Cardiovascular phenotype of the duchenne muscular dystrophy carrier female rat

Abstract

Abstract Background Duchenne muscular dystrophy (DMD) is a lethal, severely progressive muscle-wasting disease. It is an X-linked recessive disease caused by mutations in the DMD gene encoding dystrophin protein and consequently the absence of dystrophin protein synthesis. Besides the skeletal muscle abnormalities, DMD patients are often suffering cardiovascular symptoms include cardiac fibrosis, dilated cardiomyopathy, arrhythmia, and congestive heart failure. Heterozygous female DMD carriers rarely acquire skeletal muscle symptoms, however the cardiac abnormalities are often underrepresented. There are few studies to date characterizing cardiovascular abnormalities in female DMD carriers and the mechanisms underlying the cardiac manifestations are still not fully known. Aims and Methods: Therefore, the aim of this study was to investigate the cardiovascular phenotype of 9 months old female DMD carrier rats and compare with age-matched wildtype rats. Objectives included assessment of cardiac function via transthoracic echocardiography, vascular function using wire myography, cardiac fibrosis and dystrophin via histology, mRNA expression of inflammatory markers via RT-qPCR, measurements of ACE1 and ACE2 activity, as well as expression of dystrophin, SERCA, sarcolipin, BAG3, and Connexin43 (Cx43) using western blotting. The cardiac function, vascular endothelial function, ACE1 and ACE2 activity, and collagen deposition in cardiac tissue were similar between the female DMD carrier and wildtype rats. The echocardiography data revealed thinning of the left-ventricular posterior wall indicating trend towards cardiac atrophy and dysfunction (p<0.05). Wire myography data showed trends of impaired endothelium-independent relaxation and hypercontractility, suggesting the potential vascular abnormalities. Histological analysis showed thickening of intima-media layer in coronary arteries, addition to reduced dystrophin staining, further affirming dystrophin loss impairs smooth muscle cell function (p<0.05, respectively). Protein expression of dystrophin, SERCA2, and BAG3 was decreased in female DMD carrier rats in the myocardium. We also found decreased levels of Cx43 at the intercalated discs, with a concomitant increase in the phosphorylation of Serine368 on Cx43 that very often precedes ubiquitination and degradation of Cx43 channels. In agreement, higher levels of the ubiquitin ligase Nedd4 were observed in hearts of female DMD carrier rats, implicating Cx43 degradation in cardiac dysfunction observed in these animals. Conclusions These findings indicate that female DMD carrier rats do not develop severe cardiac systolic dysfunction nor dilated cardiomyopathy at the age of 9 months unlike male DMD rats as previously described(1). However, the female DMD carrier rats begin to manifest tendencies towards cardiac and vascular dysfunction in association with a changes of calcium handling protein and Cx 43 expression in the heart.