金黄色葡菌球菌脂蛋白对M1型小鼠骨髓源巨噬细胞免疫作用的影响

本研究旨在阐明金黄色葡萄球菌脂蛋白对M1型小鼠骨髓源巨噬细胞免疫作用的影响，为金黄色葡萄球菌致病性研究提供理论参考。以金黄色葡萄球菌野生株SA113（WT SA113）和SA113 lgt：：ermB脂蛋白表达缺失菌株（SA113Δlgt株）体外感染M1型小鼠骨髓源巨噬细胞，分为3组：空白对照组、WT SA113感染组（MOI：3：1）、SA113Δlgt感染组（MOI：3：1）。采用ELISA法检测M1型小鼠骨髓源巨噬细胞中肿瘤坏死因子-α（TNF-α）、白细胞介素1β（IL-1β）、趋化因子（RANTES）和白细胞介素10（IL-10）的水平，实时荧光定量PCR检测Toll样受体2（TLR2）、Toll样受体4（TLR4）和含NLR家族Pyrin域蛋白3（NLRP3）基因的表达，免疫荧光法检测脂蛋白对M1型小鼠骨髓源巨噬细胞吞噬金黄色葡萄球菌作用的影响。结果显示，与空白对照组相比，WT SA113感染组和SA113Δlgt感染组均可显著上调M1型小鼠骨髓源巨噬细胞中TNF-α、RANTES、IL-10分泌量以及WT SA113感染组TLR2、NLRP3基因表达水平（P<0.05），而TLR4基因表达量显著降低（P<0.05）；与WT SA113感染组相比，SA113Δlgt感染组M1型小鼠骨髓源巨噬细胞中TNF-α、IL-1β、RANTES、IL-10分泌量以及TLR2（12 h除外）、NLRP3基因表达水平显著降低（P<0.05）。免疫荧光结果显示，M1型巨噬细胞对SA113Δlgt株的吞噬作用显著低于对WT SA113株的吞噬作用（P<0.05）。综上，金黄色葡萄球菌的脂蛋白在M1型小鼠骨髓源巨噬细胞中主要通过激活TLR2和NLRP3受体，诱导细胞因子TNF-α、IL-1β、RANTES和IL-10的产生和释放。

Effects of Staphylococcus aureus Lipoprotein on Immunity in M1 Mouse Bone Marrow-Derived Macrophages

The aim of this study was to clarify the effect of Staphylococcus aureus lipoproteins on immunity of M1 mouse bone marrow-derived macrophages, and provide the theoretical reference for the study of Staphylococcus aureus pathogenicity.In vitro, M1 mouse bone marrow-derived macrophages was infected by WT SA113 and SA113 lgt::ermB strains (SA113Δlgt) of Staphylococcus aureus.The mice were divided into three groups:Blank control group, WT SA113 infection group (MOI:3:1), and SA113Δlgt infection group (MOI:3:1).The tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), chemokines (RANTES) and interleukin-10 (IL-10) in M1 mouse bone marrow-derived macrophages was detected by ELISA method, the expression of Toll-like receptor 2 (TLR2), Toll-like receptor 2(TLR4) and recombinant NLR family, Pyrin domain containing protein 3 (NLRP3) genes were detected by Real-time quantitative PCR, and the effect of lipoproteins on phagocytosis of Staphylococcus aureus by M1 mouse bone marrow-derived macrophages was detected by immunofluorescence.The results showed that compared with control group, both WT SA113 and SA113Δlgt infection could significantly upregulate the secretion of TNF-α, RANTES and IL-10, and WT SA113 infection could significantly increase the expression of TLR2 and NLRP3 genes in M1 mouse bone marrow-derived macrophages polarization (P<0.05), while the expression of TLR4 gene was significantly decreased (P<0.05).Compared with WT SA113 infected group, the secretion of TNF-α, IL-1β, RANTES and IL-10, and the expression of TLR2 (expect for 12 h) and NLRP3 gene in SA113Δlgt infected group were significantly decreased (P<0.05).The immunofluorescence results showed that the phagocytosis effect of M1 macrophage on SA113Δlgt strain was significantly lower than that of WT SA113 strain (P<0.05).In conclusion, the lipoprotein of Staphylococcus aureus induced the production and release of cytokines TNF-α, IL-1β, RANTES and IL-10 mainly through activation of TLR2 and NLRP3 receptors in M1 mouse bone marrow-derived macrophages.