PFT-α对结肠上皮细胞凋亡和周期的影响及机制探讨

Effects of PFT-α on proliferation and apoptosis of large intestinal epithelial cells and its mechanisms

目的探讨p53抑制剂(p-fiftythreeinhibitor-alpha,PFT-α)对结肠上皮细胞凋亡、周期的影响及机制。研究PFT-α对热化疗损伤结肠上皮细胞的影响。方法顺铂联合温热处理原代培养结肠上皮细胞30min,对比加入不同浓度PFT-α后,AnnexinV-FITC/PI染色,流式细胞仪检测细胞凋亡。PI染色检测细胞周期。Westernblot检测结肠上皮细胞CyclinB1和Cdc2(Tyr15)表达。结果不同浓度PFT-α作用于热化疗处理的结肠上皮细胞后,细胞凋亡率下降且呈剂量依赖性。流式细胞仪细胞周期分析显示在运用PFT-α后,结肠上皮细胞的G2/M延长,CyclinB1和Cdc2(Tyr15)蛋白表达随PFT-α的剂量升高而逐渐增强。结论PFT-α可能通过促进CyclinB1蛋白表达,Cdc2(Tyr15)磷酸化水平升高,降低CyclinB1/Cdc2活性,细胞停滞于G2/M期,减轻热化疗对结肠上皮细胞的损伤。

Objective To investigate the effect of PFT-α on proliferation, cell cycle and apoptosis of large intestinal epithelial cells in vitro . Methods The large intestinal epithelial cells were treated with hyperthermic （43℃） and cisplatin （ 10 mg/L） for 30 min in the absence or presence of different dosage of PFT-α. The rate of apoptosis was detected using flow cytometry （FCM） by Annexin V-FITC and propidium iodide （PI） double labeling technique. Cell cycle was analyzed by using FCM with PI labeling technique. The expression of CyclinB1 and Cdc2 （Tyr15） were measured by Western blot. Results After treatment of large intestinal epithelial cells with different concentrations of PFT a and thermochemotherapy, the apoptosis rate was decreased. FCM analysis showed an increase in G2/M phase cell fraction. Western blot displayed significantly higher protein levels of CyelinB1 and Cdc2（Tyr15） than those in the hyperthermic chemotherapy group. Conclusions PFT-α can protect large intestinal epithelial cells against thermochemotherapy-induced damages. The mechanisms of the protective effect of PFT-α is most likely due to the decreased apoptotic large intestinal epithelial cells, and the increased G2/M phase cell fraction and protein of CyclinBl.