中国组织工程研究

中国组织工程研究 ›› 2024, Vol. 28 ›› Issue (13): 2036-2041.doi: 10.12307/2024.141

• 干细胞基础实验 basic experiments of stem cells • 上一篇    下一篇

过表达miR-378a促进巨噬细胞向M2极化且抑制巨噬细胞向M1极化

杨  泉1，2，何惠宇2，3，王思凡2，3，吕尚毅2，3，周琦琪2，3，韩祥祯2，3    

  1. 新疆医科大学第一附属医院(附属口腔医院)，1口腔急诊综合科，3口腔修复科，新疆维吾尔自治区乌鲁木齐市   830054；2新疆维吾尔自治区口腔医学研究所，新疆维吾尔自治区乌鲁木齐市   830054
  • 收稿日期:2023-04-06 接受日期:2023-04-15 出版日期:2024-05-08 发布日期:2023-08-28
  • 通讯作者: 韩祥祯，硕士，主治医师，讲师，新疆维吾尔自治区口腔医学研究所，新疆维吾尔自治区乌鲁木齐市 830054 ；新疆医科大学第一附属医院(附属口腔医院)口腔修复科，新疆维吾尔自治区乌鲁木齐市 830054
  • 作者简介:杨泉，男，1994年生，新疆维吾尔自治区昌吉市人，在读硕士，主要从事口腔修复学及组织工程研究。
  • 基金资助:
    新疆维吾尔自治区自然科学基金计划青年项目(2021D01C337)，项目负责人：韩祥祯

Overexpression of miR-378a promotes macrophage M2 polarization and inhibits M1 polarization

Yang Quan1, 2, He Huiyu2, 3, Wang Sifan2, 3, Lyu Shangyi2, 3, Zhou Qiqi2, 3, Han Xiangzhen2, 3   

  1. 1Department of Oral Emergency Medicine,3Department of Prosthodontics, First Affiliated Hospital of Xinjiang Medical University, Urumqi 830054, Xinjiang Uygur Autonomous Region, China; 2Xinjiang Institute of Stomatology, Urumqi 830054, Xinjiang Uygur Autonomous Region, China
  • Received:2023-04-06 Accepted:2023-04-15 Online:2024-05-08 Published:2023-08-28
  • Contact: Han Xiangzhen, Master, Attending physician, Lecturer, Xinjiang Institute of Stomatology, Urumqi 830054, Xinjiang Uygur Autonomous Region, China; Department of Prosthodontics, First Affiliated Hospital of Xinjiang Medical University, Urumqi 830054, Xinjiang Uygur Autonomous Region, China
  • About author:Yang Quan, Master candidate, Department of Oral Emergency Medicine, First Affiliated Hospital of Xinjiang Medical University, Urumqi 830054, Xinjiang Uygur Autonomous Region, China; Xinjiang Institute of Stomatology, Urumqi 830054, Xinjiang Uygur Autonomous Region, China
  • Supported by:
    Youth Project of Natural Science Foundation of Xinjiang Uygur Autonomous Region, No. 2021D01C337 (to HXZ)

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摘要：


文题释义：

巨噬细胞极化：巨噬细胞首先被极化为促炎M1表型，M1巨噬细胞通过增强吞噬作用以及增加促炎细胞因子的产生和释放，促进先天免疫，以清除损伤部位产生的组织碎片以及来自体外的病原微生物。随后，巨噬细胞表型改变，形成以抗炎反应为主的M2表型并促进组织重塑和修复。
慢病毒：由于慢病毒载体具有许多吸引人的特点，已成为基因治疗中体外转基因传递的首选工具之一。慢病毒载体基因组一旦融入宿主细胞基因组，就可以实现长期稳定的转基因表达，并且慢病毒载体的遗传毒性更低，已成为转基因传递的首选载体。


