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目的探讨重组甲型流感病毒基质蛋白1/2(rM1/2)诱导气管上皮细胞产生干扰素-γ的作用,以及基于p38MAPK信号因子的诱导作用机制。方法将小鼠原代气管上皮细胞分成6组,分别为M1组、M2组、病毒组、M1+病毒组、M2+病毒组、正常对照组。各组分别用相应制剂干预细胞4、8、24h,抑制试验中各实验组提前1h分别加入p38抑制剂,再进行相应制剂干预。提取细胞总RNA和总蛋白,分别采用RT-PCR和Western blot方法检测IFN-γmRNA和IFN-γ、p38MAPK、P-p38MAPK的表达。结果重组甲型流感病毒M1/2作用于小鼠气管上皮细胞,作用4h,8h,24h后的半定量RT-PCR和Western blot实验结果为rM1、rM2组诱导IFN-γmRNA表达量高于正常组,表示M1/2能诱导IFN-γ的产生。Western blot结果显示rM1、rM2组诱导P-p38MAPK表达量高于正常组,用p38MAPK抑制剂SB203580后,rM1联合病毒、rM2联合病毒组诱导P-p38MAPK表达量低于病毒组,且rM1联合病毒、rM2联合病毒组诱导IFN-γmRNA表达量低于病毒组,表示M1/2能诱导p38MAPK磷酸化。结论甲型流感病毒rM1/2能够在早期诱导小鼠气管上皮细胞产生IFN-γ;该作用与p38MAPK信号因子激活有关。
Objective To investigate the effect of recombinant human influenza virus matrix protein 1/2 (rM1 / 2) on IFN-γ induced by tracheal epithelial cells and the induction mechanism based on p38MAPK signaling. Methods Primary mouse tracheal epithelial cells were divided into 6 groups: M1 group, M2 group, virus group, M1 + virus group, M2 + virus group and normal control group. The cells were treated with the corresponding agents for 4, 8, 24 hours, respectively, and each experimental group in the inhibition test was added with p38 inhibitor 1 h before the intervention. Total RNA and total protein were extracted. The expression of IFN-γmRNA, IFN-γ, p38MAPK and P-p38MAPK were detected by RT-PCR and Western blot respectively. Results The results of RT-PCR and Western blot showed that the expression of IFN-γ mRNA in rM2 group was higher than that in normal group after 4 h, 8 h and 24 h after recombinant influenza virus M1 / 2 was administered to mouse tracheal epithelial cells , Indicating that M1 / 2 can induce IFN-γ production. Western blot results showed that the expression of P-p38MAPK in rM1 and rM2 groups was higher than that in normal group. The expression of P-p38MAPK in rM1 and rM2 combined with virus group was lower than that in normal group after p38MAPK inhibitor SB203580, The expression level of IFN-γmRNA in rM2 combined virus group was lower than that in virus group, which indicated that M1 / 2 could induce the phosphorylation of p38MAPK. Conclusion Influenza A virus rM1 / 2 can induce the production of IFN-γ in mouse tracheal epithelial cells in early stage, which is related to the activation of p38 MAPK.