14-3-3η通过抑制内质网应激反应对H9c2心肌细胞的保护作用

摘要

Objective To study the effectof 14-3-3Ƞ on isoprenaline (ISO) induced H9c2 cardiomyocytes injury through inhibiting endoplasmic reticulum stress. Methods The recombinant expression vector pFLAG-14-3-3Ƞ was constructed and transfected into H9c2 cells. The cardiomyocytes injury model was established by using ISO. The H9c2 cells were randomly divided into four groups: control group, ISO group, ISO+pFLAG group, ISO+14-3-3Ƞ group. Cell apoptosis was detected by flow cytometry. Protein expression levels ofcaspase-3, caspase-12, glucose regulated protein 78 (GPR78), and CCAAT/enhancer binding protein homologous protein (CHOP) were examined by western blot. The-maleic dialdehyde (MDA) content and superoxide dismutase (SOD) activity was measured by the MDA and SOD detec-tion kit, respectively. Results After ISO stimulating H9c2 cardiomyocytes, compared with the control group, in ISO group, the cell apoptosis increased (P<0.05), the expression levels of caspase-3 and caspase-12 increased (P< 0.05), the content of MDA increased (P<0.05), the activity of SOD decreased (P<0.05), and the expression levels of GRP78 and CHOP increased (P<0.05). After reversed by pFLAG-14-3-3Ƞ transfection, dealed with ISO, compared with ISO group, in ISO+14-3-3Ƞ group, the expression levels of caspase-3 and caspase-12 decreased (P<0.05), the content of MDA decreased (P<0.05), the activity of SOD increased (P<0.05), and the expression levels of GRP78 and CHOP decreased (P<0.05). Conclusion The 14-3-3Ƞ over expression protects cardiomyocytes against ISO-induced cell in-jury via inhibiting endoplasmic reticulum stress.%目的：探讨14-3-3浊通过抑制内质网应激对异丙肾上腺素(ISO)诱导的H9c2心肌细胞损伤的保护作用。方法构建pFLAG-14-3-3Ƞ重组表达载体转染H9c2心肌细胞，使用ISO建立心肌细胞损伤模型。将培养的H9c2心肌细胞随机分为4组：正常对照组、ISO组、ISO+pFLAG组、ISO+14-3-3Ƞ组。流式细胞仪检测H9c2心肌细胞凋亡；Western blot检测 H9c2心肌细胞中半胱氨酸蛋白酶(caspase)-3、caspase-12、葡萄糖调节蛋白78(GPR78)、CCAAT/增强子结合蛋白同源蛋白(CHOP)的表达情况；采用试剂盒检测各组细胞丙二醛(MDA)含量和超氧化物歧化酶(SOD)的活性。结果 ISO刺激H9c2心肌细胞后，与正常对照组比较，ISO组细胞凋亡率显著增高(P<0.05)，caspase-3与caspase-12的表达升高(P<0.05)，MDA含量增高(P<0.05)，SOD活性降低(P<0.05)，内质网应激反应标志蛋白GPR78与CHOP的表达显著增高(P<0.05)。转染pFLAG-14-3-3Ƞ后再进行ISO处理，与ISO组比较，ISO+14-3-3Ƞ组心肌细胞的凋亡率显著降低(P<0.05)，caspase-3与caspase-12的表达明显减少(P<0.05)，MDA含量降低(P<0.05)，SOD活性增大(P<0.05)，GPR78与CHOP的表达显著降低(P<0.05)。结论14-3-3浊可以通过抑制内质网应激反应减轻ISO诱导的H9c2心肌细胞损伤。