Abstract
MicroRNAs regulate endothelial function and angiogenesis, but their implication in pericyte biology remains undetermined. A PCR array, covering a panel of 379 human microRNAs, showed microRNA-532-5p to be one of the most differentially modulated by hypoxia, which was confirmed by qPCR in both skeletal muscle and adventitial pericytes. Furthermore, microRNA-532-5p was upregulated in murine muscular pericytes early after experimentally induced ischemia, decreasing below baseline after reperfusion. Transfection of human pericytes with anti-microRNA, microRNA-mimic, or controls indicates microRNA-532-5p modulates pro-angiogenic activity via transcriptional regulation of angiopoietin-1. Tie-2 blockade abrogated the ability of microRNA-532-5p-overexpressing pericytes to promote endothelial network formation in vitro. However, angiopoietin-1 is not a direct target of microRNA-532-5p. In silico analysis of microRNA-532-5p inhibitory targets associated with angiopoietin-1 transcription indicated three potential candidates, BACH1, HIF1AN, and EGLN1. Binding of microRNA-532-5p to the BACH1 3′ UTR was confirmed by luciferase assay. MicroRNA-532-5p silencing increased BACH1, while a microRNA-532-5p mimic decreased expression. Silencing of BACH1 modulated angiopoietin-1 gene and protein expression. ChIP confirmed BACH1 transcriptional regulation of angiopoietin-1 promoter. Finally, microRNA-532-5p overexpression increased pericyte coverage in an in vivo Matrigel assay, suggesting its role in vascular maturation. This study provides a new mechanistic understanding of the transcriptional program orchestrating angiopoietin-1/Tie-2 signaling in human pericytes.
Highlights
Angiogenesis is a tightly regulated process that involves the interaction between endothelial cells (ECs) and pericytes (PCs).[1]
The present study investigates the effect of hypoxia on global miR expression in a subset of human CD34+ and CD146À PCs isolated from the saphenous vein—adventitial PCs (APCs)
We previously showed that miR-132 plays a role in the pro-angiogenic and antifibrotic activity of APCs in a murine model of myocardial infarction, respectively, through inhibition of Ras-GTPase-activating protein and methyl-CpG-binding protein 2.16 The hypoxia-miR Let-7d was found upregulated in PC-derived neurospheres and proposed to be involved in PC differentiation.[17]
Summary
Angiogenesis is a tightly regulated process that involves the interaction between endothelial cells (ECs) and pericytes (PCs).[1]. MicroRNAs (miRs) are small noncoding RNAs that function in diverse biological processes via post-transcriptional gene regulation They modulate protein expression by binding to complementary or partially complementary target sites in the 30 UTRs of mRNA, causing inhibition of gene expression.[6] miRs have been identified in cells,[6] serum, and plasma.[7] Some miRs remain within the cell-modifying transcriptional regulation, while others are released, acting like paracrine or endocrine factors.[8,9] Changes in the expression profile of several miRs have been associated with cardiovascular processes and diseases.[10,11] In particular, a class of miRs is modulated by hypoxia and implicated in the regulation of angiogenesis. A few studies have addressed the participation of miRs in the regulation of PC functions.[13,14,15,16] some data exist on the hypoxia-induced Let-7d having a role in PC differentiation,[17] no systematic analysis has been carried out to characterize the profile of hypoxia-miRs that target angiogenic genes in human PCs
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