人类疱疹病毒6A对神经细胞的嗜性及细胞周期的影响

Study on neural cell tropism of human herpesvirus 6 and its effect on neural cell cycle

目的:研究人类疱疹病毒6A对神经细胞嗜性及细胞周期的影响。方法:HHV-6A GS株感染神经细胞,倒置显微镜观察细胞病变。PCR法鉴定神经细胞中HHV-6A U22基因,实时荧光定量PCR法测定神经细胞中HHV-6A U22基因的相对含量。免疫细胞化学和Western blot检测神经细胞中HHV-6A糖蛋白gB的表达。神经细胞感染HHV-6A后MTT法测定细胞增殖的改变,流式细胞仪检测神经细胞周期的改变。结果:HHV-6A GS株感染5天后,U373细胞形态无明显变化,SK-N-SH和SHG44细胞聚集,折光性降低,细胞发生明显病变。PCR法检测到U373、SK-N-SH和SHG44细胞中均含有HHV-6A U22基因,实时荧光定量PCR法发现随着感染天数的增加,HHV-6A DNA在细胞中含量逐渐降低。免疫细胞化学和Western blot结果显示病毒糖蛋白gB在细胞中表达,其中在U373细胞和SHG44细胞中的表达量高于SK-N-SH细胞。MTT法显示HHV-6A促进U373细胞和SHG44细胞的增殖,抑制SK-N-SH细胞的增殖。流式细胞仪检测细胞周期,结果显示U373和SHG44细胞感染组与对照组相比,G1期细胞百分数减少,S和G2期细胞百分数增多;SK-N-SH细胞G1期细胞百分数增多,而S和G2期细胞百分数减少。结论:HHV-6A感染神经细胞,对神经胶质细胞的嗜性强于神经元细胞。HHV-6A通过改变神经细胞周期进程,促进神经胶质细胞增殖,而抑制神经元细胞增殖。

Objective：To study neural cell tropism of human herpesvirus 6A（HHV-6A） and its effect on neural cell cycle.Methods：The cell morphology was assessed by a light microscope.The fragment of HHV-6A U22 gene was amplified by PCR and relative amount of HHV-6A U22 gene was analyzed using quantitative real-time PCR.Expression of HHV-6A gB was examined by immunocytochemical and Western blot analysis.Cell proliferation was measured by MTT assay.Cell cycle analysis was evaluated by the propidium iodide assay.Results：On the fifth day after being infected by HHV-6A,U373 cells had no obvious change,but SK-N-SH and SHG44 cells showed cytopathic effect.HHV-6A U22 gene was detected in all three infected cell lines and its relative amounts kept reducing all the time.HHV-6A gB was positive in the three infected cell lines,furthermore,the expression levels in U373 and SHG44 were more than those in SK-N-SH.HHV-6A stimulated the proliferations of U373 and SHG44 cells,however,inhibited the proliferation of SK-N-SH cells significantly.Compared with uninfected cells,percentages of U373 and SGH44 cells were decreased in phase G1 but increased in phase S and G2 after cells were infected by HHV-6A,as to SK-N-SH,the percentages were increased in phase G1 but decreased in S and G2 phases.Conclusion：All three neural cell lines can sustain replication of HHV-6A,and moreover,HHV-6A can boost astrocyte proliferation and inhibit neuronal cell proliferation.