Background: Arrhythmogenic cardiomyopathy (ACM) is an inherited heart disease caused by defective cellular junctions, in particular cardiac desmosomes. Clinically ACM patients present with fibrofatty remodeling, cardiac dysfunction, arrhythmia and propensity for sudden death. Mutations in Plakoglobin (Pg), an important component of cell-cell junctions, were the first linked to ACM. However, the underlying pathomechanisms are only poorly understood.

Purpose: Recent findings in other ACM mouse models suggest impaired mitochondrial function, enhanced formation of reactive oxygen species (ROS) formation and dysregulated Ca²⁺ handling to play a crucial role for disease onset and progression. We used 4- and 6-week-old cardiac-restricted Pg knockout (Pg KO) mice to assess the role of Pg for cardiac function in vivo and analyzed mitochondrial energetics and excitation-contraction coupling along with their dynamic interplay.

Methods and Results: Pg KO mice largely recapitulate the human ACM phenotype: they develop early onset and progressive cardiomyocyte (CM) loss, fibrosis, biventricular dilatation and systolic dysfunction. Microscopy of cardiac tissue and isolated CMs showed concentric hypertrophic growth. High-resolution respirometry using freshly isolated mitochondria showed that mitochondrial electron transport chain function, ROS emission, and the membrane potential (∆Ψ_m) did not differ between Pg KO and WT. Isolated electrically stimulated Pg KO CMs displayed reduced diastolic sarcomere length at baseline, and after ß-adrenergic stimulation and an increase in stimulation frequency, indicating altered cellular diastolic function. Unlike in vivo, systolic cellular function was preserved.  Additionally and also in contrast to findings on organ level, an increase in the cellular arrhythmic index was observed. In opposition to other murine desmosomal KO models, cytosolic Ca²⁺ [Ca²⁺]_c concentrations did not differ between genotypes. Plotting sarcomere lengths against the respective [Ca²⁺]_c resulted in similar slopes, suggesting changes at the myofilament rather than altered Ca²⁺ affinity to underlie the observed cellular diastolic sarcomere shortening. However, IF microscopy using respective antibodies demonstrated intact myofilaments in Pg KO CMs. Analysis of mitochondrial NADP(H) and FAD autofluorescence revealed a more oxidized NAD(P)H/FAD redox state in Pg KO CMs during baseline and workload transition, suggesting modestly reduced bioenergetic adaption. ∆Ψ_m, determined by TMRM fluorescence, did not differ between genotypes. Albeit overall ROS emission in CMs using H₂DCF-DA was similar between groups, exposure to an excess amount of H₂O₂ led to an increased DCF-signal in Pg KO CMs, indicating altered endogenous anti-oxidative capacity.

Conclusion: Our data show that Pg-deficiency induced an early onset ACM like phenotype in vivo, accompanied by altered cellular diastolic function and modestly impaired bioenergetic adaption, while defects in mitochondrial function and Ca²⁺ cycling could not be detected. We therefore rule out these factors as global key drivers for disease onset and progression.