CAJ | 학술논문

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-14-3-3?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+14-3-3?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-14-3-3?后再进行ISO处理，与ISO组比较，ISO+14-3-3?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。
목적：탐토14-3-3탁통과억제내질망응격대이병신상선소(ISO)유도적H9c2심기세포손상적보호작용。방법구건pFLAG-14-3-3?중조표체재체전염H9c2심기세포，사용ISO건립심기세포손상모형。장배양적H9c2심기세포수궤분위4조：정상대조조、ISO조、ISO+pFLAG조、ISO+14-3-3?조。류식세포의검측H9c2심기세포조망；Western blot검측 H9c2심기세포중반광안산단백매(caspase)-3、caspase-12、포도당조절단백78(GPR78)、CCAAT/증강자결합단백동원단백(CHOP)적표체정황；채용시제합검측각조세포병이철(MDA)함량화초양화물기화매(SOD)적활성。결과 ISO자격H9c2심기세포후，여정상대조조비교，ISO조세포조망솔현저증고(P<0.05)，caspase-3여caspase-12적표체승고(P<0.05)，MDA함량증고(P<0.05)，SOD활성강저(P<0.05)，내질망응격반응표지단백GPR78여CHOP적표체현저증고(P<0.05)。전염pFLAG-14-3-3?후재진행ISO처리，여ISO조비교，ISO+14-3-3?조심기세포적조망솔현저강저(P<0.05)，caspase-3여caspase-12적표체명현감소(P<0.05)，MDA함량강저(P<0.05)，SOD활성증대(P<0.05)，GPR78여CHOP적표체현저강저(P<0.05)。결론14-3-3탁가이통과억제내질망응격반응감경ISO유도적H9c2심기세포손상。
Objective To study the effectof 14-3-3? on isoprenaline (ISO) induced H9c2 cardiomyocytes injury through inhibiting endoplasmic reticulum stress. Methods The recombinant expression vector pFLAG-14-3-3? 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+14-3-3? 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-14-3-3? transfection, dealed with ISO, compared with ISO group, in ISO+14-3-3? 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 14-3-3? over expression protects cardiomyocytes against ISO-induced cell in-jury via inhibiting endoplasmic reticulum stress.