M2型巨噬细胞来源的外泌体lncRNA NR_028113.1通过激活JAK2/STAT3通路促进巨噬细胞的极化  

M2 macrophage-derived exosomal lncRNA NR_028113.1 promotes macrophage polarization possibly by activating the JAK2/STAT3 signaling pathway

作　　者：张梦莹[1,2] 李志[3] 裴纬亚[1,2] 李雪琴 杨辉[1,2] 朱小龙[1,2] 吕坤 ZHANG Mengying;LI Zhi;PEI Weiya;LI Xueqin;YANG Hui;ZHU Xiaolong;LÜKun(Key Laboratory of Non-coding RNA Transformation Research of Anhui Higher Education Institution,Wannan Medical College,Wuhu 241001,China;Central Laboratory,Yijishan Hospital,Wannan Medical College,Wuhu 241001,China;Department of Rheumatology,Yijishan Hospital,Wannan Medical College,Wuhu 241001,China)

机构地区：[1]皖南医学院重大疾病非编码RNA转化研究安徽普通高校重点实验室,安徽芜湖241001 [2]皖南医学院弋矶山医院中心实验室,安徽芜湖241001 [3]皖南医学院弋矶山医院风湿免疫科,安徽芜湖241001

出　　处：《南方医科大学学报》2023年第3期393-399,共7页Journal of Southern Medical University

基　　金：国家自然科学基金(82072370,82100019,81802503);安徽省自然科学基金(2108085J44、2108085QH307);皖南医学院重点科研项目培育基金(WK2021ZF05)。

摘　　要：目的探究M2型巨噬细胞来源的外泌体lncRNA NR_028113.1对巨噬细胞极化的影响及机制。方法体外分离并培养BALB/c小鼠骨髓来源的巨噬细胞(BMDMs),IL-4诱导其向M2型巨噬细胞极化后,提取并鉴定M2细胞上清液所分泌的外泌体exosome(M2-exo),qRT-PCR检测M2外泌体中lncRNA的表达。将100μg/mL的M2-exo,对照组用等体积的PBS分别与M0巨噬细胞共孵育48 h后,qRT-PCR及Western blot检测各组细胞中Arg1、YM-1、FIZZ1、iNOS和TNF-α的表达,流式细胞术检测CD206^(+)细胞比例,Western blot检测各组细胞JAK2/STAT3蛋白磷酸化水平。设计、合成lncRNA smart silencer特异性抑制lncRNA NR_028113.1的表达,转染M2细胞48 h后提取各组细胞(exo+NC和exo+smart silencer)上清液分泌的外泌体再与M0型巨噬细胞共孵育,检测孵育后各组细胞中Arg1、YM-1、FIZZ1、iNOS和TNF-α的表达以及CD206^(+)细胞比例和JAK2/STAT3信号通路蛋白磷酸化水平。结果lncRNA NR_028113.1在M2型巨噬细胞外泌体中高表达(P<0.05)。相比于PBS对照组,与M2-exo共培养的M0巨噬细胞中Arg1、YM-1和FIZZ1表达均显著上调(P<0.05),iNOS和TNF-α表达显著下调(P<0.05),CD206^(+)细胞所占比例显著增加(P<0.01),JAK2/STAT3蛋白磷酸化水平显著增加(P<0.05)。抑制M2-exo中lncRNA NR_028113.1的表达后,共培养的M0巨噬细胞中Arg1、YM-1和FIZZ1表达显著下调(P<0.05),iNOS和TNF-α表达显著上调(P<0.05),CD206^(+)细胞所占比例显著减少(P<0.05),JAK2/STAT3蛋白磷酸化水平显著减少(P<0.05)。结论M2型巨噬细胞来源的外泌体lncRNA NR_028113.1可显著促进巨噬细胞向M2型极化,其机制可能与激活JAK2/STAT3信号通路相关。Objective To explore the effect of M2 macrophage-derived exosomal lncRNA NR_028113.1 on macrophage polarization and its possible mechanism.Methods Bone marrow-derived macrophages(BMDMs)from BALB/c mice were isolated and cultured in vitro.After IL-4 treatment to induce M2 macrophage polarization,exosomes(M2-exo)were extracted from the supernatant of M2 macrophages and identified.The expression of lncRNA in M2-exo was detected by qRT-PCR.BMDMs were co-cultured with M2-exo(100μg/mL)or PBS for 48 h,and the changes in cellular expression levels of Arg1,YM-1,FIZZ1,iNOS and TNF-αwere detected using qRT-PCR and Western blotting.The percentage of CD206^(+)cells was analyzed using flow cytometry,and the phosphorylation levels of JAK2/STAT3 proteins were detected using Western blotting.A lncRNA smart silencer was designed to specifically inhibit the expression of lncRNA NR_028113.1 in the M2 macrophages,from which exosomes were extracted and co-cultured with BMDMs for 48 h.The mRNA expression levels of Arg1,YM-1,FIZZ1,iNOS and TNF-α,CD206^(+)cell percentage and the phosphorylation levels of JAK2/STAT3 proteins were detected using qRT-PCR,flow cytometry and Western blotting.Results LncRNA NR_028113.1 was highly expressed in the exosomes of M2 macrophages(P<0.05).Co-culture with M2-exo significantly increased mRNA expressions of M2 macrophage marker genes Arg1,YM-1 and FIZZ1(P<0.05),lowered the expressions of iNOS and TNF-α(P<0.05),and increased CD206^(+)cell percentage and JAK2/STAT3 protein phosphorylation level in BMDMs(P<0.05).After inhibiting the expression of lncRNA NR_028113.1 in M2 macrophages,the extracted M2-exo caused significant down-regulation of the mRNA expressions of Arg1,YM-1 and FIZZ1 and up-regulation of iNOS and TNF-αmRNA(P<0.05),resulting also in signi-ficantly reduced CD206^(+)cell percentage and lowered phosphorylation levels of JAK2/STAT3 proteins in co-cultured BMDM(P<0.05).Conclusions M2 macrophage-derived exosomal lncRNA NR_028113.1 can significantly promote M2 polarization of macrophages possi

关 键 词：外泌体 lncRNA 巨噬细胞极化 JAK2 STAT3 

分 类 号：R392[医药卫生—免疫学]