Molecular Basis of Abnormal Conduction in Mice Over-expressing Endothelin-1

Binary transgenic (BT) mice with doxycycline (DOX)-suppressible cardiac-specific over-expression of endothelin 1 (ET 1) exhibit progressive heart failure, QRS prolongation, and death following DOX withdrawal. However, the molecular basis and reversibility of the electrophysiological abnormalities in this model were not known. Here we assess the mechanisms underlying ET 1 mediated electrical remodelling, and its role in heart failure. Prior attempts to prevent this model of ET-1 induced cardiomyopathy with ET receptor antagonism were not beneficial. We now propose to evaluate the effectiveness of blocking the synthesis of ET-1 with CGS 26303, a dual inhibitor of endothelin converting enzyme (ECE) and neutral endopeptidase. BT vs. littermate control mice were withdrawn from DOX and serially studied with ultrasound biomicroscopy, octapolar catheters, multi-electrode epicardial mapping, histopathology, Western blot, immunohistochemistry and qRT-PCR. Prolonged ventricular activation and depressed rate of ventricular activation were detected as early as 4 wks after transgene activation, when structure and function of the heart remained unaffected. By 8 wks of ET-1 over-expression, biventricular systolic and diastolic dysfunction, myocardial fibrosis, cardiomyocyte hypertrophy, prolonged ventricular activation and repolarization, depressed rate of ventricular activation, and abnormal atrioventricular nodal function were observed. Within 4 wks of ET-1 induction, reduction were observed in connexin-43 mRNA, protein, and phosphorylation, Nav1.5 mRNA and protein, Na+ conductance, K+ channel interacting protein-2 mRNA and Kv4.2 mRNA. Chromatin immunoprecipitation revealed that nuclear factor κB preferentially binds to Cx43 and Nav1.5 promoters. Importantly, the associated electrophysiological abnormalities at this time point were reversible upon suppression of ET 1 over-expression and completely prevented the development of structural and functional remodelling. Treatment with CGS-26303 (5 mg/kg/day) failed to improve survival, or hemodynamic and contractile decline. ET-1-mediated ventricular conduction delays correlates with gap junction and ion channel remodelling, and precedes heart failure. The sequence and reversibility of this phenotype suggest that a primary abnormality in electrical remodelling may contribute to the pathogenesis of heart failure. CGS 26303 failed to prevent this cardiomyopathic phenotype. These data suggest that chronically high levels of bigET-1, as seen in heart failure, may induce increased ECE activity and/or non-ECE ET-1 synthesis, thus circumventing the efficacy of ECE blockade in this model.Ph