摘要
目的:评价黄芪甲苷(AS-Ⅳ)对脂多糖(LPS)调节巨噬细胞M1/M2型极化的影响及其相关机制.方法:体外培养小鼠单核巨噬细胞系RAW264.7,采用随机数字表法分为3组:对照组(C组)、LPS组、AS-Ⅳ干预(AS-Ⅳ)组.C组细胞正常培养,LPS组加入终浓度为100 ng/mL的LPS处理24 h,AS-Ⅳ组加入终浓度为100 ng/mL的LPS和200 μg/mL的AS-Ⅳ处理24 h.利用ELISA法检测白细胞介素(IL)-1β、IL-10和IL-18水平,qRT-PCR法检测CD86、CD11c、CD206和精氨酸酶-1(Arg-1)的mRNA表达,流式细胞术检测CD86(+)和CD206(+)细胞百分比,Western blot法检测NOD样受体热蛋白结构域相关蛋白3(NLRP3)、凋亡相关斑点样蛋白(ASC)和半胱氨酸蛋白酶-1(caspase-1)的蛋白表达.结果:与C组比较,LPS组IL-1β和IL-18水平、CD86和CD11c mRNA表达、CD86(+)细胞百分比、NLRP3、ASC和caspase-1蛋白表达升高,IL-10水平、CD206和Arg-1 mRNA表达、CD206(+)细胞百分比降低(P＜0.05);与LPS组比较,AS-Ⅳ组IL-1β和IL-18水平、CD86和CD11c mRNA表达、CD86(+)细胞百分比、NLRP3、ASC和caspase-1蛋白表达降低,IL-10水平、CD206和Arg-1 mRNA表达、CD206(+)细胞百分比升高(P＜0.05).结论:AS-Ⅳ能够抑制LPS诱导的巨噬细胞M1型极化并促进其向M2型极化,其机制与下调NLRP3炎症小体表达有关.
Abstract
Objective To evaluate the effect of Astragaloside Ⅳ(AS-Ⅳ)on lipopolysaccharide(LPS)-induced macrophages M1/M2 polarization and related mechanism.Methods Mouse monocyte-derived macrophages RAW264.7 were cultured in vitro and were randomly divided into 3 groups:control group(group C),LPS group(group LPS)and AS-Ⅳ treatment group(group AS-Ⅳ).In group C,cells were cultured in the common culture medium.In group LPS,100 ng/mL LPS was added into the culture medium for 24 h incubation.In group AS-Ⅳ,100 ng/mL LPS and 200 µg/mL AS-Ⅳ were added into the culture medium for 24 h incubation.The levels of IL-1 β,IL-10 and IL-18 in culture medium were measured by ELISA assay.The mRNA levels of CD86,CD11c,CD206 and Arginase-1(Arg-1)were measured by qRT-PCR assay.The percentage of CD86(+)and CD206(+)cells was assessed by flow cytometry.The expression levels of nucleotide-binding oligomerization domain-like receptor containing pyrin domain 3(NLRP3)and apoptosis-associated speck-like protein(ASC)were detected by Western blot assay.Results Compared with group C,the levels of IL-1 β and IL-18,the mRNA levels of CD86 and CD1 1c,the percentage of CD86(+)cells and the expression levels of NLRP3,ASC and caspase-1 were increased,while the level of IL-10 and the mRNA levels of CD206 and Arg-1,and the percentage of CD206(+)cells were decreased in group LPS(P＜0.05).Compared with group LPS,the levels of IL-1β and IL-18,the mRNA levels of CD86 and CD11c,the percentage of CD86(+)cells and the expression levels of NLRP3,ASC and caspase-1 were decreased,while the level of IL-10 and the mRNA levels of CD206 and Arg-1,and the percentage of CD206(+)cells were increased in group AS-Ⅳ(P＜0.05).Conclusion AS-Ⅳ can inhibit LPS-induced macrophages M1 polarizaion and promote macrophages M2 polarizaion,whose mechanism may be associated with down-regulating NLRP3 inflammasome expression.
维普
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