CD4^＋T细胞活化诱导细胞凋亡在原发性胆汁性肝硬化小鼠模型中的作用研究  

Defective activation induced cell death of CD4^＋T cells in primary biliary cirrhosis murine model

作　　者：蒋廷旺[1] 邓安梅[1] 吴传勇[1] 陈波[1] 周晔[1] 钱琤[1] 谷明莉[1] 陈燕[1] 仲人前[1] 

机构地区：[1]第二军医大学附属长征医院实验诊断科全军临床免疫中心,上海200003

出　　处：《中华微生物学和免疫学杂志》2008年第5期431-434,共4页Chinese Journal of Microbiology and Immunology

基　　金：国家863计划项目（2006AA022496）;国家自然科学基金（30671840,30772017）;上海市优秀学科带头人基金（07XD14013）;上海市优秀青年医学人才培养计划、长征医院“重大临床研究”和“优秀青年骨干人才培养计划”资助

摘　　要：目的研究CD4^＋T细胞活化诱导细胞凋亡（activation induced cell death，AICD）在poly I：C诱导的原发性胆汁性肝硬化（研marybiliarycirrhosis，PBC）小鼠模型中的作用。方法30只C57BL／6雌性小鼠随机分为模型组和对照组，模型组小鼠腹腔注射poly I：C5mg／kg，对照组小鼠注射等体积无菌PBS，16周后通过测定血清抗线粒体抗体（antimitochondrial antibody,AMA）、碱性磷酸酶（alkali phosphatase，ALP）及肝脏HE染色验证模型。磁珠分离小鼠脾脏CD4^＋T细胞，分别以ConA和anti—CD3体外诱导细胞凋亡；实时荧光定量PCR测定T细胞中凋亡相关基因Fas、FasL和TRAIL（tumor necrosis factor-related apoptosis—inducing ligand）的表达；Westernblot检测CD4^＋T细胞中抗凋亡基因Bcl-2的表达。结果poly I：C注射16周后模型组小鼠血清AMA均为阳性，同时肝组织汇管区出现不同程度的炎性细胞浸润，而对照组AMA均为阴性，肝组织未出现明显病变。模型组小鼠血清ALP[（110．4±18．3）U／L]显著高于对照组[（52．2±15．4）U／L]，P〈0．001；模型组小鼠脾CD4^＋T细胞AICD显著低于对照组（P〈0．001），同时定量PCR结果表明：FasLmRNA水平较对照组有所降低（P〈0．05），而两组Fas水平差异无统计学意义（P〉0．05），TRAIL水平则显著低于对照组（P〈0．001）；Westernblot结果表明模型鼠的抗凋亡蛋白Bcl-2的表达较对照组显著增高。结论TH1细胞的凋亡缺陷可能在PBC小鼠模型的发病机制中有重要作用，该缺陷可能是由于自身免疫机制引起凋亡相关分子Fas体系及TRAIL的表达变化引起，同时通过上调Bcl-2的表达抑制自身反应性T细胞的凋亡。Objective To study the activation induced celt death （AICD） of CD4^＋T cells in primary biliary cirrhosis（PBC）murine model induced by poly I： C. Methods Thirty female C57BL/6 mice were divided into model and control group randomly, and the former were injected with 5 mg/kg of poly I： C, the later with PBS. PBC mice were detected 16 weeks after injection. CD4^＋T cells isolated from spleen were stimulated in vitro by Con A and anti-CD3, and the apoptosis were determined by Annexin-V and PI staining. The expression of Fas, FasL and TRAIL were assayed by relative quantitative real-time PCR. Bcl-2 was detected by Western blot. Results Compared with control group, the portal areas of mice in model group were infiltrated with mononuclear cells obviously. The positive rate of serum antimitochondrial antibody （AMA） and the level of alkali phosphatase （ALP） were higher than that in control group （P 〈 0. 001 ）. AICD of splenic CD4^＋T cells in model group was lower than that of control group （ P 〈0.001 ）. The mRNA of FasL and TRAIL in model mice was down-regulated. Simultaneously, the anti-apoptosis protein Bcl-2 was up-regulated in model group. Conclusion These observations suggest that a defect in AICD of auto-reactive TH1 cells may contribute to the pathogenesis of PBC model. Furthermore, this defect in AICD may results from the change of Fas/FasL, TRAIL pathway and the up-regulation of Bcl-2.

关 键 词：聚肌胞苷酸 肝硬化 胆汁性 凋亡 模型 小鼠 

分 类 号：R575.2[医药卫生—消化系统]