胎盘滋养层细胞的感染和凋亡与HBV的宫内传播机制  

Relationship between the infection and apoptosis of placental trophoblastic cells and the mechanism of hepatitis B virus intrauterine transmission

作　　者：白菡[1] 张琳[1] 何丽霞[2] 冯国和[1] 石理兰[1] 窦晓光[1] 赵桂珍[1] 

机构地区：[1]中国医科大学附属盛京医院感染科,辽宁省沈阳市110004 [2]中国医科大学附属盛京医院妇产科,辽宁省沈阳市110004

出　　处：《世界华人消化杂志》2007年第15期1695-1700,共6页World Chinese Journal of Digestology

基　　金：辽宁省自然科学基金;No.9910500706~~

摘　　要：目的:通过HBV感染体外培养的人绒毛膜滋养层细胞,探讨HBV宫内感染发生的机制.方法:对人早孕绒毛膜滋养层细胞进行原代培养和对人滋养层细胞系JEG-3进行传代培养,将HBV感染血清与原代及传代培养细胞共同孵育8-48h,倒置显微镜观察细胞形态及细胞间连接,细胞免疫荧光和免疫组织化学染色方法检测滋养层细胞中HBsAg和HBcAg的表达,荧光定量PCR方法检测被感染的滋养层细胞中的HBV DNA,TUNEL方法检测滋养层细胞的凋亡.结果:与HBV阳性血清共同孵育对滋养层细胞的形态和细胞间连接无明显影响;细胞免疫荧光和免疫组织化学染色结果显示,滋养层细胞与HBV感染血清共同孵育(8,24,48h)后,滋养层细胞可以出现HBsAg和HBcAg的阳性表达,24 h时HBsAg阳性细胞数量最多(8h:18.0%±3.67%;24h:30.6%±2.88%;48 h:24.8%±4.21%);荧光定量PCR方法检测到被感染的滋养层细胞中HBV DNA的存在;TUNEL结果显示,与HBV感染血清共同孵育可导致滋养层凋亡细胞数量逐渐增加(24h:18.6%±3.05%;48h:26.8%±2.86%:P<0.01).结论:滋养层细胞的感染可能是HBV通过胎盘屏障的途径之一;HBV感染可以诱导滋养层细胞产生凋亡,这可能是胎盘屏障阻止HBV宫内感染的一种保护性机制.AIM： To study the mechanism of hepatitis B virus （HBV） intrauterine transmission by infecting the cultured human trophoblastic cells in vitro. METHODS： Primary cultured trophoblastic cells from human first-trimester placenta and serial subcultivation of JEG-3 cell line were investigated. The primary cultured and subcultured cells were cultured in the HBV-infected serum for 8 to 48 hours. The appearance of the cells and intercellular conjunctions were observed under inverted microscope. The protein expression of HBsAg and HBcAg were detected by cell immunofluorescence and immunohistochemical staining. HBV DNA of the infected trophoblastic cells was detected by fluorescent quantitative polymerase chain reaction （FQ-PCR）. Cell apoptosis was tested by TUNEL technique. RESULTS： The appearance of the cells and intercellular conjunction were not obviously affected after co-culturing with HBV-positive serum. Cell immunofluorescence and immunohistochemistry showed that HBsAg and HBcAg can be detected in the trophoblastic cells after co-culturing with HBV-infected serum for 8, 24 and 48 hours, and HBsAg-positive cells were at the most level after 24-hour culturing （8 h： 18.0% ± 3.67%; 24 h： 30.6% ± 2.88%; 48 h： 24.8% ± 4.21%）. FQ-PCR demonstrated the existence of HBV DNA in the infected ceils. TUNEL exhibited that the co-culturing resulted in an increase of cell apoptosis （24 h： 18.6 % ± 3.05%; 48 h： 26.8% ±2.86%; P 〈 0.01）. CONCLUSION： The infection of trophoblastic cells may be one of the routes for HBV passing through the placental membrane. HBV infection may induce the apoptosis of trophoblastic cells, which may be a protective mechanism for placental membrane to block up the vertical transmission of HBV.

关 键 词：乙型肝炎病毒 人绒毛膜滋养层细胞系JEG-3 细胞免疫荧光 免疫组化 荧光定量聚合酶链式反应 原位缺口末端标记技术 官内传播 

分 类 号：R512.62[医药卫生—内科学]