Abstract 14420: MicroRNA-214 Regulates Stem Cell Differentiation Into Smooth Muscle Cells by Targeting Quaking
Background: Smooth muscle cell (SMC) differentiation is a critical process during vascular development and SMC proliferative related vascular disease. Several microRNAs have been reported to be involved in SMC differentiation. Recently, the microRNA-214 (miR-214) was shown to regulate cell cycle progression and apoptosis. However, very little is known about the functional role of miR-214 in SMC differentiation. The present study investigated the role of miR-214 during SMC differentiation from stem cells and verified its targets.
Methods and results: Mouse embryonic stem cells (ES cells) were seeded on collagen IV-coated flasks and cultured in the absence of Leukemia Inhibitory Factor (LIF) in differentiation medium for 4 to 8 days in order to induce SMC differentiation. We found that miR-214 was significantly upregulated during SMC differentiation by using real-time RT-PCR analysis. Furthermore, enforced expression of miR-214 by its mimic in differentiating ES cells was found to induce expression of SMC-specific markers. Similarly, Western blot analysis revealed the increased protein production of these genes in the cells overexpressing miR-214. The knock-down of miR-214 by its antagomiR on the other hand, inhibited expression of smooth muscle differentiation specific markers at both RNA and protein levels. Furthermore, we have found that Quaking (QKI), a protein critical for vascular development and remodeling, is a novel target of miR-214 during SMC differentiation from ES cells. Overexpression of miR-214 was found to repress QKI in differentiating ES cells at both RNA and protein levels, whereas miR-214 antagomir resulted in increased QKI expression. Luciferase assay showed miR-214 substantially inhibited QKI-3’-UTR-luciferase activity in differentiating ES cells, but not mutant QKI-3’-UTR-luciferease reporter. Finally, we tested the expression of miR-214 in mouse femoral artery denudation injury model and found that miR-214 was significantly down-regulated in the wire injury-induced neointimal formation.
Conclusion: Using genomic and functional studies, we have discovered miR-214 controls differentiation via regulating QKI expression during ES cell differentiation towards SMCs.
- © 2013 by American Heart Association, Inc.