.12%、6.00%±0.25%、10.05%±0.53%和22.30%±1.25%,TSA+PIX组明显高于其他各组(均P＜0.05).药物作用后24 h,Ark2和KLE细胞的TSA+PTX组caspase-9、PARP的裂解明显增加;Ark7.和AN3细胞的TSA+PTX组MMP消失率(16.80%±0.92%、11.28%±0.78%)明显高于TSA组(4.96%±0.47%、6.46%±0.62%)和PTX组(5.34%±0.45%、5.61%±0.56%)(均P＜0.05);Ark2和KLE细胞的TSA+FIX组乙酰化微管蛋白表达明显增加.结论 TSA和PTX可通过促进子宫内膜癌细胞中乙酰化微管蛋白表达和增强微管稳定性而产生协同抗肿瘤作用.

Effects of trichostatin A and paclitaxel on apoptosis and microtubule stabilizatian in endometrial carcinoma cells: an in vitro research

Objective To investigate the effects of trichostatin A (TSA) and paclitaxel (PTX) on the apoptosis and microtubulin stabilization in human endometrial carcinoma cells and its mechanism. Methods Human endometrial carcinoma cells of the line Ark7., KLE and AN3 were cultured in the presence OfTSA (TSA group), or PTX(PTX group) , or TSA plus PTX (TSA +PTX group) respectively. The growth curve was obtained by trypan-blue exclusion assay. Apoptosis was observed by annexin V and Hoechst staining. Perturbation of mitochondrial membrane potential (MMP) was detected by flow cytometry. Western blotting was used to detect the protein expression of caspaae-9, poly ADP-riboae polymerase (PARP), and acetylated microtubulin. Results The growth of the Ark2, KLE, and AN3 cells of the TSA, PTX, and TSA + PTX group, especially in the latter group, was inhibited. The Ark2 cell apoptotic rates 4 days later of the TSA, PTX, TSA + FTX, and control group were 4.25%±0.25%, 12.12% ±0.62%, 16.56%±0.74%, and 46.78%±2.68% respectively by annexin V staining, and 3.39%±0.12%, 6.00%±0.25%, 10.05%±0.53%, and 22.30%±1.25% respectively by Hoechst staining. The apeptotie rates of the TSA + PTX group by both staining methods were both significantly higher than those of the other groups (all P＜0.05). Lysis of caspase-9 and PARP in the Ark2 and KLE cells increased greatly 24 hours after the TSA and PTX treatment. The disappearance rate of MMP in the Ark2 and AN3 cells of the TSA +PTX groups were 16.80%±0.92% and 11.28%±0.78% respectively, significandy higher than that of the PTX group (5.34%±0.45% and 5.61%±0.56% respectively) and TSA group (4.96%± 0.47% and 6.46%±0.62% respectively, all P＜0.05). The expression of acetylated mieretubulin was increased in the Ark2 and KLE cells of the TSA + PTX groups. Condusions Synergy of TSA and PTX inhibits the cell growth and induces apoptosis. The acetylation of non-histone protein induced by histone deacetylase inhibitor is one of the possible mechanisms of its anti-cancer effects.