激光共聚焦技术证实HCV NS5在人胎盘滋养层细胞的定位表达

The laser confocal microscopy confirms hepatitis C virus NS5 expression in human placental trophoblast cells

目的胎盘屏障是营养物质以及某些药物、病原体、激素等从母体进入胎儿的必经之路。体外分离培养滋养层细胞是研究其功能及母体与胎儿之间物质交换的具体分子机制的细胞学基础。本研究的目的是探讨体外分离培养的人体胎盘滋养层细胞感染丙型肝炎病毒(HCV)后HCV NS5在滋养层细胞中的定位表达。方法采用胰蛋白酶消化法及Percoll密度梯度分离法分离培养人胎盘组织中滋养层细胞,以HCV RNA阳性血清对滋养层细胞进行体外感染试验,应用RT-PCR法定性及定量检测感染后细胞培养上清中HCV RNA,用激光共聚焦技术观察免疫荧光染色后HCV NS5在滋养层细胞中的定位。结果HCV感染后的细胞培养上清中可间断检测到HCV RNA,而对照组始终未检出;激光共聚焦观察显示HCV NS5主要定位于细胞核周围。结论感染HCV的人滋养层细胞中存在HCV NS5表达。此研究为进一步深入研究HCV的母婴传播分子机制奠定了实验基础。

Objective Placental barrier is the nutrients and certain drugs,pathogens,hormones from the mother into the baby＇s only way.Isolated and cultured trophoblast cells in vitro to study its function and material exchange between maternal and fetal specific molecular mechanism of the cellular basis.The purpose of this study is to explore the in vitro isolation and culture of human placental trophoblast cells infected with hepatitis C virus（HCV） after HCV NS5 in trophoblast cell-specific expression.Methods Trophoblastic cells have been prepared from human term placentae digested with trypsin,and centrifuged on Percoll gradient.Trophoblastic cells were incubated in HCV positive serum,and then the HCV infection of syncytiotroblasts was detected with RT-PCR.Ultrastructure characteristics were observed with laser confocal microscopy.Results The HCV RNA was intermittently detected in the supernatent of the culture medium during the 40 days culture.The antibody of HCV NS5 could be observed around the nucleus with laser confocal microscopy.Conclusion Infected with HCV present in human trophoblast cells expressing HCV NS5.To further in-depth study of HCV molecular mechanism of mother to child transmission to lay an experimental basis.