CAJ | 학술논문

目的观察丙酮酸乙酯(EP)对SD大鼠乳鼠心肌细胞缺氧复氧(SIR)损伤的影响,并探讨内质网应激(ERS)在这一过程中的作用.方法乳鼠心肌细胞经EP(5或10 mmol/L)处理2h后,接受模拟缺氧复氧(SIR)损伤(缺氧2h,复氧4h),噻唑蓝比色(MTT)法检测细胞活力,专用试剂盒检测细胞培养液中丙二醛(MDA)和超氧化物歧化酶(SOD)含量,Western blot检测ERS相关蛋白葡萄糖调节蛋白78(GRP78)、CCAAT/增强子结合蛋白同源蛋白(CHOP)表达变化.随后,EP处理时加入ERS激动剂毒胡萝卜素(THA),观察其对EP抗心肌细胞SIR损伤的影响.结果 5、10 mmol/L EP均可以显著升高心肌细胞SIR后细胞活力(分别增加到0.424±0.016和0.517±0.020),降低培养液中MDA水平[分别降低到(16.17±1.75)和(11.45±1.20) μmol/L],但SOD水平升高[分别升高到(58.61 ±4.70)和(72.60 ±4.35) U/L,与SIR组比较,P＜0.05].SIR后细胞中GRP78和CHOP表达水平升高(与对照组比较,P＜0.05),EP还可以逆转SIR上调的GRP78和CHOP表达水平(与SIR组比较,P＜0.05).进一步研究结果显示,ERS激动剂THA可以逆转EP对心肌细胞的保护作用,降低心肌细胞活力,增加GRP78和CHOP表达水平(与EP+ SIR组比较,P＜0.05).结论 EP可显著减轻乳鼠心肌细胞的SIR损伤,其机制与抑制异常激活的ERS反应有关.
목적 관찰병동산을지(EP)대SD대서유서심기세포결양복양(SIR)손상적영향,병탐토내질망응격(ERS)재저일과정중적작용.방법 유서심기세포경EP(5혹10 mmol/L)처리2h후,접수모의결양복양(SIR)손상(결양2h,복양4h),새서람비색(MTT)법검측세포활력,전용시제합검측세포배양액중병이철(MDA)화초양화물기화매(SOD)함량,Western blot검측ERS상관단백포도당조절단백78(GRP78)、CCAAT/증강자결합단백동원단백(CHOP)표체변화.수후,EP처리시가입ERS격동제독호라복소(THA),관찰기대EP항심기세포SIR손상적영향.결과 5、10 mmol/L EP균가이현저승고심기세포SIR후세포활력(분별증가도0.424±0.016화0.517±0.020),강저배양액중MDA수평[분별강저도(16.17±1.75)화(11.45±1.20) μmol/L],단SOD수평승고[분별승고도(58.61 ±4.70)화(72.60 ±4.35) U/L,여SIR조비교,P＜0.05].SIR후세포중GRP78화CHOP표체수평승고(여대조조비교,P＜0.05),EP환가이역전SIR상조적GRP78화CHOP표체수평(여SIR조비교,P＜0.05).진일보연구결과현시,ERS격동제THA가이역전EP대심기세포적보호작용,강저심기세포활력,증가GRP78화CHOP표체수평(여EP+ SIR조비교,P＜0.05).결론 EP가현저감경유서심기세포적SIR손상,기궤제여억제이상격활적ERS반응유관.
Objective To explore the effect of ethyl pyruvate (EP) on the simulated ischemia reperfusion (SIR) injury of neonatal rat cardiomyocyteand the role of endoplasmic reticulum stress (ERS)in this procedure.Methods The cultured neonatal rat cardiomyocytes,pretreated with EP (5 or 10 mmol/L) for 2 h,were subjected to SIR injury (hypoxia for 2 h,reoxygenation for4 h).Methyl thiazolyl tetrazolium (MTT) method was used to detect the cell viability,special kits were used to detected the levels of malondialdehyde (MDA) and superoxide dismutase (SOD) in culture medium.Western blotting was used to detect the expression of ERS related molecules glucose regulated protein 78 (GRP78) and CCAAT/enhancer-binding protein homologous protein (CHOP).Further,EP treatment was added with the ERS activator thapsigargin (THA),the effect of THA on the protection of EP against SIR was observed.Results Both 5 and 10 mmol/LEP treatment dramatically increased the cell viability (0.424 ±0.016 and 0.517 ± 0.020),downregulated the increased MDA level [(16.17 ± 1.75) and (11.45 ± 1.20) μmol/L] and decreased SOD level [(58.61 ± 4.70) and (72.60 ± 4.35) U/L] in the culture medium (vs.the SIR group,P ＜ 0.05).Compared with the control group,the level of GRP78 and CHOP increased significantly in the SIR group (P ＜ 0.05),while EP treatment significantly decreased the level of GRP78 and CHOP (vs.the SIR group,P ＜ 0.05).Further,the ERSactivator THA reversed the protection of EP,decreased the cell viability (vs.the EP + SIR group,P ＜ 0.05),upreguleted the expression of GRP78 and CHOP (vs.the EP + SIR group,P ＜ 0.05).Conclusion EP treatment can protect against SIR injury of neonatal rat cardiomyocyte,this effect is associated with inhibition of ERS.