背景：M2型巨噬细胞具有降低炎症因子及促进组织愈合的功能，因此，如何调节巨噬细胞M2极化是近年来研究的热点，研究发现部分miRNAs具有此功能。

目的：探讨miR-378a对Raw264.7巨噬细胞系极化的影响。
方法：首先使用脂多糖和干扰素γ协同诱导巨噬细胞M1极化，使用白细胞介素4诱导巨噬细胞M2极化，qRT-PCR检测各型细胞中内源性miR-378a的表达，验证miR-378a是否参与巨噬细胞的极化。通过慢病毒载体将miR-378a转染至巨噬细胞，筛选出miR-378a稳定表达的细胞系，使用脂多糖和干扰素γ协同诱导巨噬细胞M1极化，使用白细胞介素4诱导巨噬细胞M2极化，ELISA检测巨噬细胞培养基上清液中M1/M2极化相关细胞因子水平，qRT-PCR检测M1/M2型巨噬细胞极化特征及相关细胞因子的mRNA表达。

结果与结论：①诱导巨噬细胞极化后，各组Raw264.7细胞中内源性miR-378a的表达量均升高；②与未转染组比较，转染miR-378a组诱导巨噬细胞M1极化后促炎性细胞因子诱导型一氧化氮合酶、肿瘤坏死因子α、白细胞介素6、白细胞介素1β表达显著降低(P < 0.05)，细胞上清液中诱导型一氧化氮合酶、肿瘤坏死因子α、白细胞介素6水平也明显降低(P < 0.05)；③与未转染组比较，转染miR-378a组诱导巨噬细胞M2极化后CD206、白细胞介素10、精氨酸酶Ⅰ的表达显著升高(P < 0.05)，细胞上清液中CD206、白细胞介素10水平也明显升高(P < 0.05)；④结果表明：过表达miR-378a促进巨噬细胞向M2极化且抑制巨噬细胞向M1极化。

https://orcid.org/0009-0009-5367-1725 (杨泉) 

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

关键词: miR-378a, 巨噬细胞, 极化, 慢病毒, 炎症因子

Abstract: BACKGROUND: M2 macrophages have the function of reducing inflammatory factors and promoting tissue healing. Therefore, how to regulate M2 polarization of macrophages has been a hot research topic in recent years, and some miRNAs have been found to have this function.
OBJECTIVE: To investigate the effects of miR-378a on the polarization of the Raw264.7 macrophage cell line.
METHODS: The M1 polarization of macrophages was induced by lipopolysaccharide and interferon-γ. Interleukin-4 induced M2 polarization and the expression of endogenous miR-378a in each cell type was detected using qRT-PCR to verify whether miR-378a was involved in the polarization of macrophages. By transfection with lentivirus as the vector of overexpression of miR-378a, the stable expression of miR-378a cell lines was screened. Macrophage M1 polarization was induced synergically by lipopolysaccharide and interferon-γ. Macrophage M2 polarization was induced by interleukin-4. The levels of M1/M2 polarization-related cytokines in the supernatant of the macrophage culture medium were determined by enzyme-linked immunosorbent assay. qRT-PCR was used to detect the polarization characteristics of M1/M2-type macrophages and the mRNA expression levels of related cytokines. 
RESULTS AND CONCLUSION: (1) The expression level of endogenous miR-378a in Raw264.7 cells of each group increased after macrophage polarization. (2) Compared with the non-transfected group, the expressions of proinflammatory cytokine-induced nitric oxide synthase, tumor necrosis factor-α, interleukin-6 and interleukin-1β in macrophage M1 induced polarization were significantly decreased in the miR-378a transfection group (P < 0.05); the levels of inducible nitric oxide synthase, tumor necrosis factor-α and interleukin-6 in cell supernatant were also significantly decreased (P < 0.05). (3) Compared with the non-transfected group, the expressions of CD206, interleukin-10 and arginase-I in macrophage M2 induced polarization were significantly increased (P < 0.05); the levels of CD206 and interleukin-10 in cell supernatant were also significantly increased (P < 0.05) in the miR-378a transfection group. (4) It is indicated that overexpression of miR-378a promotes the M2 polarization of macrophages and inhibits the M1 polarization of macrophages. 

Key words: miR-378a, macrophage, polarization, lentivirus, inflammatory factor

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