摘要目的 观察热休克蛋白70 (HSP70)在心肌内质网应激(ERS)诱导的细胞凋亡中的作用,探讨HSP70调控内质网应激诱导细胞凋亡、分泌和钙信号转导的分子机制.方法 应用转基因技术和反义寡核苷酸导入技术诱导和抑制HSP70基因在SD乳鼠心肌细胞的表达,建立细胞培养和缺氧模型,测定细胞凋亡率,提取内质网测定HSP70对ERS导致的内质网内外游离钙、活性氧(ROS)含量的变化及内质网钙三磷酸腺苷酶(SERCA)和钙敏感受体(CaSR)的表达.结果 空白组早期细胞凋亡率为(9.28±0.83)％、晚期细胞凋亡率为(28.60 ±1.83)％、ROS生成率为80.0％,HSP70基因转染组早期细胞凋亡率为(4.30±0.72)％、晚期细胞凋亡率为(11.6±1.66)％、ROS生成率为38.5％,HSP70反义寡核苷酸处理组早期细胞凋亡率为(29.3±2.54)％、晚期细胞凋亡率为(34.4±2.43)％、ROS生成率为77.0％,表明HSP70显著抑制细胞凋亡和ROS的产生;HSP70基因转染组CaSR蛋白表达水平明显小于HSP70反义寡核苷酸处理组,而SERCA差异无统计学意义,这表明HSP70显著抑制CaSR的表达,但并不影响SERCA的表达.结论 HSP70通过抑制Ca2电流从而抑制CaSR的表达,并且在ROS诱导的Ca2电流中起重要作用.

abstractsObjective To observe the role of heat shock protein 70 (HSP70) in endoplasmic reticulum (ER) stress-induced cell apoptosis,and to investigate the molecular mechanism underlying the HSP70 regulating ER stress induced-apoptosis,secretion and calcium signal transduction.The fidings provide new intervention targets and theoretical basis of subcellular molecular application for myocardial protection.Methods We used SD neonatal rat cardiomyocytes to establish a hypoxia cell model,and then we further detected the role of HSP70 in cell apoptosis,production of reactive oxygen species (ROS),expression of calcium sensing receptor (CaSR) and sarco endoplasmic reticulum Ca2+-ATPase (SERCA) in ER caused by ER stress,by overexpression or downexpression of HSP70.Results The early apoptosis rate of the blank group [(9.28 ± 0.83) ％],late apoptosis rate [(28.60 ± 1.83) ％],ROS formation rate is 80.0％,HSP70 gene transfection group early apoptosis rate [(4.30 ± 0.72) ％],late apoptosis rate [(11.6 ± 1.66) ％],ROS formation rate is 38.5％,HSP70 treated with antisense oligonucleotides early apoptosis rate [(29.3 ± 2.54) ％],late apoptosis rate [(34.4 ± 2.43) ％],ROS formation rate of 77.0％,respectively,suggesting that HSP70 significantly inhibited the cell apoptosis and the production of ROS.The expression of CaSR in HSP70 gene transfection group was significantly lower than that in HSP70 antisense oligonucleotide group,but SERCA showed no significant difference,indicating that HSP70 significantly inhibited the expression of CaSR,but it had no influence on the expression of SERCA.Conclusion HSP70 inhibited the Ca2+ current via inhibiting the expression of CaSR,and the JNK pathway might play important roles in cell apoptosis and ROS-induced Ca2+ current.