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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 regulation of GLP-1 analogues exendin (EX-4) on the disturbance of calcium signaling in neonatal neonatal rat cardiomyocytes induced by high glucose and to explore the cardioprotective mechanism of GLP-1. Methods Newborn neonatal rat cardiomyocytes were isolated and cultured 24 hours later and were randomly divided into control group, high glucose group and high glucose + EX-4 group. 25 mmol / L mannitol, 25 mmol / L glucose and 25 mmol / L glucose +10 nmol / L EX-4, continue to train for 24 h. Laser confocal microscopy was used to detect the distribution frequency of spontaneous calcium sparks, field transient amplitude of calcium stimulation and calcium storage capacity in each group. L-type calcium current was recorded using patch-clamp technique. Results The frequencies of spontaneous calcium sparks in control group, high glucose group and high glucose + EX-4 group were (1.35 ± 0.07), (3.86 ± 0.27) and (1.99 ± 0.15) / (μm · s), respectively The amplitude of calcium transients in field stimulated calcium transients were 3.95 ± 0.08, 3.34 ± 0.07 and 4.09 ± 0.12, respectively. The L-type calcium channels were 4.95 ± 0.18, 3.26 ± 0.09, 4.59 ± 0.15 and 10 mV respectively The current densities were -12.46 ± 0.58, -11.28 ± 0.55 and -13.09 ± 0.64, respectively. The frequency of calcium spark release and the amplitude of calcium transient in high glucose group were higher than those in control group, and the volume of calcium stores and current densities were lower than those in control group (P <0.05 or <0.01). The firing rate of calcium sparks in high glucose + EX-4 group was significantly lower than that in high glucose group (P <0.05 or <0.01) , No significant difference with the control group. Conclusion EX-4 can enhance the L-type calcium channel current in cardiac myocytes and decrease the Ca leakage in sarcoplasmic reticulum, so as to effectively correct the disturbance of calcium regulation in cardiomyocytes induced by high glucose and reduce the myocardial injury induced by high glucose.