Antagomir-mediated silencing of endothelial cell specific microRNA-126 impairs ischemia-induced angiogenesis

Abstract

MicroRNAs are negative regulators of gene expression that play a key role in cell-type specific differentiation and modulation of cell function and have been proposed to be involved in neovascularization. Previously, using an extensive cloning and sequencing approach, we identified miR-126 to be specifically and highly expressed in human endothelial cells (EC). Here, we demonstrate EC-specific expression of miR-126 in capillaries and the larger vessels in vivo. We therefore explored the potential role of miR-126 in arteriogenesis and angiogenesis. Using miR-reporter constructs, we show that miR-126 is functionally active in EC in vitro and that it could be specifically repressed using antagomirs specifically targeting miR-126. To study the consequences of miR-126 silencing on vascular regeneration, mice were injected with a single dose of antagomir-126 or a control ‘scramblemir’ and exposed to ischemia of the left hindlimb by ligation of the femoral artery. Although miR-126 was effectively silenced in mice treated with a single, high dose (HD) of antagomir-126, laser Doppler perfusion imaging did not show effects on blood flow recovery. In contrast, quantification of the capillary density in the gastrocnemius muscle revealed that mice treated with a HD of antagomir-126 had a markedly reduced angiogenic response. Aortic explant cultures of the mice confirmed the role of miR-126 in angiogenesis. Our data demonstrate a facilitary function for miR-126 in ischemia-induced angiogenesis and show the efficacy and specificity of antagomir-induced silencing of EC-specific microRNAs in vivo.

Regulation of Vascular Endothelial Growth Factor Signaling by miR-200b

“…Currently, miR-126, the miR-17-92 cluster, let-7b and -7f, miR130a, miR-210, miR-378, and miR-296 are known as proangiogenic miRNAs, while anti-angiogenic miRNAs include miR-221/miR-222, miR-328, miR-15b, miR-16, miR-20a, and miR-200b (Chan et al, 2011;Wu et al, 2009). Among proangiogenic miRNAs, miR-126 is highly expressed in human endothelial cells (van Solingen et al, 2009;Wang et al, 2008). Recent studies showed that pro-angiogenic function of miR-126 is in part mediated by down-regulating Sprouty-related protein SPRED1 and phosphoinositol-3 kinase regulatory subunit 2 (PIK3R2/p85-β), which are inhibitors of angiogenic signaling (Fish et al, 2008;Wang et al, 2008).…”

MicroRNA miR-126-5p Enhances the Inflammatory Responses of Monocytes to Lipopolysaccharide Stimulation by Suppressing Cylindromatosis in Chronic HIV-1 Infection

“…1B and C). As the purity of CD14 ϩ monocytes in our study exceeded 97% based on CD14 staining, the predominant miR-126-5p quantified was likely derived from monocytes, although we were unable to exclude the possibility of contamination by predominant endothelial cells (ECs), which have been reported as the main source of the miR-126-3p/5p cluster (35)(36)(37)(38).…”

“…Indeed, a significant increase in several miRNAs in monocytes was observed in HIV-1 infection, among which miR-126-5p was the most upregulated member, especially in proinflammatory CD14 ϩ CD16 ϩϩ monocytes. The miR-126-3p/5p cluster is expressed predominately in ECs (35)(36)(37)(38) and can exert their functions in non-ECs (34,56,57). miR-126-5p has been shown to correlate with metastasis-free survival in breast cancer patients (33) and to promote endothelial proliferation and prevent atherosclerosis by suppressing delta-like 1 (Dlk1) (58).…”

Role of microRNAs in peripheral artery disease (Review)

“…miR-126 has been found to mediate CXCL12 production, which was enriched in apoptotic bodies, and induced vascular protection by repressing the function of the regulator of G protein signaling 16 (21). In addition, the beneficial role of miR-126 has been elucidated using the specificity of antagomir-induced silencing of miR-126 in vivo (22).…”