IFITM3调控流感病毒感染小鼠肺组织NLRP3炎症小体活化的研究

Study on the regulation of IFITM3 in activation of NLRP3Inflammasome in the lung of mice infected with influenza virus

目的探讨干扰素诱导的跨膜蛋白3(Interferon-induced transmembrane protein 3,IFITM3)在流感病毒感染过程中对肺组织NLRP3(NOD-like receptor thermal protein domain associated protein 3,NLRP3)炎症小体活化的调控作用,为流感的防治提供新的思路。方法构建IFITM3敲除小鼠(Ifitm3^(-/-)小鼠),用A/AiCHi/1968/2(H3N2)病毒分别感染Ifitm3^(-/-)小鼠与野生型小鼠,连续14 d记录体重变化并计算存活率。在感染后第0、1、3和5 d采集小鼠肺组织,通过病理组织切片考察小鼠肺组织的病理损伤,采用Western blot、RT-qPCR、Elisa法检测小鼠肺组织的NLRP3炎症小体激活状况。采用SPSS 20.0软件进行独立样本t检验,以P<0.05为差异有统计学意义。结果经A/AiCHi/1968/2(H3N2)病毒感染后,Ifitm3^(-/-)小鼠相比野生型小鼠表现出高的死亡率和更严重的肺组织炎症反应。较野生型小鼠而言,Ifitm3^(-/-)小鼠肺组织中NLRP3炎症小体效应蛋白NLRP3和caspase-1的基因和蛋白表达水平显著上升(P<0.05),同时NLRP3炎症小体下游因子IL-1β的mRNA拷贝数及分泌含量亦明显升高(P<0.05)。结论敲除IFITM3基因促使流感病毒感染小鼠肺组织NLRP3炎症小体的活化程度升高,首次发现在流感病毒感染时IFITM3参与了NLRP3炎症小体活化的调控过程,为IFITM3拮抗流感病毒的分子机制提供了新的线索。

Objective To explore the regulation of interferon-induced transmembrane protein 3 (IFITM3)in the activation of NLRP3 (Nod-like receptor thermal Protein Domain associated protein 3)inflammasome in lung tissues during influenza virus infection,and to provide new ideas for prevention and treatment of influenza.Methods IFITM3 knockout mice (Ifitm3^(-/-) mice) were constructed.A/AiCHi/1968/2 (H3N2) virus was used to infect Ifitm3^(-/-) and wild-type mice,respectively.Body weight changes were recorded and survival rates were calculated for 14consecutive days.Lung tissues of mice were collected on day 0,1,3 and 5 after infection,and pathological damage of lung tissue in mice was investigated through pathological tissue section.Western blot,RT-qPCR and Elisa were used to detect the activation of NLRP3 inflammasome in lung tissue of mice.SPSS 20.2 software was used for independent sample t test,and P<0.05 was considered as statistically significant.Results Ifitm^(3-/-)mice infected with A/AiCHi/1968/2 (H3N2)virus showed higher mortality and more severe lung tissue inflammation than wild-type mice.Compared with wild-type mice,the gene and protein expression levels of NLRP3 and caspase-1(two effector proteins of NLRP3 inflammasome) in lung tissues of Ifitm3^(-/-) mice significantly increased (P<0.05).The mRNA copy number and secretion of IL-1β,a downstream factor of NLRP3 inflammasome,also significantly increased at the same time (P<0.05).Conclusions IFITM3 gene knockout promoted the activation of NLRP3 inflammasome in lung tissues of mice infected with influenza virus.It was found for the first time that IFITM3 was involved in the regulation of NLRP3 inflammasome activation.These results provide a new clue for the molecular mechanism of IFITM3 antagonism against influenza virus.