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目的建立小鼠成纤维细胞系NIH-3T3预热适应细胞模型,探讨应激与适应对细胞活性和热休克蛋白90(HSP90)合成的影响。方法通过预热适应(42℃,20 min)建立应激适应细胞模型,并通过再次热应激时(44℃,40min)细胞膜损伤指标、DNA损伤指标的变化综合评价适应效果。以Western blot法检测应激及适应对细胞内HSP90合成的影响。结果结合预热适应后再次热应激所致的细胞膜损害和HSP90合成情况,初步确定预热适应后6 h为最佳应激保护时间。预热适应6 h后,再次热应激时,培养液中乳酸脱氢酶(LDH)漏出变化率为15．4%±2．6%,对照组为41．2%±5．1%；DNA受损细胞所占百分比为15．1%,较直接热应激组(26．3%)轻。OD_(HSP90)/OD_(control)变化趋势显示热应激40min后细胞内HSP90含量均呈下降趋势,直接热应激组为0．82±0．18,预热适应组为1．70±0．52,预热适应+热应激组为1．41±0．16。结论通过对NIH-3T3细胞进行预热处理,确定应激保护的时间点,建立了细胞应激适应模型；初步确认HSP90在该模型中的保护作用。
OBJECTIVE: To establish a mouse model of NIH-3T3 preconditioning and to investigate the effects of stress and adaptation on cell viability and the synthesis of heat shock protein 90 (HSP90). Methods The model of stress adaptation was established by preheating adaptation (42 ℃, 20 min). The effect of adaptation was evaluated by the changes of cell membrane damage index and DNA damage index after reheating (44 ℃, 40 min). The effect of stress and adaptation on the synthesis of intracellular HSP90 was detected by Western blot. Results Combined with the damage of cell membrane and the synthesis of HSP90 caused by reheat after reheat, the optimum time of preconditioning protection was preliminarily determined at 6 hours. After preheating for 6 h, the rate of change of lactate dehydrogenase (LDH) leakage in culture solution was 15.4% ± 2.6% and 41.2% ± 5.1% in the control group after heat stress again. The percentage of DNA damaged cells was 15.1%, which was lighter than that of direct heat stress group (26.3%). The change trend of OD_ (HSP90) / OD_control showed that the content of HSP90 in the cells decreased after heat stress for 40min, which was 0.82 ± 0.18 in the direct heat stress group and 1.70 ± 0 in the preheat adaptation group .52, preheat adaptation + heat stress group was 1.41 ± 0.16. Conclusion NIH-3T3 cells pre-heat treatment to determine the time point of stress protection, established a model of cell stress adaptation; preliminary confirmed HSP90 in the model of protective effect.