Abstract 351: Downregulation of Connexin43 by MicroRNA-130a Results in Cardiac Arrhythmias
MicroRNAs (miRs) are endogenous ~22 nucleotide RNA molecules that are now recognized as critical regulators of diverse physiological and pathological processes; however, studies of miRs and arrhythmogenesis remain sparse. Connexin43 (Cx43), a major cardiac gap junction protein, is critical for maintaining normal cardiac electrical conduction. One aspect of arrhythmia pathogenesis that has elicited great interest has been changes in Cx43 expression. Computational analysis of messenger RNAs predicted Cx43 to be a target for miR-130a. We hypothesized that overexpression of miR-130a in adult cardiomyocytes would result in cardiac arrhythmias. To study this, we used an inducible (tetracycline-off) bigenic system to study miR-130a overexpression in the adult mouse heart.
12 weeks after induction of miR-130a, 100% (16/16) of transgenic mice demonstrated a loss of sinus rhythm and irregular ventricular activity suggestive of atrial fibrillation on surface electrocardiograms (ECG). Nonsustained ventricular tachycardia was also observed during ECG recordings. Atrial contractions could not be detected by pulse wave Doppler. Intracardiac electrocardiography confirmed the presence of atrial arrhythmias. Western blot analysis demonstrated a 90% reduction in Cx43 levels and immunofluorescent staining confirmed a near complete loss of Cx43 in transgenic compared to littermate control hearts (n=12). Morphology of the intercalated disc was evaluated by electron microscopy and showed a loss of gap junctions in transgenic hearts. To validate Cx43 as a direct target of miR-130a, we performed in vitro target assays in 3T3 fibroblasts, known to express miR-130a. Using a luciferase reporter fused to the 3’UTR of Cx43 compared to a control (SV40) 3’UTR, we found a 55% ± 6% reduction in luciferase activity with the Cx43 3’UTR (p<0.01, n=9). Addition of an antisense miR-130a inhibitor resulted in a dose dependent loss of inhibitory activity of the Cx43 3’UTR reporter, with no changes seen in the control vector.
In conclusion, we have identified an unappreciated role of miR-130a as a direct regulator of Cx43. Overexpression of miR-130a results in the development of a variety of important cardiac arrhythmias including atrial fibrillation and ventricular tachycardia.
- © 2013 by American Heart Association, Inc.