艾塞那肽对高糖所致新生乳鼠心肌细胞钙信号紊乱的调控作用

Regulating effect of exenatide on disorder of calcium signal in neonatal rat cardiomyocytes induced by high glucose

目的观察GLP-1类似物艾塞那肽(EX-4)对高糖所致新生乳鼠心肌细胞钙信号紊乱的调控作用,探讨GLP-1的心脏保护机制。方法分离新生乳鼠心肌细胞,培养24 h后随机分为对照组、高糖组和高糖+EX-4组,分别加入25 mmol/L甘露醇、25 mmol/L葡萄糖、25 mmol/L葡萄糖+10 nmol/L EX-4,继续培养24 h。使用激光共聚焦显微技术检测各组心肌细胞自发钙火花发放频率、场刺激钙瞬变幅度、钙库容量;使用膜片钳技术记录L型钙通道电流。结果对照组、高糖组、高糖+EX-4组的心肌细胞自发钙火花发放频率分别为(1.35±0.07)、(3.86±0.27)、(1.99±0.15)个/(μm·s),场刺激钙瞬变幅度钙瞬变幅度分别为3.95±0.08、3.34±0.07、4.09±0.12,钙库容量分别为4.95±0.18、3.26±0.09、4.59±0.15,10 m V时的L型钙通道电流密度分别为-12.46±0.58、-11.28±0.55、-13.09±0.64,高糖组心肌细胞钙火花发放频率、钙瞬变幅度较对照组升高,钙库容量、电流密度较对照组降低(P<0.05或<0.01),高糖+EX-4组钙火花发放频率、钙瞬变幅度较高糖组降低,钙库容量、电流密度较高糖组升高(P<0.05或<0.01),与对照组比较无统计学意义。结论 EX-4能够增强心肌细胞L型钙通道电流,减少肌浆网钙漏流,从而有效纠正高糖所致心肌细胞钙调控紊乱,降低高糖所致心肌损伤。

Objective To investigate the regulating effect of GLP-1 analogue exenatide（exenatide-4,EX-4） on the disorder of calcium signal in neonatal rat cardiomyocytes induced by high glucose and the cardioprotective mechanism.Methods The neonatal rat cardiomyocytes were isolated and cultured for 24 h,and then were randomly divided into 3groups：the control group,high glucose group and high glucose＋EX-4 group which were separately treated with 25 mmol/L mannitol,25 mmol/L high glucose and 25 mmol/L high glucose＋10 mmol/L EX-4.The frequency of spontaneous Ca2＋spark in cadiomyocyte,field-stimulated calcium transient amplitude and calcium storage capacity were detected by laser scanning confocal microscopy.L-type calcium channel current was recorded by using patch-clamp technique.Results The frequencies of spontaneous Ca^（2＋） spark in cadiomyocytes of the control group,high glucose group and high glucose＋EX-4group were（1.35±0.07）,（3.86±0.27） and（1.99±0.15）/（μm·s）,respectively.Field-stimulated calcium transient amplitude was 3.95±0.08,3.34±0.07 and 4.09±0.12.The calcium storage capacities were 4.95±0.18,3.26±0.09 and 4.59±0.15,respectively.L-type calcium channel current density at 10 m V were-12.46±0.58,-11.28±0.55,and-13.09±0.64.In the high glucose group,the frequency of Ca^2＋spark and the Ca^2＋ transient amplitude were higher than those of the control group（P〈0.05 or P〈0.01）,and calcium storage capacity and L type calcium current density were lower than those in the control group（P〈0.05 or P〈0.01）.In the high glucose＋EX-4 group,the frequency of Ca^2＋ spark and the amplitude of Ca^2＋ transient were higher than those of the high glucose group（P〈0.05 or P〈0.01）,and calcium storage capacity and L type calcium current density were higher than those of the high glucose group（P〈0.05 or P〈0.01）,while there was no statistically significant difference as compared with those of the control group.Conclusions EX-4 can increase L-type Ca^2＋ current in cardiomyocytes,and reduce the Ca^2＋leakage current in the plasma,thus effectively correcting the disorder of Ca^2＋ regulation in cardiomyocytes induced by high glucose.Therefore,EX-4has good myocardial protective effect on patients with diabetes mellitus.