不同亚型甲型流感病毒在单个核细胞内复制情况研究

Replication of different influenza A virus subtypes in mononuclear cells

目的研究不同亚型的甲型流感病毒在单个核细胞内的复制情况,探讨其免疫应答机制。方法①细胞培养:复苏A549、MDCK细胞后,用DMEM培养液常规培养;外周血分离得到单个核细胞,用RPMI1640常规培养;②空斑形成试验:用空斑试验检测病毒A/Shantou/169/2006(H1N1)和A/Shantou/602/2006(H3N2)的病毒原始滴度;检测流感病毒感染单个核细胞后的病毒滴度变化。结果流感病毒感染单核细胞后,上清液的病毒滴度下降,36、48、72 h病毒滴度<10 PFU/mL;而其细胞裂解液病毒滴度上升,滴度由9.5×10~4 PFU/mL上升至1.63×10~5 PFU/mL。流感病毒感染淋巴细胞,其细胞上清和裂解液病毒滴度均下降,其中上清液H3N2病毒滴度在48、72 h均<10 PFU/mL;裂解液病毒滴度则由4.8×10~5 PFU/mL下降至1.8×10~3 PFU/mL。结论不同亚型的甲型流感病毒在单核细胞和淋巴细胞中的复制存在差异。单核细胞可以吞噬流感病毒但不能直接灭活流感病毒,而淋巴细胞却可以直接抑制流感病毒的复制。

Objective This article was to investigate the replication of different subtypes of influenza A virus in mononuclear cells and to explore its immune response mechanism.Methods Cell culture:A549 and MDCK cells were recovered and cultured in DMEM medium.Then mononuclear cells were isolated from peripheral blood and cultured in RPMI1640 medium.Plaque formation test:the original viral titers were detected by a plaque formation test for an original titers from virus A/Shantou/169/2006(H1 N1)and A/Shantou/602/2006(H3 N2).After mononuclear cells were infected by influenza virus,the change from viral titer was detected by a plaque formation test.Results When monocytes were infected by influenza virus,the virus titer in the supernatant decreased,and the titer was less than 10 PFU/mL at 36 h,48 h and 72 h.However,the virus titer in the cell lysate increased from 9.5×10~4 PFU/mL to 1.63×10~5 PFU/mL at 48 h.When lymphocytes were infected by influenza virus,the virus titer decreased in both supernatant and lysate,in which the titer of H3 N2 in supernatant was less than 10 PFU/mL at 48 h and 72 h,and the titer in the lysate decreased from 4.8×10~5 PFU/mL to1.8×10~3 PFU/mL at 48 h.Conclusion There appeared some difference for the replication of influenza A virus subtypes between monocytes and lymphocytes.The virus was engulfed by monocytes but not directly inactivated it,while the replication of virus could be effectively inhibited by lymphocytes.