南方医科大学学报

南方医科大学学报 ›› 2020, Vol. 40 ›› Issue (10): 1472-1479.doi: 10.12122/j.issn.1673-4254.2020.10.13

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高盐刺激可诱导巨噬细胞极化从而促进肾脏成纤维细胞的增及表型转化

陆 静，白志勋，匡晓燕，李 玲   

  • 出版日期:2020-10-20 发布日期:2020-10-20

High-salt exposure induces macrophage polarization to promote proliferation and phenotypic transformation of co-cultured renal fibroblasts

  • Online:2020-10-20 Published:2020-10-20

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摘要： 目的 探讨高盐诱导单核巨噬细胞极化，以及极化后巨噬细胞对肾脏纤维细胞株（NRK-49F）增殖及表型转化的影响。方法 培养大鼠骨髓单核巨噬细胞及NRK-49F，随后予以高盐（Na+161 mmol/L）刺激单核巨噬细胞2 h。采用RT-qPCR检测M0、M1、M2各型巨噬细胞表面标记物，同时分别收集正常及高盐组巨噬细胞培养基，予以RT-qPCR及Elisa分别检测培养基中IL-6及TGF-β的mRNA表达与蛋白表达。随后建立巨噬细胞与NRK-49F的Transwell小室共培养体系。采用EdU及Transwell分别检测NRK-49F增殖及迁移能力。予以Western blot检测NRK-49F中collagen I、collagen III及α-平滑肌肌动蛋白(α-SMA)相对表达情况。结果 RT-qPCR结果显示，与对照组相比，高盐组细胞中表达M2型巨噬细胞表面标记物甘露糖受体（MR）和精氨酸酶（Arg）基因的mRNA水平显著升高（P<0.05）。EdU及Transwell结果显示，共培养体系中，高盐处理巨噬细胞组上层NRK-49F增殖、迁移能力增强（P<0.05）。Western blot结果显示，高盐处理组细胞培养基可诱导NRK-49F中collagen I、collagen III及和α-SMA蛋白表达增强（P<0.05）。此外，RT-qPCR及Elisa结果显示高盐处理组单核巨噬细胞培养基中IL-6与TGF-β1均显著高表达（P<0.05）。结论 高盐处理可诱导单核巨噬细胞像M2型极化并分泌IL-6与TGF-β1，从而诱导NRK-49F增殖及表型转化。

关键词: 高盐, 巨噬细胞, 肾脏成纤维细胞株, 细胞增殖, 表型转化

Abstract: Objective To investigate high-salt exposure-induced polarization of mononuclear macrophages and the changes in proliferation and phenotypic transformation of renal fibroblasts in a co-culture system. Methods Cultured mononuclear macrophages were exposed to high salt (161 mmol/L Na + ) for 2 h and the surface markers of M0, M1 and M2-type macrophages were detected with RT-qPCR. The culture medium of the macrophages in normal and high-salt groups was collected for detection of the mRNA and protein levels of IL-6 and TGF-β1 using RT-qPCR and ELISA. A co-culture system of high salt-exposed macrophages and renal fibroblasts (NRK-49F) was established using a Transwell chamber, and the changes in proliferation and migration of NRK-49F cells were examined using EdU assay and Transwell assay, respectively. Western blotting was performed to detect the expressions of collagen I, collagen III and collagen α-SMA in NRK-49F cells. Results The high salt-exposed macrophages showed significantly increased mRNA levels of M2-type macrophage surface markers mannose receptor and arginase (P<0.05). The results of EdU and Transwell assays showed that NRK-49F cells co-cultured with high salt-exposed macrophages exhibited significantly increased proliferation and migration ability (P<0.05). Co-culture with high salt-exposed macrophages resulted in significantly enhanced protein expressions of collagen I, collagen III and α-SMA in NRK-49F cells (P<0.05) and significantly increased levels of IL-6 and TGF-β1 in the culture medium (P<0.05). Conclusion High-salt exposure induces polarization of mononuclear macrophages into M2-type macrophages and promotes secretion of IL-6 and TGF-β1 by the macrophages to induce the proliferation and phenotypic transformation of NRK-49F cells. 

Key words: high salt, macrophages, renal fibroblasts, cell proliferation, phenotypic transformation