南方医科大学学报

南方医科大学学报 ›› 2023, Vol. 43 ›› Issue (3): 393-399.doi: 10.12122/j.issn.1673-4254.2023.03.08

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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

张梦莹，李 志，裴纬亚，李雪琴，杨 辉，朱小龙，吕 坤   

  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，皖南医学院弋矶山医院中心实验室，风湿免疫科，安徽 芜湖 241001
  • 出版日期:2023-03-20 发布日期:2023-03-20

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

ZHANG Mengying, LI Zhi, PEI Weiya, LI Xueqin, YANG Hui, ZHU Xiaolong, LÜ Kun   

  1. Key Laboratory of Non- coding RNA Transformation Research of Anhui Higher Education Institution, Wannan Medical College, Wuhu 241001, China; Central Laboratory, Department of Rheumatology, Yijishan Hospital, Wannan Medical College, Wuhu 241001, China
  • Online:2023-03-20 Published:2023-03-20

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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100 μg/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。

关键词: 外泌体；lncRNA；巨噬细胞极化；JAK2；STAT3

Abstract: 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 (100 μg/mL) or PBS for 48 h, and the changes in cellular expression levels of Arg1, YM-1, FIZZ1, iNOS and TNF-α were 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-α mRNA (P<0.05), resulting also in significantly 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 possibly by activating the JAK2/STAT3 signaling pathway.

Key words: exosomes; long non-coding RNA; macrophage polarization; JAK2; STAT3