Preliminary Studies On Infection Of HBV In Human Placental Trophoblasts

Aim Vertical transmission is a kind of very important route of HBV transmission. In the route, HBV could infect fetus through placentae .which is intrauterine transmission.It was thought that intrauterine transmission was very difficult to be prevented for its mechanism not being known clearly. There were only a few domestic and overseas studies about this problem, but our department have performed macroscopic and microcosmic studies in ten years and gotten some important results which suggested that HBV intrauterine transmission might occur either through placental tears with transfusion of maternal blood into fetal circulation namely hematogenous transmission or progressive infection of every placental layer until the virus got to fetus, which was named "transplacental cellular traffic" .Then the latter was researched more systemically.According to the hypothesis of transplacentalcellular traffic, trophoblast cells as the outermost covering of placental barrier should be infected firstly, moreover some evidences of HBV infection in the cells has been gained by morphological and pathological methods in sections of placent tissue. But the process and the mechanism of HBV infection in trophoblast cells or reproduction as well as expression of the virus in the cells could not be researched by these methods. So the experiments on cultured trophoblast cells should be performed.In the present study , we were about to verify evidences of HBV infection in the cells by morphological and pathological methods and fumble about a feasible method of HBV infection in vitro by experiment on a hepatical cell line namely HepG2 cells,then tested the virus infection in cultured primary trophoblast cells in vitro. This study could be the base of coming further study about the mechanism of HBV infection in trophoblast cells.Methods (1)41 continuous pregnant women who were HBsAg positive in their sera and their neonates were be selected as our subjects, then their venous blood was collected from elbow vein and femoral vein respectively to detect HBsAg and HBeAg. Their placental tissue was collected to detect HBsAg in the cells by immunohistochemical method too.(2)The cultured HepG2 cells were incubated with HBV positive serum(HBsAg, HBeAg, HBV DNA positive), 2%PEG, 1%DMSO. Then HBsAg was be detected in supernatant and cells by ELISA and immunohistochemical method respectively and HBV DNA was be detected in cells by PCR to analyze the results of infection. (3)The experiment of HBV infectionin cultured primary trophoblast cells in vitro were performed by the same methods, moreover transmission electron microscopy and immunoelectron microscopic method were used to confirm the results. Result (1)HBsAg was found in placent tissue sections of 30 women in the 41 subjects .Positive signal is well-distribute minute brown drops which located in the cytoplasm of decidual cells, trophoblast cells , Hofbauer cells and endothelial cells. The frequency and intensity of positive signal were decreasing from decidual cells to endothelial cells. 〩BsAg was found in the supernatant and cells of the cultured HepG2 cells which incubated with HBV positive serum -. 2%PEG and 1%DMSO as well as with HBV positive serum and 2%PEG by ELISA. HBsAg could also be found in the cytoplasm of these cells by immunohistochemical method. On the other hand, HBV DNA was be found in these cells by PCR too. (3)In the experiment of HBV infection in cultured primary trophoblast cells in vitro, the results was similar to that of the experiment on HepG2 cells. HBsAg and HBV DNA were also found in supernatant and cells respectively by ELISA , immunohistochemical method and PCR. The results were confirmed by transmission electron microscopy and immunoelectron microscopic method.Conclusions According to some reports and our studies, HBV could infect trophoblast cells in placenta tissue .Cultured primary trophoblast cells might also be infected by HBV if the cells were incubated with HBV positive serum, 2%PEG and 1%DMSO or with HBV positive serum and 2%PEG in...