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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 investigate the effect of influenza virus infected macrophages D111 on the expression of IFN-γ-associated MHC molecules and the levels of cytokines IL-6, IL-12 and IL-18 in vitro by using Notch signal blocker (GSI). Methods RAW264.7 macrophages were infected with A / H1N1 influenza virus and incubated with different concentrations of GSI for 48 hours. The total RNA was extracted and the supernatant was collected. The expression of MHC and the expression of MHC were detected by real-time fluorescence quantitative PCR. Factor level. Results Compared with the DMSO control group, the expression of MHC-ⅠmRNA in RAW 264.7 infected with influenza virus had no significant change (P> 0.05) and the expression of MHC-ⅡmRNA decreased (P <0.05). The Pearson correlation analysis showed that Dll1 mRNA expression was positively correlated with MHC-Ⅱ (r = 0.816, P <0.05). The level of IL-12 and IL-18 secreted by influenza A virus-infected RAW264.7 cells after GSI treatment was lower than that of DMSO control group (P <0.05), while the level of IL-6 was higher than that of control group. IL-12 levels were significantly lower in 0.5, 1, and 5 μmol / L GSI groups (24.15 ± 0.35, 21.15 ± 4 and 14.75 ± 0.7 pg / ml, respectively) than those in DMSO control group / ml] (P <0.05). The level of IL-18 in the group of 5μmol / L GSI [(75 ± 7.07) pg / ml] was lower than that in the DMSO control group [(114.75 ± 4.5) pg / ml] The Pearson correlation analysis showed that Dll1 mRNA expression was positively correlated with IL-12 (r = 0.972, P <0.05) and negatively correlated with IL-6 (r = -0.767, P <0.05). Conclusions Different Dll1 gene expression levels affect the secretion of MHC-Ⅱ and IL-12 and IL-18 on the surface of IFN-γ-dependent macrophages, which may affect IFN-γ-mediated anti-influenza virus immune response. Macrophage Dll1 is immunologically related to anti-influenza virus.