上海交通大学学报（医学版）

上海交通大学学报（医学版） ›› 2017, Vol. 37 ›› Issue (7): 914-.doi: 10.3969/j.issn.1674-8115.2017.07.006

• 论著（基础研究） • 上一篇    下一篇

CaMKKβ 通过激活 AMPK/JAK2/STAT3 信号促进小鼠单核巨噬细胞向 M2 表型转换

孔祥歆 1, 2，刘卉芳 2#，陈凤玲 2#   

  1. 1. 蚌埠医学院，蚌埠 233030；2. 上海交通大学 医学院附属第九人民医院内分泌科，上海 201999
  • 出版日期:2017-07-28 发布日期:2017-08-25
  • 通讯作者: ?陈凤玲，电子信箱 cfl1993@126.com。刘卉芳，电子信箱：lhf_404@163.com。# 共同通信作者
  • 作者简介:?孔祥歆（1989—）女，硕士生；电子信箱：kongxiangxin1106@126.com
  • 基金资助:
    国家自然科学基金青年基金（81400802）；上海交通大学医学院附属第九人民医院优秀青年骨干培养计划（jyyq08201607）

CaMKKβ promotes mouse macrophage M2 polarization by activating AMPK/JAK2/STAT3 signaling

KONG Xiang-xin1,2, LIU Hui-fang2#, CHEN Feng-ling2#   

  1. 1. Bengbu Medical College, Bengbu 233030, China; 2. Department of Endocrinology, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 201999, China
  • Online:2017-07-28 Published:2017-08-25
  • Supported by:
    National Natural Science Foundation of China, 81400802; Excellent Youth Training Program, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, jyyq08201607

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摘要： 目的 · 探明钙离子/ 钙调蛋白依赖性蛋白激酶激酶β（CaMKKβ）在白介素4（IL-4）诱导的RAW264.7 细胞极化过程中的 作用和机制。方法 · RT-PCR 和 ELISA 检测 IL-4 诱导 RAW264.7 细胞极化水平，Western blotting 测定极化过程中CaMKKβ、AMPK、 JAK2、STAT3 蛋白表达及磷酸化水平。慢病毒为载体的 shRNA 稳定干扰 RAW264.7 细胞 CaMKKβ 表达，RT-PCR 和 ELISA 检测 IL-4 诱导下细胞极化水平的改变， Western blotting 检测 AMPK、JAK2、STAT3 蛋白及磷酸化蛋白的表达。分别特异性阻断 AMPK、JAK2 和 STAT3 蛋白活性，经 RT-PCR 检测 IL-4 诱导下 RAW264.7 细胞极化水平的改变。结果 · IL-4 主要诱导 RAW264.7 细胞向 M2 型巨噬 细胞极化（P<0.05），且 CaMKKβ、AMPK、JAK2、STAT3 的磷酸化水平明显升高（均P<0.05）。 shRNA 稳定干扰RAW264.7 细胞 CaMKKβ 表达后，IL-4 促进巨噬细胞向 M2 型极化作用降低（P<0.05），且 AMPK、JAK2 和 STAT3 的磷酸化水平降低（均 P<0.05）。 分别阻断 AMPK、JAK2 和 STAT3，巨噬细胞向 M2 型极化减少（均 P<0.05）。 结论 · CaMKKβ 通过激活 AMPK/JAK2/STAT3 信号通 路，促进 IL-4 诱导的巨噬细胞 M2 型极化。

关键词: &ensp, CaMKK&beta, AMPK, JAK2, STAT3, 巨噬细胞极化

Abstract:

 Objective · To investigate the role and mechanism of CaMKKβ in RAW264.7 macrophage M2 polarization induced by interleukin-4 (IL-4). 
 Methods · IL-4 induced RAW264.7 polarization was measured by RT-PCR and ELISA. Western blotting was used to analyze the total and phosphorylated protein levels of CaMKKβ, AMPK, JAK2 and STAT3 during polarization. A lentiviral vector carrying an shRNA targeting the CaMKKβ gene was successfully constructed. The expression of M1 and M2 markers and the expression of phosphorylated AMPK, JAK2, STAT3 were detected by RTPCR and Western blotting respectively. The protein activity of AMPK, JAK2 and STAT3 were selectively blocked, and then the effect of IL-4 on macrophage polarization was detected by RT-PCR.  Results · IL-4 significantly polarized RAW264.7 cells towards M2 macrophages (P<0.05), and the phosphorylated protein expression of CaMKKβ, AMPK, JAK2, STAT3 was significantly increased (all P<0.05). The expression of M2 markers induced by IL-4 was decreased and the phosphorylation of AMPK, JAK2 and STAT3 was also decreased after knockdown of CaMKKβ by shRNA (all P<0.05). The polarization of macrophages to M2 was decreased after the activities of AMPK, JAK2 and STAT3 proteins were blocked respectively (all P<0.05).  Conclusion · CaMKKβ promotes IL-4 induced M2 macrophage polarization via activation of AMPK/JAK2/STAT3 pathway.

Key words:  CaMKKβ, AMPK, JAK2, STAT3, macrophage polarization