CAJ | 학술논문

目的:观察外源性精胺对缺血-再灌注损伤诱导的乳鼠心肌细胞凋亡的影响并探讨其可能机制.方法:原代培养的乳鼠心肌细胞缺氧、无血清孵育2 h,再复氧和复血清培养24 h以模拟心肌缺血-再灌注损伤,并于再灌注期给予10 μmol/L的外源性精胺.利用流式细胞仪检测心肌细胞的凋亡率;电镜观察细胞超微结构变化;应用RT-PCR、Western blotting方法和免疫荧光的方法观察Fas、FasL mRNA的转录和蛋白的表达.结果:模拟缺血-再灌注后心肌细胞凋亡率为27.4%±1.8%,明显高于对照组(5.7%±0.3%),细胞超微结构出现凋亡、坏死等改变,Fas、FasL的转录和蛋白表达均明显上调,与对照组比,Fas mRNA和FasL mRNA转录分别增加了2.2倍和2.4倍(P<0.01);Fas和FasL 的蛋白表达分别增加了1.7倍和1.9倍(P<0.01);10 μmol/L的外源性精胺干预后心肌细胞凋亡率降低为21.7%±1.3%,Fas、FasL的转录和蛋白表达也明显被抑制(与单纯模拟缺血-再灌注组比,P<0.05或P<0.01).结论:外源性精胺可通过抑制Fas死亡受体途径而减轻模拟缺血-再灌注诱导的心肌细胞凋亡.
목적:관찰외원성정알대결혈-재관주손상유도적유서심기세포조망적영향병탐토기가능궤제.방법:원대배양적유서심기세포결양、무혈청부육2 h,재복양화복혈청배양24 h이모의심기결혈-재관주손상,병우재관주기급여10 μmol/L적외원성정알.이용류식세포의검측심기세포적조망솔;전경관찰세포초미결구변화;응용RT-PCR、Western blotting방법화면역형광적방법관찰Fas、FasL mRNA적전록화단백적표체.결과:모의결혈-재관주후심기세포조망솔위27.4%±1.8%,명현고우대조조(5.7%±0.3%),세포초미결구출현조망、배사등개변,Fas、FasL적전록화단백표체균명현상조,여대조조비,Fas mRNA화FasL mRNA전록분별증가료2.2배화2.4배(P<0.01);Fas화FasL 적단백표체분별증가료1.7배화1.9배(P<0.01);10 μmol/L적외원성정알간예후심기세포조망솔강저위21.7%±1.3%,Fas、FasL적전록화단백표체야명현피억제(여단순모의결혈-재관주조비,P<0.05혹P<0.01).결론:외원성정알가통과억제Fas사망수체도경이감경모의결혈-재관주유도적심기세포조망.
AIM: To explore the effects and possible mechanism of exogenous spermine on the apoptosis of primary cultured neonatal cardiomyocytes induced by simulated ischemia-reperfusion (I/R) injury. METHODS: To establish a model of simulated I/R, the primary cultured neonatal rat cardiomyocytes were incubated in ischemia-mimetic solution (under the conditions of hypoxia plus serum deprivation) for 2 h, and re-incubated the cells in normal culture medium for 24 h. The apoptotic cell death was assayed by flow cytometry. The morphological alterations of the cells were observed under transmission electron microscope. The transcription and expression of Fas and FasL were determined by the methods of RT-PCR, Western blotting and immunofluorescence. RESULTS: The cells exposed to I/R underwent significant apoptosis, and the percentage of apoptotic cells was 27.4%±1.8%, much higher than that in normal group (5.7%±0.3%). In I/R group the evident histopathological changes were observed and the myocardial transcription and expression of Fas and FasL were significantly upregulated. Compared to normal group, mRNA expression of Fas and FasL increased 2.2 folds and 2.4 folds, respectively, and their proteins increased 1.7 folds and 1.9 folds at 24 h of reperfusion respectively (P<0.01). Pretreatment with 10 μmol/L spermine significantly inhibited apoptosis of I/R injured cells, and the percentage of apoptotic cells was 21.7%±1.3% (P<0.01, as compared to I/P group). Spermine also suppressed the expression of Fas and FasL significantly (P<0.05 or P<0.01, as compared to I/P group). CONCLUSION: Spermine plays anti-apoptotic effect on the cultured neonatal myocardial cells under the condition of I/R injury by suppressing the expression of Fas/FasL.