4,4＇-二异硫氰基茋-2,2＇-二磺酸对高糖诱导心肌细胞损伤的保护作用

Protective effect of 4,4’-diisothiocyanostilbene-2,2’-disulfonic acid on high glucose-induced cardiomyocyte injury

目的：探讨高糖诱导心肌细胞损伤过程中氯离子浓度的变化以及氯离子通道阻断剂4,4’-二异硫氰基茋-2,2’-二磺酸（DIDS）对受损心肌细胞的作用。方法原代培养乳鼠心肌细胞，应用葡萄糖浓度为33mmol/L DMEM培养基，作用72h，构建心肌细胞损伤模型。实验分为对照组、高糖组、高糖＋DIDS组和DIDS组。用MTT法检测细胞存活率，流式细胞仪检测细胞凋亡率，氯离子荧光探针（MQAE）检测细胞内氯离子浓度。结果高糖作用下的心肌细胞存活率呈浓度及时间依赖性降低（P＜0.05），流式细胞仪检测显示受损心肌细胞以凋亡性损伤为主。MQAE检测结果显示，与对照组比较，高糖组细胞内氯离子浓度显著下降（P＜0.05）。心肌细胞存活率及细胞内氯离子浓度，DIDS组与对照组比较差异均无统计学意义（P＞0.05）。与高糖组比较，高糖＋DIDS组心肌细胞存活率显著升高，细胞内氯离子浓度下降明显减少，差异均有统计学意义（P＜0.05）。结论氯离子通道参与了高糖诱导的心肌细胞损伤过程，DIDS可在一定程度上减轻以凋亡为主的心肌细胞损伤。

Objective To investigate the change in chloride ion （Cl-） concentration in high glucose-induced cardiomyocyte injury and the role of 4,4’-diisothiocyanostilbene-2,2’-disulfonic acid （DIDS）, a chloride channel blocker, in the injury. Methods Primarily cultured neonatal rat cardiomyocytes were exposed to glucose （33mmol/L） for 72h to establish the cell model of high glucose-induced injury. The cardiomyocytes were randomly divided into 4 groups：control group, high glucose group, high glucose＋DIDS group and DIDS group. MTT assay was applied to detect cell viability, and flow cytometry was used to measure cell viability and apoptosis. N-（Ethoxycarbonylmethyl）-6-methoxyquinolinium bromide （MQAE） was used to examine the intracellular chloride concentration. Results High glucose decreased the cardiomyocyte viability in a dose- and time-dependent manner （P〈0.05）. Cardiomyocyte injury was mainly in the form of apoptosis. Compared to control group, high glucose resulted in a significant decrease in intracellular Cl-in the cardiomyocytes （P〈0.05）. There was no significant difference in cardiomyocyte viability and intracellular Cl-between the DIDS treated cells and control cells （P〉0.05）. While, DIDS treatment improved the cardiomyocyte viability and attenuated the decrease in intracellular Cl- in cardiomyocytes after high glucose induction （P〈0.05）. Conclusion Cl- is involved in the high glucose-induced cardiomyocyte injury. DIDS attenuates the injury which mainly presents as apoptosis.