Abstract 13272: Cardiomyocytes Expressing Arrhythmogenic Right Ventricular Cardiomyopathy Mutant Plakoglobin Exhibit Abnormal Responses to Mechanical Stress without Change in Mechanical Properties
Introduction: Arrhythmogenic right ventricular cardiomyopathy (ARVC) can be caused by mutations in desmosomal proteins. It is widely believed, but not proven, that abnormal intercellular adhesion is a critical aspect of disease pathogenesis. Accordingly, we characterized the effects of two ARVC-causing mutations in the desmosomal gene encoding plakoglobin (JUP) on biomechanical properties of cardiac myocytes.
Methods & Results: Neonatal rat ventricular myocytes (NRVM) were transfected with adenoviruses to express either of two different ARVC-causing mutations in JUP (2057del2 & S39_K40insS). Cell-cell adhesion strength was evaluated by lifting a confluent monolayer of NRVM using dispase dissociation, shearing the resultant myocyte sheet, and quantifying the number of fragments following shear. Cell stiffness was characterized with atomic force microscopy. The resulting force-deflection plots were used to extract the cellular elastic modulus. Interestingly, no differences in cell-cell adhesion strength or stiffness were observed in cells expressing mutant JUP, relative to controls. We next examined whether mutant JUP affects cell viability and the expression of JUP at cell junctions in response to physiologically relevant shear stresses (forces in the plane of the cell layer that mimic those in contracting myocardium). Control NRVM exhibited 3-fold increases in junctional JUP signal in response to oscillatory fluid shear (p=0.021; 0.6 dynes/cm2 peak shear for 4 hours; n=3) relative to unsheared controls. NRVM infected to express mutant JUP exhibited no significant increase in junctional JUP; additionally cells expressing the 2057del2 JUP mutant exhibited a 2.14-fold increase in apoptosis, relative to unsheared controls (p = 0.013; n=4). Inhibition of GSK-3β rescued junctional JUP upregulation in cells expressing mutant JUP, and returned apoptosis to control levels.
Conclusions: Our results demonstrate that ARVC-causing mutations in JUP do not alter the mechanical properties of myocyte stiffness or adhesion, but affect shear-induced junctional JUP levels and viability in NRVM. These results suggest that ARVC-causing mutations affect myocyte responses to mechanical stimuli rather than the biomechanical properties of the cells.
- © 2012 by American Heart Association, Inc.