脂肪来源干细胞衍生的外泌体 miR-215-5p 通过抑制. 来减轻足细胞的上皮-间充质转化。

Exosomal miRNA-215-5p Derived from Adipose-Derived Stem Cells Attenuates Epithelial-Mesenchymal Transition of Podocytes by Inhibiting .

机构信息

Department of Nephrology, Zhejiang Provincial People's Hospital, Zhejiang 310014, China.

People's Hospital of Hangzhou Medical College, Zhejiang 310014, China.

出版信息

Biomed Res Int. 2020 Feb 21;2020:2685305. doi: 10.1155/2020/2685305. eCollection 2020.

DOI:10.1155/2020/2685305

PMID:32149094

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC7057016/

Abstract

BACKGROUND

Podocyte migration is actively involved in the process of podocyte loss and proteinuria production, which is closely associated with the development of diabetic nephropathy (DN). Exosomes from adipose-derived stem cells (ADSCs-Exos) effectively inhibit podocyte apoptosis in the treatment of DN. However, how ADSCs-Exos affect the migration of podocytes is obscure. This study is aimed at exploring the regulatory role of ADSCs-Exos on cell migration and the underlying mechanism.

METHODS

ADSCs-Exo was authenticated by transmission electron microscopy (TEM), western blotting, and flow cytometry. Cell viability and migration ability of podocytes were measured by CCK8 and Transwell assays, respectively. Relative expressions of miRNAs and mRNAs were determined by qRT-PCR. The transmitting between PKH26-labeled exosome and podocytes was evaluated by IF assay. Dual luciferase reporter assay was employed to detect the relationship between miR-215-5p and .

RESULTS

The exposure to serum from DN patient (hDN-serum) significantly inhibited cell viability of podocytes, but ADSCs-Exo addition notably blunts cytotoxicity induced by the transient stimulus of hDN-serum. Besides, ADSCs-Exo administration powerfully impeded high glucose- (HG-) induced migration and injury of podocyte. With the podocyte dysfunction, several miRNAs presented a significant decline under the treatment of HG including miR-251-5p, miR-879-5p, miR-3066-5p, and miR-7a-5p, all of which were rescued by the addition of ADSCs-Exo. However, only miR-251-5p was a key determinant in the process of ADSCs-Exo-mediated protective role on podocyte damage. The miR-251-5p inhibitor counteracted the improvement from the ADSCs-Exo preparation on HG-induced proliferation inhibition and migration promotion. Additionally, miR-215-5p mimics alone remarkably reversed HG-induced EMT process of podocyte. Mechanistically, we confirmed that ADSCs-Exos mediated the shuttling of miR-215-5p to podocyte, thereby protecting against HG-induced metastasis, possibly through inhibiting the transcription of ZEB2.

CONCLUSION

ADSCs-Exo has the protective effect on HG-evoked EMT progression of podocytes thru a mechanism involving ZEB2. Potentially, the ADSCs-Exo preparation is a useful therapeutic strategy for improving podocyte dysfunction and DN symptoms clinically.

摘要

背景

足细胞迁移积极参与足细胞丢失和蛋白尿产生的过程，这与糖尿病肾病（DN）的发展密切相关。脂肪来源干细胞（ADSCs）衍生的外泌体（ADSCs-Exos）在治疗 DN 中有效抑制足细胞凋亡。然而，ADSCs-Exos 如何影响足细胞的迁移尚不清楚。本研究旨在探讨 ADSCs-Exos 对细胞迁移的调节作用及其潜在机制。

方法

通过透射电子显微镜（TEM）、Western blot 和流式细胞术对 ADSCs-Exo 进行鉴定。通过 CCK8 和 Transwell 测定分别测量足细胞的细胞活力和迁移能力。通过 qRT-PCR 测定 miRNA 和 mRNA 的相对表达。通过 IF 测定评估 PKH26 标记的外泌体与足细胞之间的传递。采用双荧光素酶报告基因检测 miR-215-5p 与. 之间的关系。

结果

DN 患者血清（hDN-血清）的暴露显著抑制了足细胞的细胞活力，但 ADSCs-Exo 的添加显著减弱了 hDN-血清短暂刺激引起的细胞毒性。此外，ADSCs-Exo 给药强力抑制高糖（HG）诱导的足细胞迁移和损伤。随着足细胞功能障碍，几种 miRNA 在 HG 处理下显著下降，包括 miR-251-5p、miR-879-5p、miR-3066-5p 和 miR-7a-5p，所有这些 miRNA 都被 ADSCs-Exo 的添加所挽救。然而，只有 miR-251-5p 是 ADSCs-Exo 介导的足细胞损伤保护作用过程中的关键决定因素。miR-251-5p 抑制剂拮抗了 ADSCs-Exo 制剂对 HG 诱导的增殖抑制和迁移促进的改善作用。此外，miR-215-5p 模拟物单独显著逆转了 HG 诱导的足细胞 EMT 过程。机制上，我们证实 ADSCs-Exos 通过将 miR-215-5p 转运到足细胞中来介导对 HG 诱导的转移的保护作用，可能通过抑制 ZEB2 的转录。