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  • Endothelial progenitor cells-secreted extracellular vesicles containing microRNA-93-5p confer protection against sepsis-induced acute kidney injury via the KDM6B/H3K27me3/TNF-α axis.

Endothelial progenitor cells-secreted extracellular vesicles containing microRNA-93-5p confer protection against sepsis-induced acute kidney injury via the KDM6B/H3K27me3/TNF-α axis.

Experimental cell research (2020-07-18)
Zhonghua He, Haixia Wang, Lingju Yue
ABSTRACT

The pivotal pathogenetic role of microRNAs (miRs) in sepsis-induced acute kidney injury (AKI) has been demonstrated in mounting evidence. The functions of the target cells are regulated through the release of cells-encapsulated extracellular vesicles (Evs) into the extracellular space. The present study aims to elucidate the clinical significance as well as biological function of the endothelial progenitor cell (EPC)-derived Evs containing miR-93-5p in sepsis-induced AKI. We first established a cellular sepsis-induced AKI mouse model by treatment with lipopolysaccharide (LPS), and tested ectopic expression and depletion experiments in the model. Evs derived from miR-93-5p inhibitor-transfected EPCs (Evs/miR-93-5p inhibitor) were isolated, and co-cultured with HK2 cells to explore the effects of EPC-derived Evs overexpressing miR-93-5p on LPS-induced HK2 cell injury. The interaction between miR-93-5p and lysine (K)-specific demethylase 6B (KDM6B) was identified using dual-luciferase reporter assay, and ChIP was used to validate the relationship between KDM6B and tumor necrosis factor-α (TNF-α). Mice were made septic by cecal ligation and puncture (CLP), and then injected with Ev/miR-93-5p inhibitor to explore its functions in vivo. The results found that miR-93-5p and histone H3 Lys27 trimethylation (H3K27me3) were downregulated while KDM6B was upregulated in LPS-treated HK2 cells. EPC-derived Evs alleviated LPS-induced HK2 cell injury, while Ev/miR-93-5p inhibitor potentiated the cell injury in vitro. miR-93-5p was found to directly target KDM6B. Silencing KDM6B induced H3K27me3, inhibiting the activation of TNF-α, thereby weakening LPS-induced HK2 cell injury. EPC-derived Evs containing miR-93-5p attenuated multiple organ injury, vascular leakage, inflammation, and apoptosis in septic mice. In conclusion, the present study demonstrated that endothelial protection from EPC-derived Evs carrying miR-93-5p in sepsis-induced AKI, which was mediated by regulation KDM6BH/3K27me3/TNF-α axis.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
GSK-J4, ≥98% (HPLC)
Sigma-Aldrich
Protease Inhibitor Cocktail, for use with mammalian cell and tissue extracts, DMSO solution
Sigma-Aldrich
β-N-Acetylglucosaminidase Assay Kit, sufficient for 50 reactions (1 mL), sufficient for 500 reactions (100 μL)