Abstract 16692: MicroRNA 1825 Acts as a Master Regulator Inducing Proliferation of Adult Cardiomyocytes and Regenerates Ischemic Myocardium
Background: While cardiomyocytes (CMs) in zebrafish and neonatal mice are known to proliferate and regenerate the infarcted heart, this capacity is lost in adult mammals. From our unpublished RNA sequencing data using rat neonatal CMs (NCM) transfected with miR-1825 we identified miR-99a and miR-199a to be downstream effectors of miR-1825. In this study we aim to elucidate the mechanism and relative contribution of each of these miRs in inducing adult CM (ACM) proliferation.
Methods and Results: CMs were isolated from 8-12 week old adult rat hearts and transfected with miR-cel-67 (control), miR-1825, miR-99a, and miR-199a. To label proliferating CMs EDU was added daily. On day 5 cells were fixed and stained for EDU, and Troponin I (TnI) (CM marker). MiR-1825 transfected CMs showed 50±5% increase in proliferation compared to control 11.2±3%, while miR-99a and miR-199a showed an increase of 14±3% and 16±2%, respectively. To identify the relative contribution of these downstream miRs additional experiments using respective miRNA inhibitors were carried out revealing proliferation of 40±4% with miR-1825+anti-miR-99a and 22±3% with miR-1825+anti-miR-199a, which were lower than miR-1825 alone. These results indicate that miR-1825 works both independently as well as through miR-99a and miR-199a. To elucidate a mechanism for miR-1825 mediated proliferation of ACM, we performed protein analysis and found a significant reduction in expression of p16, while cyclin-D1 was significantly increased in miR-1825 transfected ACMs. To further identify in-vivo proliferative potential, following LAD ligation we transduced 12 week old (C57BL/6) mice hearts with miR-1825 and miR-cel-67(control). EDU was administered IP up to day 12 post surgery. Four weeks after LAD ligation mice were euthanized and hearts were perfused and harvested for histochemical studies. A significant increase in proliferating ACMs was seen in heart receiving miR-1825 (11.4±3.6% ) vs miR-cel-67 control (1.35± 0.5%).
Conclusion: We determine that miR-1825 acts as a master regulator of downstream miRs, and induces proliferation of ACMs through regulation of cyclin-D1 and p16. Overall, this project determines a hierarchical role of miR-1825 and elucidates a novel mechanism of ACM proliferation.
Author Disclosures: R. Pandey: None. S. Durrani: None. S. Jiang: None. U. Savani: None. R.P. Ahmed: None.
- © 2014 by American Heart Association, Inc.