中介素通过激活AMPK信号通路对脂多糖诱导巨噬细胞极化的影响

目的探讨中介素(intermedin,IMD)对脂多糖(lipopolysaccharide,LPS)诱导小鼠单核巨噬细胞系RAW 264.7极化的影响及其作用机制。方法RAW 264.7细胞随机分为对照组、LPS组、LPS+IMD组、LPS+IMD+CC(AMPK抑制剂Compound C)组。Real time-PCR法检测TNF-α、CD86、iNOS、Arg^-1、CD206 mRNA表达,Western blot法检测p-AMPK、AMPK、TNF-α、IL-6和IL^-10蛋白表达,流式细胞术检测巨噬细胞亚型,ELISA法检测培养基上清IL-6和TNF-α浓度。结果与对照组及LPS组比较,IMD处理可增加AMPK磷酸化水平,增加p-AMPK/AMPK比值;与对照组相比,LPS诱导可导致巨噬细胞发生M1极化,M1型标志分子CD86、TNF-α及iNOS mRNA表达升高,M2型标志分子CD206、Arg^-1 mRNA表达降低,上调促炎因子TNF-α、IL-6表达,降低抑炎因子IL^-10表达,使M1型细胞数量增加,细胞上清中TNF-α、IL-6分泌增加;而IMD处理可抑制LPS诱导的M1极化,AMPK抑制剂Compound C组处理可在一定程度上拮抗这一作用。结论IMD通过激活AMPK信号通路抑制LPS诱导的巨噬细胞M1型极化。

Intermedin inhibits lipopolysaccharide-induced polarization of macrophages by activating AMPK signaling pathway

Aim To evaluate the mechanism by which intermdin(IMD)inhibits lipopolysaccha ride(LPS)-induced polarization in RAW264.7 cells.Methods RAW264.7 cells were divided into control groups,LPS groups,LPS+IMD groups,LPS+IMD+Compound C groups.The mRNA expressions of tumor necrosis factor-α,(TNF-α,),CD86,inducible nitric oxide synthase(iNOS),Arginase-1(Arg^-1)and CD206 were detected by Realtime-PCR.The protein expressions of p-AMPK,AMPK,TNF-α,intereukin-6(IL-6)and intereukin-10(IL^-10)were detected by Western blot.The proportion of CD86+M1 type cells was detected by Flow cytometry.In addition,the expression levels of supernatant cytokines,including IL-6 and TNF-α,were detected by ELISA.Results Compared with control and LPS groups,IMD treatment could up-regulate the expression level of p-AMPK and the ratio of p-AMPK/AMPK.LPS promoted M1 polarization,since the expressions of CD86,TNF-αand iNOS increased,while the expressions of CD206 and Arg^-1 decreased by LPS induction.The proportion of M1 type cells increased and the secretion of TNF-α,IL-6 in the cell supernatant increased.And IMD treatment could inhibit the polarization of M1 induced by LPS.These effects were reversed by Compound C,an inhibitor of AMPK.Conclusion IMD can inhibit the M1-type polarization of LPS-induced macrophages by activating AMPK signaling pathway.