摘要
目的 探讨细胞毒T淋巴细胞 (CTL)经Fas/FasL介导的细胞凋亡在柯萨奇病毒B3(CVB3)病毒性心肌炎 (VM)小鼠发病中的作用。方法 采用电镜技术及原位末端标记法 (TUNEL)检测小鼠心肌组织的细胞凋亡 ,同时采用免疫组化、逆转录 聚合酶链反应 (RT PCR)及原位杂交 3种方法检测心肌炎小鼠不同时期心肌组织中Fas与FasL基因转录与蛋白的表达。结果 (1)各期病鼠心肌组织中的TUNEL阳性的心肌细胞凋亡检出率为 79% ,中、重度VM小鼠的检出率明显高于轻度VM小鼠 (P <0 .0 5 )。平均心肌细胞凋亡百分率为 (8± 6 ) %。感染后第 7～ 14天心肌细胞凋亡百分率明显高于第 2 1～ 2 8天 (P <0 .0 5 )。(2 )感染后第 7～ 14天 ,病鼠心肌组织中 ,主要表达于心肌细胞的FasmRNA及蛋白和主要表达于浸润淋巴细胞的FasLmRNA及蛋白均明显增强 (P <0 .0 1)。FasL蛋白表达水平与心肌病变积分呈正相关 (r=0 .89,P <0 .0 1)。
Objective The cytotoxic T lymphocyte (CTL) mediated cytotoxic reaction is one of the important pathogenesis of virus myocarditis (VM). CTL can induce apoptosis in cardiac muscle cells through Fas/Fas ligand (FasL) pathway. Apoptosis was proved exist in VM. The study, therefore, investigated the role of apoptosis and Fas/ FasL in the development of VM. Methods One hundred and twenty five Balb/c mice were included in the experiment. Twenty mice in experimental group inoculated with 10 9 TCID Coxackie virus B 3(CVB 3) and five mice in control group injected with saline were sacrificed 7, 10, 14, 21 and 28 days post inoculation (p.i.). Light microscopy, electronic microscopy and terminal transferase mediated dUTP biotin nick end labeling (TUNEL) assays were used to detect the inflammation, necrosis and apoptosis in myocardium. The expressions of Fas and FasL protein in myocardium were determined by immunohistochemistry. Fas mRNA and FasL mRNA were analyzed by reverse transcription polymerase chain reaction (RT PCR) and in situ hybridization. Results (1) The incidence of VM was 86%. The typical myocardial lesions including myocardial necrosis and lymphocyte infiltration at the 7th day (p.i.) were observed, much more obviously at the 10th to 14th day, and recovered gradually from the 21th to 28th day (p.i.). The mice in control group did not show any sign of inflammation. (2)Apoptosis myocytes were seen in five of ten myocarditis mice by electronic microscopy. Apoptosis cells including myocytes, endothelial cells and infiltrating cells appeared in the myocardium of myocarditis mice. The detection rate of TUNEL positive myocytes was 79%. The ratio of TUNEL positive myocytes was much higher in mice with moderate to severe VM than mice with mild VM ( P <0.05). TUNEL positive myocytes were mainly distributed in tela subendocardiaca, subepicardial layer and around the inflammatory foci. The average percent of apoptosis myocytes was (8±6)%. The percent of apoptosis myocytes increased significantly after the infection 7 to 14 days than 21 to 28 days post inoculation ( P <0.05). (3) In experimental group, Fas mRNA and protein expressed mainly in myocytes, and FasL mRNA and protein expressed mainly in infiltrating lymphocytes and increased remarkably from 7 to 14 days compared with control group( P <0.01). The dynamic changes of FasL protein expression showed a significantly positive correlation with the changes of myocardial histopathologic scores ( r = 0.89, P <0.01). Conclusion The cytotoxic T lymphocyte mediated apoptosis in myocardium through Fas/FasL pathway might play an important role in the development of VM.
出处
《中华儿科杂志》
CAS
CSCD
北大核心
2000年第7期405-408,共4页
Chinese Journal of Pediatrics
