A Novel Role for microRNA-214 in Inflammatory Smooth Muscle Cell Differentiation from Adventitia Stem/progenitor Cells and Arterial Remodelling

PhD ThesisBackground: Cardiovascular disease (CVD) is still the leading cause of death in the developed as well as developing country. Atherosclerosis and neointima formation considered as chronic inflammatory diseases, in which many cells including inflammatory smooth muscle cell (iSMC) were involved. ISMC differentiation from vascular adventitia stem/progenitor cells (AdSPCs) has been recently recognised as a critical determinant in cardiovascular diseases. One study have recently reported an important role for microRNA-214 (miR-214) in mature vascular SMC functions and injury-induced neointima formation. However, little is known about the functional involvements of miR-214 in iSMC differentiation from AdSPCs and its contribution to adverse arterial remodelling. Purpose: In this PhD study, I aimed to study the functional importance of miR-214 and its target gene in iSMC differentiation from AdSPCs and neointima SMC hyperplasia. Results: miR-214 expression was significantly increased during SMC differentiation from AdSPCs in response to TGFβ1. miR-214 gain/loss-offunction assays showed that miR-214 plays an important role in SMC differentiation from AdSPCs. By co-incubating AdSPCs with TGFβ1 and TNFα, AdSPCs were induced to differentiate toward iSMCs as evident by a decreased expression level of SMC-specific genes, but an increased level of inflammatory genes in iSMCs. Importantly, such phenotype could be reverted by miR-214 over-expression. I observed that sonic hedgehogglioma- associated oncogene 1 (Shh-GLI1) signal was closely modulated during iSMC differentiation, and demonstrated that Suppressor of Fused (SuFu) is the functional target of miR-214 controlling iSMC generation from AdSPCs. Further mechanistic studies revealed that increased amount of the GLI1 protein was translocalized into nuclei in miR-214 over-expressing or 6 SuFu knockdown cells, and the consensus sequence (GACCACCCA) of GLI1 within gene promoters of smooth muscle alpha-actin (SMαA) and serum response factor (SRF) was required for their regulation by miR-214 and SuFu. Additionally, Sufu also functions as a positive regulator for inflammatory gene regulations in iSMCs. In vivo, I demonstrated that locally enforced expression of miR-214 in the injured vessels significantly reduced SuFu expression level, iSMC generation, and inhibited neointima SMC hyperplasia after injury. Importantly, I observed that perivascularly transplanted AdSPCs significant contribute to neointima SMC hyperplasia by differentiating to iSMCs, and miR-214 over-expression could reverse such an effect. Finally, a decreased expression level of miR-214, but increased expression level of Sufu was observed in diseased human arteries. Conclusions/Implications: This thesis reports an unexplored role for miR- 214 in iSMC differentiation from AdSPCs and controlling neointima iSMC hyperplasia. These findings provide new insights into the therapeutic effect of miR-214 in vascular diseases