Downregulation of connexin43 by microRNA-130a in cardiomyocytes results in cardiac arrhythmias

作者:O**oume Appledene; Calway Tyler; Broman Michael; McSharry Saoirse; Earley Judy; Kim Gene H*
来源:Journal of Molecular and Cellular Cardiology, 2014, 74: 53-63.
DOI:10.1016/j.yjmcc.2014.04.024

摘要

MicroRNAs (miRNAs) are now recognized as critical regulators of diverse physiological and pathological processes; however, studies of miRNAs and arrhythmogenesis remain sparse. Connexin43 (0(43), a major cardiac gap junction protein, has elicited great interest in its role in arrhythmias. Additionally, Cx43 was a potential target for miR-130a as predicted by several computational algorithms. This study investigates the effect of miR-130a overexpression in the adult heart and its effect on cardiac rhythm. Using a cardiac-specific inducible system, transgenic mice demonstrated both atrial and ventricular arrhythmias. We performed ventricular-programmed electrical stimulation and found that the alpha MHC-miR130a mice developed sustained ventricular tachycardia beginning 6 weeks after overexpression. Western blot analysis demonstrated a steady decline in Cx43 after 2 weeks of overexpression with over a 90% reduction in Cx43 levels by 10 weeks. Immunofluorescent staining confirmed a near complete loss of Cx43 throughout the heart. To validate Cx43 as a direct target of miR-130a, we performed in vitro target assays in 3T3 fibroblasts and HL-1 cardiomyocytes, both known to endogenously express miR-130a. Using a luciferase reporter fused to the 3'UTR of Cx43, we found a 52.9% reduction in luciferase activity in 3T3 cells (p < 0.0001) and a 47.6% reduction in HL-1 cells (p = 0.0056) compared to controls. Addition of an antisense miR-130a inhibitor resulted in a loss of inhibitory activity of the Cx43 3'UTR reporter. We have identified an unappreciated role for miR-130a as a direct regulator of Cx43. Overexpression of miR-130a may contribute importantly to gap junction remodeling and to the pathogenesis of atrial and ventricular arrhythmias.

  • 出版日期2014-9