目的:研究梯度热打击对体外培养肠黏膜上皮细胞活性和增殖的影响。方法:肠黏膜上皮细胞株IEC-6经培养后分为正常对照组(37℃、5%CO2培养箱培养)及39℃、41℃、43℃热打击组(分别在相应温度培养箱中培养),相差显微镜观察各组细胞培养1h的形态学改变,CCK8法比较各组细胞培养0、1、3、5、7h的细胞活性和24h增殖率,流式细胞术研究细胞周期的改变。结果:与正常对照组比较,各热打击组各个时相细胞形态变圆,伪足变短,细胞间隙增大,活力下降(P〈0.01)。24h细胞增殖实验显示,与正常对照组比较,39℃组7h及41℃和43℃组各时相增殖率显著下降(P〈0.01),呈时间及温度依赖关系。流式细胞术检查显示,热打击组细胞呈现细胞周期G0/G1和G2/M期阻滞。结论:热打击对IEC-6细胞具有细胞毒效应,可抑制IEC-6细胞的增殖,造成细胞周期G0/G1和G2/M期阻滞。
Objective: To investigate the effect of gradient heat stress on the viability and proliferation of intestinal epithelial cells in vitro. Methods: The cultured intestinal epithelial cells IEC-6 were divided into normal control group(37 ℃ )and 39 ℃ , 41 ℃ , 43 ℃ heat stress groups. The IEC-6 cells in different groups were cultured in a 5 % carbon dioxide incubator, and they were exposed to the different temperature in vitro. Phase contrast microscope was used to observe the morphology of IEC-6 1 hour after thermal stimulation, and CCK 8 assay was used to determine cellular viability after 0, 1, 3, 5, 7 hours of thermal stimulation, as well as 24-hour proliferation rate of each group. Flow cytometry was used to investigate the influence of heat stress on the change of IEC-6 cell cycle. Results: Compared to the normal control group, cells of each heat stress group became rounded, with shorter pseudopod, increase in cellular interspace, and declination of vitality(P〈0.01). The assay of 24-hour proliferation indicated that the proliferation rate of the cells undergone 7 hours of exposure to 39 ℃ group and that of each phase of cells underg one 41 ℃ and 43℃ were significantly decreased compared to the normal control group(P〈0.01), and the decrease was time-and thermal-dependent. With flow cytometry, arrest of cell cycle at G0/G1 and G2/M phases was observed in heat stress groups. Conclusions: Heat stress has eytotoxic effect on intestinal epithelial cells IEC-6. It inhibits cellular proliferative capacity, and arrests cell cycle at G0/G1 and G2/M phases.