CAJ | 학술논문

目的探讨重组腺相关病毒(rAAV)介导大鼠血红素氧合酶-1(rHO-1)基因转染对心肌缺血/再灌注(I/R)损伤大鼠心肌细胞凋亡的影响.方法 95只体重225～250 g的雄性SD大鼠随机分为假手术组(SO组,n=8)、生理盐水组(NS组,,n=29)、rAAV-荧光蛋白组(rAAV-EGFP组,n=29)及rAAV-rHO-1组(n=29).NS组、rAAV-EGFP组和rAAV-rHO-1组分别于大鼠左心室前后壁共取4点分别注入生理盐水、rAAV-EGFP或rAAV-rHO-1病毒液共600 μl.在基因转染后3个月,各组处死3只大鼠,取注射部位心肌,荧光显微镜下观察荧光蛋白的表达并计算转染率;用免疫组化染色和逆转录-聚合酶链反应(RT-PCR)检测HO一1的蛋白和mRNA表达.采用结扎左冠状动脉前降支30 min、再灌注120 min建立心肌I/R模型;成模后处死大鼠测定其心肌梗死面及心肌细胞凋亡指数(AI),光镜下观察心肌组织病理学改变.结果荧光显微镜下仅rAAV-EGFP组可见心肌有荧光蛋白表达,转染率为(53.5±2.0)%.与SO组比较,NS组、rAAV-EGFP组和rAAV-rHO-1组Al增加(P均<0.01),与NS组和rAAV-EGFP组比较,rAAV-rHO-1组HO-1的蛋白及mRNA表达增加,心肌梗死面积减小,A1减少(P均<0.01);NS组及rAAV-EGFP组心肌组织病理学损伤较SO组和rAAV-rHO-1组为重.NS组与rAAV-EGFP组间上述指标比较无统计学意义.结论 rAAV介导的rHO-1基因转染大鼠心肌细胞后,能够显著减少心肌细胞凋亡,从而减轻心肌I/R损伤.
목적 탐토중조선상관병독(rAAV)개도대서혈홍소양합매-1(rHO-1)기인전염대심기결혈/재관주(I/R)손상대서심기세포조망적영향.방법 95지체중225～250 g적웅성SD대서수궤분위가수술조(SO조,n=8)、생리염수조(NS조,,n=29)、rAAV-형광단백조(rAAV-EGFP조,n=29)급rAAV-rHO-1조(n=29).NS조、rAAV-EGFP조화rAAV-rHO-1조분별우대서좌심실전후벽공취4점분별주입생리염수、rAAV-EGFP혹rAAV-rHO-1병독액공600 μl.재기인전염후3개월,각조처사3지대서,취주사부위심기,형광현미경하관찰형광단백적표체병계산전염솔;용면역조화염색화역전록-취합매련반응(RT-PCR)검측HO일1적단백화mRNA표체.채용결찰좌관상동맥전강지30 min、재관주120 min건립심기I/R모형;성모후처사대서측정기심기경사면급심기세포조망지수(AI),광경하관찰심기조직병이학개변.결과 형광현미경하부rAAV-EGFP조가견심기유형광단백표체,전염솔위(53.5±2.0)%.여SO조비교,NS조、rAAV-EGFP조화rAAV-rHO-1조Al증가(P균<0.01),여NS조화rAAV-EGFP조비교,rAAV-rHO-1조HO-1적단백급mRNA표체증가,심기경사면적감소,A1감소(P균<0.01);NS조급rAAV-EGFP조심기조직병이학손상교SO조화rAAV-rHO-1조위중.NS조여rAAV-EGFP조간상술지표비교무통계학의의.결론 rAAV개도적rHO-1기인전염대서심기세포후,능구현저감소심기세포조망,종이감경심기I/R손상.
Objective To investigate the effect of rat hemeoxygenase-1 (rHO-1) gene carried by recombinant adeno-assoeiated virus (rAAV) on myocardial isehemia/reperfusion (I/R) injury in rats. Methods Ninety-five healthy male Sprague-Dawley (SD) rats weighing 225～250 g were randomly divided into four groups: sham operation group (Ⅰ, n=8); normal saline group (Ⅱ , n=29); rAAV-EGFP (enhan ced green fluorescent protein)group (Ⅱ, n=29)and rAAV-rHO-1 group (Ⅳ, n=29). In Ⅱ ,III and Ⅳ groups,600μl of normal saline,rAAV-EGFP or rAAV-rHO-1 was injected intramyocardially at four sites on the anterior and posterior walls of left ventricle. After 3 months, 3 animals in each group were sacrificed. EGFP-expression in heart sections was observed under fluorescence microscope. The expression of HO-1 in the injected myocardium was detected by immunohistoehemistry and reverse transcription-polymerase chain reaction (RT-PCR). The remaining animals in the four groups were anesthetized, traeheostomized and mechanically ventilated. I/R of myocardium was producing by blocking the left anterior descending branch of coronary artery (LAD) for 30 minutes followed by 120 minutes reperfusion. After the successful reproduction of the model, the animals were killed and their hearts were harvested for determination of myocardial infarct size, apoptotic index (AI), and pathology changes in myocardial tissue. Results The expression of EGFP was detected in group Ⅲ only, and transfeetion efficiency was (53.5±2.0)%. AI was significantly higher in groupⅡ, group Ⅲ and group Ⅳ than in group I (all P<0. 01). The expression of HO-1 mRNA and protein was significantly higher, and the infarct size and AI, were significantly lower in group Ⅳ than in group Ⅱ and group Ⅲ (all P<0. 01). The degree of damage to myocardial tissue was significantly severer in group Ⅱ and group Ⅲ than in group Ⅰ and group Ⅳ. There was no significant dift'erenee between group Ⅱand group Ⅲ. Conclusion rAAV-mediated rHO-1 gene transfeetion may attenuate myocardium I/R injury by inhibiting apoptosis of cardiomyocyte in rats.