高糖对INS-1细胞损伤存在记忆效应

摘要

Objective To investigate the cellular memory of previous high glucose exposure in rat islet cell line (INS-1) and explore the possible mechanism. Methods INS-1 cells were exposed to a high glucose (33.3 mmol/L) culture for 48 h followed by further culture in the presence of 11.1 mmol/L glucose in the culture medium for 3 or 5 days. The levels of bax and caspase-3 Mrna were measured by real-time PCR, the production of reactive oxygen species (ROS) was assayed using the dihydroethidium probe, and the cell viability was detected by MTT assay. Results High glucose exposure of the cells for 48 h resulted in significantly increased ROS production and bax and caspase-3 Mrna expressions and lowered cell viability (P＜0.01). In cells cultured in 11.1 mmol/L glucose following previous high glucose exposure, the ROS production and bax and caspase-3 Mrna expressions still maintained the high levels (P< 0.05) while the cell viability remained significantly lower than the control cells (P<0.001). Conclusion High glucose causes persistent changes in cell viability and apoptosis-related gene expressions even after recovery of normoglycemia, the mechanism of which is probably related to increased ROS production.%目的 观察大鼠胰岛细胞对高糖损伤是否存在记忆效应,并对其机制进行初步探讨.方法 以INS-1(大鼠胰岛瘤细胞株)细胞作为研究对象,观察INS-1细胞经历48 h持续高糖(33.3 mmol/L)刺激后,纠正高糖状态,以11.1 mmol/L糖浓度继续培养,检测细胞活力,凋亡相关基因(bax,caspase-3)的mRNA水平和活性氧(ROS)水平的变化.INS-1细胞处理组分为:对照组(11.1 mmol/L葡萄糖×7 d);高糖组(33.3 mmol/L葡萄糖×2 d);3 d记忆组(33.3 mmol/L葡萄糖×2 d+11.1mmol/L葡萄糖×3 d);5 d记忆组(33.3 mmol/L葡萄糖×2 d+11.1 mmol/L葡萄糖×5 d).Real-Time PCR检测bax,caspase-3的mRNA水平,Dihydrocthidium(DHE探针)检测ROS水平,MTT法测定细胞活力.结果 高糖组细胞活力较对照组显著降低(P＜0.001).而ROS水平,bax,Caspase-3的mRNA水平较对照组显著增高(P＜0.05).在3 d记忆组及5 d记忆组,细胞活力较对照组显著降低(P＜0.001),ROS水平,bax,Caaspase-3的mRNA水平较对照组显著增高(P＜0.05).结论 高糖对INS-1细胞的损伤存在记忆效应,主要表现为细胞活力的持续降低,促凋亡基因的持续高表达.ROS水平的持续升高可能与记忆效应产生有密切关系.