P.309Mechanistic insights into Duchenne muscular dystrophy-associated cardiomyopathy

Abstract

Duchenne muscular dystrophy (DMD) is an X-linked neuromuscular disorder that is associated with progressive muscle wasting and cardiomyopathy. Utilizing cardiac MRI, we have demonstrated that adult DMD patients have small hearts with very low LV mass as compared to patients with non-ischemic cardiomyopathy or healthy patients. Thus, we hypothesize that DMD-associated cardiomyopathy develops secondary to dysregulation of cardiac signaling pathways that modulate cardiac growth, namely cardiomyocyte proliferation. Utilizing the mdx mouse, a model of DMD, decreased heart/body weight ratios were noted from birth (P1: Hrt/BW 5.6 ± 0.16 vs. 4.8 ± 0.18 NTg vs mdx p<0.005 n=32). NTg and mdx hearts have similar cardiomyocyte cell size (WGA staining: 0.08 ± 0.001 vs 0.08 ± 0.002 NTg vs mdx p<0.05; n=3). FACS analysis indicated mdx hearts have fewer cardiomyocytes (35% reduction; 51.7e ± 4.9e vs. 33.8e ± 3.5e NTg vs mdx p<0.05 n=6). Transcriptome profiling demonstrated that molecular pathways governing cardiac proliferation were significantly reduced in mdx hearts (Ki67: 4.8 ± 0.3 vs 2.0 ± 0.1 NTg vs mdx p<0.005 n=3), with a corresponding increase in cardiac atrophy gene expression at later time points (Foxo3 1.4 ± 0.2 vs 2.8 ± 0.3 NTg vs mdx p<0.005 n=3). Likewise, immunohistochemical (IHC) analyses showed significantly reduced proliferation in mdx hearts (Ki67: 13.7 ± 1.2 vs. 6.0 ± 1.0 NTg vs mdx p<0.005, n=3; pH3: 118 ± 5.3 vs 59 ± 4.5 p<0.005 n=3). RNA-Seq data revealed a disruption of the Yap signaling pathway, an important co-transcriptional mediator of cardiomyocyte proliferation, in mdx P4 hearts. By quantitative RT-PCR, cell cycle targets of Yap were reduced in mdx hearts. Finally, western blot and IHC revealed less nuclear localization of Yap in mdx cardiomyocytes. Collectively, this data suggests a novel mode of cardiac remodeling that contributes to the development of DMD-associated cardiomyopathy.