感染流感病毒巨噬细胞Dll1对分泌IFN-γ相关因素的影响

The effects of Dll1 expression on factors for IFN-γ secretion by macrophages infected with the influenza virus

目的通过体外途径,应用Notch信号阻断剂(GSI),研究流感病毒感染巨噬细胞Dll1对分泌IFN-γ相关MHC分子表达及细胞因子IL-6、IL-12、IL-18水平的影响。方法用A/H1N1流感病毒感染RAW264.7巨噬细胞,并加入不同浓度GSI与细胞共孵育48h,分别提取细胞总RNA及收集培养上清液,采用实时荧光定量PCR法检测MHC分子表达及细胞因子水平。结果与DMSO对照组比较,感染流感病毒的RAW 264.7在GSI作用下表面分子MHC-ⅠmRNA表达水平无明显变化(P>0.05),而MHC-ⅡmRNA表达降低(P<0.05)。经Pearson相关性分析Dll1mRNA表达与MHC-Ⅱ呈正相关(r=0.816,P<0.05)。感染流感病毒的RAW264.7细胞在GSI作用后分泌的IL-12和IL-18水平低于DMSO对照组(P<0.05),而IL-6水平高于对照组。0.5、1和5μmol/L GSI组IL-12含量[分别为(24.15±0.35)、(21.15±4)和(14.75±0.7)pg/ml]均低于DMSO对照组[(53±2.83)pg/ml](P<0.05);5μmol/L GSI组IL-18含量[(75±7.07)pg/ml]低于DMSO对照组[(114.75±4.5)pg/ml](P<0.05)。经Pearson相关性分析,Dll1mRNA表达与IL-12呈正相关(r=0.972,P<0.05),与IL-6呈负相关(r=-0.767,P<0.05)。结论不同Dll1基因表达水平影响分泌IFN-γ相关巨噬细胞表面MHC-Ⅱ及IL-12、IL-18的水平,从而影响IFN-γ介导的抗流感病毒免疫应答反应。巨噬细胞Dll1与抗流感病毒免疫相关。

Objective To use GSI to block Notch signals in vitro in order to study the effects of Dll1 expression on the factors for IFN-γsecretion,i.e.the expression of MHC molecules and the secretion of the cytokines IL-6,IL-12,and IL-18,by macrophages infected with the influenza virus. Methods RAW264.7macrophages were infected with influenza A/H1N1 virus,and cells were incubated with different concentrations of GSI for 48 h.Total RNA extracted from cultured cells and the supernatant were used to detect levels of MHC expression and cytokines. Results Compared to control cells treated with DMSO,RAW264.7macrophages infected with the influenza virus had lower levels of MHC-II mRNA but not MHC-I mRNA.Pearson correlation analysis indicated that expression of Dll1 mRNA was correlated with MHC-II（r=0.816,P〈0.05）.In RAW264.7macrophages infected with the influenza virus and treated with GSI,the levels of IL-12 and IL-18 decreased while the level of IL-6increased compared to levels in the control cells treated with DMSO.The level of IL-12[（24.15±0.35）,（21.15±4）,（14.75±0.7）pg/ml]was lower in cells treated with 0.5,1,or 5μmol/L of GSI than in control cells treated with DMSO [（53±2.83）pg/ml]（P〈0.05）;The level of IL-18[（75±7.07）pg/ml]increased in cells treated with 5μmol/L of GSI compared to the control cells treated with DMSO [（114.75±4.5）pg/ml]（P〈0.05）.Pearson correlation analysis indicated that the expression of Dll1 mRNA correlated with the level of IL-12（r=0.972,P〈0.05）and was inversely correlated with the level of IL-6（r=-0.767,P〈0.05）. Conclusion Different levels of Dll1 expression affect the expression of MHC-II and the levels of IL-12 and IL-18 in macrophages and thus their secretion of IFN-γ,affecting the IFN-mediated immune response to the influenza virus.Macrophage Dll1 is associated with immunity from the influenza virus.