CAJ | 학술논문

目的探讨心肌特异性高表达热休克蛋白27(Hsp27)对内毒素血症所致心功能不全的影响,并初步探讨其机制.方法所有实验均在南京医科大学第一附属医院老年医学科研究室和南京大学模式动物研究所完成.(1)心肌特异性高表达Hsp27转基因鼠(Hsp27Tg)基因型和表达鉴定:分别采用PCR法和western blot法;(2)心功能测定:HspZ7 Tg和野生型(WT)对照鼠均被腹腔注射内毒素(LPS,10 mg/kg),24 h后以超声心动图测定心功能,n=6/组;(3)死亡率:Hsp27Tg和WT鼠腹腔注射内毒素(LPS,20 mg/kg),密切监测70 h内的累积死亡率,n=37/WT,n=27/Hsp27Tg;(4)NF-kB活性测定:心肌组织样本采用凝胶迁移电泳法,细胞样本采用双报告基因法,n=4/组;(5)多组间的两两比较采用ANOVA和Seheffe检验,生存曲线分析采用log-rank检验,P＜0.05设为相差显著性界值.结果 (1)Hsp27 Tg小鼠心肌有丰富的Hsp27表达,WT则没有;(2)LPS使WT和Hsp27 Tg鼠心功能均显著下降,但与WT比较,Hsp27 Tg的心功能显著改善(P＜0.01或P＜0.05),其中EF增加27.3%.FS增加37.1%;(3)至LPS注射后70 h,WT和Hsp27Tg小鼠死亡率分别为37.84%和11.11%,与WT鼠比较,Hsp27TG生存率显著提高(P＜0.05);(4)Hsp27显著抑制LPS诱导的NF-kB激活(P＜0.01或P＜0.05).结论 Hsp27心肌特异性高表达显著抑制内毒素血症所致的心功能不全,改善生存率,其机制可能与Hsp27下调LPS诱导的NF-kB激活有关.
목적 탐토심기특이성고표체열휴극단백27(Hsp27)대내독소혈증소치심공능불전적영향,병초보탐토기궤제.방법 소유실험균재남경의과대학제일부속의원노년의학과연구실화남경대학모식동물연구소완성.(1)심기특이성고표체Hsp27전기인서(Hsp27Tg)기인형화표체감정:분별채용PCR법화western blot법;(2)심공능측정:HspZ7 Tg화야생형(WT)대조서균피복강주사내독소(LPS,10 mg/kg),24 h후이초성심동도측정심공능,n=6/조;(3)사망솔:Hsp27Tg화WT서복강주사내독소(LPS,20 mg/kg),밀절감측70 h내적루적사망솔,n=37/WT,n=27/Hsp27Tg;(4)NF-kB활성측정:심기조직양본채용응효천이전영법,세포양본채용쌍보고기인법,n=4/조;(5)다조간적량량비교채용ANOVA화Seheffe검험,생존곡선분석채용log-rank검험,P＜0.05설위상차현저성계치.결과 (1)Hsp27 Tg소서심기유봉부적Hsp27표체,WT칙몰유;(2)LPS사WT화Hsp27 Tg서심공능균현저하강,단여WT비교,Hsp27 Tg적심공능현저개선(P＜0.01혹P＜0.05),기중EF증가27.3%.FS증가37.1%;(3)지LPS주사후70 h,WT화Hsp27Tg소서사망솔분별위37.84%화11.11%,여WT서비교,Hsp27TG생존솔현저제고(P＜0.05);(4)Hsp27현저억제LPS유도적NF-kB격활(P＜0.01혹P＜0.05).결론 Hsp27심기특이성고표체현저억제내독소혈증소치적심공능불전,개선생존솔,기궤제가능여Hsp27하조LPS유도적NF-kB격활유관.
Objective To study the effects of heat shock proetin 27(Hsp27)on the cardiac dysfunction induced by endotoxemia.Method All experiments were performed in the geriatric lab of the First Affiliated Hospital Of Nanjing Medical University,and in the Animal Model Center of Nanjing University.The genotyping of the transgenic mice with cardiac-specific overexpression of Hsp27(Hsp27 Tg)was assayed by PCR and the expression of Hsp27 was determined by western blot.Hsp27 Tg and its wild type littermates(WT)were intraperitoneally injected with LPS(10 mg/kg),and 24 hours later,cardiac function was measured by echocardiography(n=6/group).The accumulated mice mortality was recorded within 70 hous after intraperitoneal injection of LPS(20mg/kg)(n=37/WT,n=27/Hsp27Tg).The NF-kB activity for cardiac-tissue samples was analyzed by electrophoretic mobility shift assay(ENSA)and for cell culture samples by dual-reporter gene assay(n=4/group).The comparison of multiple groups was performed by one-way analysis of variance(ANOVA),and comparison of two groups was performed by Scheffe-test.Survival curves were analyzed by the log-rank test.P＜0.05 wns considered to be significant.Results The high expression of Hsp27 exhibited in myocardium of Hsp27 Tg,whereas not in myocardium of WT.LPS significantly reduced the cardiac function both in Hsp27 Tg and WT.However,compaled with LPS-treated WT,cardiac function was more significantly improved as evidenced by the increases of EF by 27.33%and FS by 37.09%(P＜0.01 or P＜0.05).Seventy hours after LPS injection,the mortality was 11.11% in Hsp27 Tg and 37.84% in WT.Compared with WT,the survival rate of Hsp27 Tg significantly increased(P＜0.05).The NF-kB activation was significantly inhibited by Hsp27(P＜0.01 or P＜0.05).Conclusions The high cardiac-specific expression of Hsp27 significantly inhibits cardiac dysfunction induced by endotoxemia,and at the same time improve the survival rate.The mechanism may be connected with Hsp27 downregulating NF-kB-activation induced by LPS.