论文部分内容阅读
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 the effect of human herpesvirus 6A on neuronal cell tropism and cell cycle. Methods: HHV-6A GS strains were infected with neurons and the cells were observed under inverted microscope. The HHV-6A U22 gene was identified by PCR and the relative content of HHV-6A U22 gene in neurons was determined by real-time fluorescence quantitative PCR. Immunocytochemistry and Western blot were used to detect the expression of gH in HHV-6A glycoprotein in nerve cells. The changes of cell proliferation were measured by MTT assay after nerve cells were infected with HHV-6A, and the cell cycle was detected by flow cytometry. Results: After infected with HHV-6A GS strain for 5 days, the morphology of U373 cells did not change significantly. SK-N-SH and SHG44 cells aggregated, the refraction decreased, and the cells had obvious pathological changes. The HHV-6A U22 gene was detected by PCR in both U373, SK-N-SH and SHG44 cells. Real-time quantitative PCR showed that the content of HHV-6A DNA decreased gradually with the increase of infection days. Immunocytochemistry and Western blot showed that the glycoprotein gB was expressed in U373 cells and SHG44 cells compared with SK-N-SH cells. MTT assay showed that HHV-6A promoted the proliferation of U373 cells and SHG44 cells and inhibited the proliferation of SK-N-SH cells. The cell cycle was detected by flow cytometry. The results showed that the percentage of cells in G1 phase and the percentage of cells in S phase and G2 phase increased in U373 and SHG44 infected cells compared with the control group. The percentage of cells in G1 phase of SK-N-SH cells increased and the percentage of S And G2 cells decreased. CONCLUSION: HHV-6A infects nerve cells and has more neurotrophic effects on glial cells than neurons. HHV-6A can inhibit the proliferation of neurons by changing the process of nerve cell cycle and promoting glial cell proliferation.