MicroRNA-326-5p enhances therapeutic potential of endothelial progenitor cells for myocardial infarction

Xiaoting Li,Ting Zhao,Yongbing Chen,Zhiwei Zhang,Wentao Yang,Lei Chen,Xiang Xue,Yuejun Sun

doi:10.1186/s13287-019-1413-8

Abstract

BackgroundOur study sought to investigate the therapeutic effects and mechanisms of miR-326-5p-overexpressing endothelial progenitor cells (EPCs) on acute myocardial infarction (AMI).MethodsMouse EPCs were isolated, purified, and identified by flow cytometry and uptake of DiI-ac-LDL. The target gene of miR-326-5p was predicted using target prediction algorithms and verified by dual-luciferase reporter assay, RT-qPCR, and Western blot. After EPCs were transfected with the agomir or antagomir of miR-326-5p, tube formation assay and Matrigel plug angiogenesis assay were conducted in four groups (NC, miR-326-5p agomir, miR-326-5p antagomir, and miR-326-5p agomir+Wnt1 agonist). In addition, a mouse model of MI was established and treated with the injection of miR-326-5p-EPCs, miR-326-5p-EPCs+ Wnt1 agonist, EPCs-NC, or PBS/control into the peri-infarcted myocardium. Subsequently, cardiac function was monitored by echocardiography at 7 and 28 days postoperatively. Finally, the infarcted hearts were collected at 28 days, and the size of myocardial infarction was measured by Masson’s trichrome staining and the neovascularization in the peri-infarcted area was examined through immunofluorescence staining.ResultsLuciferase reporter assay indicated that Wnt1 was a direct target of miR-326-5p. Using RT-qPCR and Western blot analysis, we further demonstrated that the expression level of Wnt1 was negatively correlated with miR-326-5p expression in EPCs. Both in vitro study of tube formation assay and in vivo investigation of subcutaneous Matrigel plug assay revealed that the miR-326-5p agomir could significantly enhance the angiogenic capacity of EPCs, and this effect was partially inhibited by Wnt1 agonist. Meanwhile, miR-326-5p antagomir could obviously reduce the the angiogenic capacity of EPCs in vivo compared with that in the NC group. Moreover, the transplantation of miR-326-5p-overexpressing EPCs in the ischemic hearts of mice significantly enhanced the angiogenesis in the peri-infarcted zone and improved the cardiac function. However, the enhanced capacity of angiogenesis of miR-326-5p-overexpressing EPCs was remarkably neutralized by Wnt1 agonist, accompanied by the decreased improvement in cardiac function.ConclusionmiR-326-5p significantly enhanced the angiogenic capacity of EPCs. Transplantation of miR-326-5p-overexpressing EPCs improved cardiac function for AMI therapy, which can be a novel strategy for enhancing therapeutic angiogenesis in ischemic heart diseases.

Highlights

Our study sought to investigate the therapeutic effects and mechanisms of miR-326-5p-overexpressing endothelial progenitor cells (EPCs) on acute myocardial infarction (AMI)
Ander et al further confirmed in the PROCELL study [7] that EPCs could be significantly mobilized from the bone marrow into the peripheral circulation after AMI, which was positively correlated with the level of vascular cell adhesion molecule (VCAM)-1 in plasma
The surface markers of EPCs were further identified through flow cytometry, which were positive for CD31, CD133, and VE-cadherin and negative for CD11b, Flk-1, and CD45 (Fig. 1c)

Summary

Introduction

Our study sought to investigate the therapeutic effects and mechanisms of miR-326-5p-overexpressing endothelial progenitor cells (EPCs) on acute myocardial infarction (AMI). Previous studies have reported that several types of stem cells safely and efficiently improved the neovascularization in the local ischemic region [3], especially endothelial progenitor cells (EPCs) [4, 5]. Ander et al further confirmed in the PROCELL study [7] that EPCs could be significantly mobilized from the bone marrow into the peripheral circulation after AMI, which was positively correlated with the level of vascular cell adhesion molecule (VCAM)-1 in plasma. Direct transplantation of EPCs into the ischemic myocardium has been conducted and exhibited cardiac protective effects, including augmenting neovascularization, decreasing inflammatory apoptosis, and improving cardiac function [4, 10]

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Results

Conclusion