Role of Pannexin Channels in Arrhythmogenesis and Cardiac Fibrosis in Models of Duchenne Muscular Dystrophy

Vanderbilt University Medical Center, Nashville TN

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Abstract

Project Summary Cardiovascular disease is the primary cause of death for patients with Duchenne muscular dystrophy (DMD). Arrhythmia and cardiac fibrosis leading to dilated cardiomyopathy are the primary mechanisms of cardiac mortality. Pannexins (Pxs), which are large conductance ion and small molecule channels, have been implicated in other fibroproliferative diseases and are thought to be arrhythmogenic in other model of cardiac disease. Loss of dystrophin, the primary defect in DMD, leads to elevated intracellular calcium (Ca2+) which is also a primary effector of Pxs. The goal of this project is to investigate the role of Pxs in the development of cardiac fibrosis and arrhythmogenesis in mdx transgenic mouse model of DMD (Aim 1) human DMD induced pluripotent stem cells (iPSC) cardiomyocytes (Aim 2). Aim 1A is to identify if Px current contributes to arrhythmogenesis via transient inward current (Iti) and delayed after depolarizations (DADs) in mdx mice using patch clamp electrophysiology, pharmacological blockade, and mdxPx1-/- and mdxPx1-/-Px2-/- transgenic lines. The primary end points will be single channel conductance, average open probability, reversal potential and total current integral. Aim 1B is to test if Px channels facilitate triggered propagated electrical activity and cardiac fibrosis in intact mdx and transgenic hearts using telemetric, molecular techniques (RT-PCR/Western Blot), and histologic evaluation. The primary end points will be rate of premature ventricular contractions (PVCs) and ventricular tachycardia (VT), gross/histologic burden of fibrosis, and relative expression of ATP and other fibrotic mediators. Aim 2A is to test if Px channels contribute to arrhythmogenesis via Iti and DADs in human DMD iPSC cardiomyocytes using patch clamp electrophysiology and pharmacological blockade. The primary end points will be single channel conductance, average open probability, reversal potential and total current integral. Aim 2B is to identify the mechanism of Px channels in modulating mediators of cardiac fibrosis in DMD iPSC cardiomyocytes using transwell co-culture with fibroblasts, pharmacological blockade, and molecular techniques (RT-PCR/Western Blot). The primary end points will be cell migration index and relative expression of ATP and other fibrotic mediators. Cumulatively, this investigation will help to determine what role Pxs play in the development of cardiac fibrosis and pro-arrhythmic electrophysiological changes in DMD. Pxs may represent a novel therapeutic target for delaying or preventing cardiac fibrosis and arrhythmia in patients with DMD.