Progressive Reduction in Right Ventricular Contractile Function Attributable to Altered Actin Expression in an Aging Mouse Model of Arrhythmogenic Cardiomyopathy

Circulation. 2022 May 24;145(21):1609-1624. doi: 10.1161/CIRCULATIONAHA.120.049261. Epub 2022 Apr 19.

Abstract

Background: Arrhythmogenic cardiomyopathy (ACM) is an inherited genetic disorder of desmosomal dysfunction, and PKP2 (plakophilin-2) has been reported to be the most common disease-causing gene when mutation-positive. In the early concealed phase, the ACM heart is at high risk of sudden cardiac death before cardiac remodeling occurs because of mistargeted ion channels and altered Ca2+ handling. However, the results of pathogenic PKP2 variants on myocyte contraction in ACM pathogenesis remain unknown.

Methods: We studied the outcomes of a human truncating variant of PKP2 on myocyte contraction using a novel knock-in mouse model with insertion of thymidine in exon 5 of Pkp2, which mimics a familial case of ACM (PKP2-L404fsX5). We used serial echocardiography, electrocardiography, blood pressure measurements, histology, cardiomyocyte contraction, intracellular calcium measurements, and gene and protein expression studies.

Results: Serial echocardiography of Pkp2 heterozygous (Pkp2-Het) mice revealed progressive failure of the right ventricle (RV) in animals older than 3 months. By contrast, left ventricular function remained normal. ECGs of 6-month-old anesthetized Pkp2-Het mice showed normal baseline heart rates and QRS complexes. Cardiac responses to β-adrenergic agonist isoproterenol (2 mg/kg) plus caffeine (120 mg/kg) were also normal. However, adrenergic stimulation enhanced the susceptibility of Pkp2-Het hearts to tachyarrhythmia and sudden cardiac death. Histological staining showed no significant fibrosis or adipocyte infiltration in the RVs and left ventricles of 6- and 12-month-old Pkp2-Het hearts. Contractility assessment of isolated myocytes demonstrated progressively reduced Pkp2-Het RV cardiomyocyte function consistent with RV failure measured by echocardiography. However, aging Pkp2-Het and control RV myocytes loaded with intracellular Ca2+ indicator Fura-2 showed comparable Ca2+ transients. Western blotting of Pkp2-RV homogenates revealed a 40% decrease in actin, whereas actin immunoprecipitation followed by a 2,4-dinitrophenylhydrazine staining showed doubled oxidation level. This correlated with a 39% increase in troponin-I phosphorylation. In contrast, Pkp2-Het left ventricular myocytes had normal contraction, actin expression and oxidation, and troponin-I phosphorylation. Last, Western blotting of cardiac biopsies revealed that actin expression was 40% decreased in RVs of patients with end-stage ACM.

Conclusions: During the early concealed phase of ACM, reduced actin expression drives loss of RV myocyte contraction, contributing to progressive RV dysfunction.

Keywords: aging; arrhythmogenic right ventricular dysplasia; plakophilins; sarcomeres.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, N.I.H., Extramural

MeSH terms

  • Actins
  • Aging
  • Animals
  • Arrhythmogenic Right Ventricular Dysplasia* / pathology
  • Cardiomyopathies* / genetics
  • Death, Sudden, Cardiac
  • Disease Models, Animal
  • Humans
  • Mice
  • Plakophilins / genetics
  • Troponin I

Substances

  • Actins
  • Plakophilins
  • Troponin I