内皮高表达脂多糖相关因子1抑制巨噬细胞炎症反应

Endothelial overexpressed lipopolysaccharide-associated factor 1 inhibits inflammatory response of macrophages

目的本实验旨在观察内皮高表达脂多糖相关因子1(endothelial overexpressed lipopolysaccharide-associated factor 1,EOLA1)对巨噬细胞炎症反应的影响及可能机制。方法培养小鼠巨噬细胞,脂多糖(lipopolysaccharide,LPS)刺激激活炎症信号通路,检测肿瘤坏死因子(TNF-α)、白介素6(IL-6)和EOLA1转录水平变化,采用化学阻断剂在不同位点阻断炎症反应信号通路,再检测TNF-α、IL-6和EOLA1转录变化。巨噬细胞过表达mEOLA1,RT-qPCR检测细胞M1、M2极化类型标志基因mRNA水平,以流式细胞术检测巨噬细胞表型改变,Western blot检测M1型巨噬细胞标记物iNOS表达。结果LPS刺激后巨噬细胞释放炎症因子TNF-α、IL-6增加,而EOLA1表达下调;不同位点阻断炎症反应信号通路后炎症介质的表达明显受抑制,而EOLA1的表达上升;在巨噬细胞中高表达EOLA1,再行LPS刺激,检测到TNF-α、IL-6增加并不明显,流式细胞检测显示高表达EOLA1的巨噬细胞在LPS刺激后仍保持非炎症极化的M2型,细胞表达iNOS下降。结论EOLA1具有负调节巨噬细胞炎症相关通路活化的作用,其表达增加可以促进巨噬细胞从促炎症的M1型向促愈合的M2型转变,减少局部炎症程度。

To evaluate the effects of endothelial overexpressed lipopolysaccharide-associated factor 1(EOLA1)on inflammatory release and the possible mechanism,mouse macrophages were cultured and stimulated by LPS at different time,and the levels of TNF-α,IL-6 and EOLA1 were detected and compared before and after TLRs/NF-κB pathway blockage.Macrophages overexpressing mEOLA1 were stimulated with LPS,and then the phenotypic changes of the macrophages were detected by flow cytometry and the expression of iNOS,a marker of M1 macrophages,was detected by Western blotting.Data showed that LPS stimulation increased the levels of TNF-αand IL-6,but reduced the expression of EOLA1 in macrophages.After blocking TLRs/NF-κB pathway,the expression of inflammatory mediators were inhibited and the expression of EOLA1 was increased.High expression of EOLA1 significantly inhibited the response of macrophages to LPS stimulation,and macrophages still maintained M2 type after inflammatory stimulation,while the expression of iNOS decreased.In conclusion,EOLA1 inhibits the response activity of macrophages to inflammatory stimuli and reduce local inflammatory response.