黄芪甲苷调控内皮祖细胞外泌体miR-126-5p对高糖受损内皮细胞生物学功能及血管生长因子的影响OA

Objective:To investigate the effects of astragaloside Ⅳ(AS-Ⅳ)-regulated miR-126-5p in exosomes derived from endothelial progenitor cells(EPCs)on the biological functions and vascular growth factors of high-glucose-damaged human umbilical vein endothelial cells(HUVECs).Methods:Human EPCs and HUVECs were isolated,cultured and characterised.Characterised EPCs were treated with PBS or AS-Ⅳ,respectively,and cultured for 24 hours before collecting EPC exosomes.Their morphology was observed using transmission electron microscopy.EPCs were transfected with a miR-126-5p inhibitor or a inhibitor-NC,followed by treatment with AS-Ⅳ and exosome extraction.The levels of miR-126-5p in the exosomes were detected using real-time fluorogenic quantitative PCR(RT-qPCR).Following the establishment of a high-glucose injury model in successfully identified HUVECs,the cells were divided into high-glucose(HG)group,HG+EPCexos group,HG+AS-Ⅳ-EPCexos group,HG+miR-126-5p inhibitor+AS-Ⅳ-EPCexos group,and HG+inhibitor NC+AS-Ⅳ-EPCexos group,with a control group also included,making a total of six groups.After 48 hours of culture,cell proliferation,migration,tube formation and apoptosis were assessed using the CCK-8 assay,Transwell assay,Matrigel tube formation assay and flow cytometry,respectively.Western blotting was used to detect the protein expression of vascular endothelial growth factor A(VEGF-α),vascular endothelial growth factor B(VEGF-β),vascular endothelial growth factor C(VEGFc),fibroblast growth factor(FGF)and angiopoietin-(Ang-1).RT-qPCR was used to detect the expression of VEGFa mRNA,VEGFb mRNA,VEGFc mRNA,FGF mRNA and Ang-1 mRNA.Results:The exosomes exhibited a round or oval microvesicular structure.The HG+AS-Ⅳ-EPCexos group showed higher miR-126-5p expression than that HG+EPCexos group(P＜0.05).The HG+miR-126-5p inhibitor+AS-Ⅳ-EPCexos group showed lower miR-126-5p expression than HG+AS-Ⅳ-EPCexos group(P＜0.05).Compared with the control group,the HG group exhibited reduced relative viability,migration and tubule formation capacity in HUVECs,along with increased apoptosis rates and down-regulated expression of vascular growth factors(P＜0.05).Compared with the HG group,the HG+AS-Ⅳ-EPCexos group exhibited increased relative viability,migration and tubule formation capacity in HUVECs,along with reduced apoptosis rates and up-regulated expression of vascular growth factors(P＜0.05).Compared with the HG+AS-Ⅳ-EPCexos group,the HG+miR-126-5p inhibitor+AS-Ⅳ-EPCexos group exhibited reduced relative viability,migration and tube formation capacity in HUVECs,increased apoptosis rates,and down-regulated expression of vascular growth factors(P＜0.05).Conclusion:AS-Ⅳ can upregulate the expression of miR-126-5p in EPC-derived exosomes,thereby improving the angiogenic function of HUVECs damaged by high glucose.