Abstract 15187: Shear Stress-regulated Mir-27b Controls Pericyte Recruitment in vitro and in vivo and Contributes to Vessel Maturation

Abstract

Vessel maturation involves recruitment of mural cells. Laminar shear stress is a major trigger for vessel maturation. However, the molecular mechanisms by which shear stress affects recruitment of pericytes are unclear. MicroRNAs are small non-coding RNAs, which post-transcriptionally control gene expression. Since shear stress regulates various miRs, we hypothesize that flow-induced miRs inhibit repulsive cues and facilitate mural cell coverage. Laminar shear stress for 72h induces the up-regulation of miR-27b (2.8±0.24-fold vs static, p<0.05) in cultured endothelial cells (ECs) and in mouse femoral artery segments that were exposed to physiological shear stress ex vivo (1.5±0.14-fold vs no flow, p<0.05). Predicted targets for miR-27b include members of the semaphorin (SEMA) family (known to regulate repulsive signaling) and angiopoietin-2 (Ang2), which causes vessel destabilization. MiR-27b overexpression reduces SEMA6A (63.5±13.5%), SEMA6D (58±26%), and Ang2 (51.5±11%) expression. To determine whether miR-27b controls pericyte recruitment, we tested the effect of endothelial miR-27b on adhesion of pericytes. Overexpression of miR-27b increased the adhesion of pericytes (p<0.05) and inhibition of miR-27b reduced pericyte adhesion in vitro (p<0.05). In an in vitro matrigel assay, overexpression of miR-27b increased pericyte coverage of endothelial cell tubes (155±45%, p<0.05), whereas inhibition of miR-27b reduced pericyte coverage (81±15%). In addition, overexpression of miR-27b in vitro increases barrier function (p<0.05) in endothelial cell-pericyte co-culture assays whereas inhibition of miR-27b decreases barrier function (p<0.05). Silencing of SEMA6A and SEMA6D rescued the reduced pericyte adhesion that is caused by miR-27 inhibition (p<0.05). Furthermore, inhibition of SEMA6D increases barrier function. In vivo inhibition of miR-27a/b significantly increased edema (p<0.05) in the uterus which was associated with a significantly reduced pericyte coverage of the vessels (p<0.05). Taken together, our data demonstrate that shear stress regulate miR-27b, which targets Ang2 and semaphorins, thereby controlling pericyte adhesion and coverage in vitro and in vivo.