Abstract 4359395: Cardiac Function is Preserved in a Tissue-Engineered Model of LGMD2B

Abstract

Background: Loss of dysferlin results in the rare, currently untreatable muscular dystrophy known as Limb Girdle Muscular Dystrophy 2B (LGMD2B). In LGMD2B mice, skeletal muscle undergoes progressive myopathy, whereas cardiac deficits only arise with advanced age, stress, or injury – suggesting the heart may harbor protective mechanisms that could guide future therapy developments. While dysferlin’s roles in membrane repair, Ca 2+ handling, and metabolism are well characterized in skeletal muscle, its function in the heart is poorly defined. We therefore engineered 3D LGMD2B cardiac and skeletal muscle tissues (“cardio- and myobundles”) to compare dysferlin’s differential roles in cardiac vs. skeletal muscle. Methods: Three healthy (HLT) and three LGMD2B human induced pluripotent stem cell (hiPSC) lines were differentiated into cardiomyocytes (hCMs) and muscle progenitor cells to generate cardio- and myobundles. After 2 weeks of culture, we performed isometric force tests, Ca 2+ transient imaging, and optical mapping of action potential propagation. To probe membrane repair capacity, osmotic shock injury (OSI) was induced with ~30 mOsm medium for 5 min followed by 15 min recovery, with contractile force recorded every minute. Tissues were also immunostained for sarcomere structure and dysferlin localization. Results: Both HLT and LGMD2B cardio- and myobundles exhibited aligned, cross-striated sarcomeric structure with dysferlin predominantly localized at the plasma membrane. Dysferlin-deficiency in myobundles resulted in a ~2-fold decrease in specific force generation and Ca 2+ transient amplitude. In contrast, loss of dysferlin did not affect cardiobundle specific force generation, Ca 2+ transient amplitude, conduction velocity, or action potential duration. Following OSI, cardiobundles lost >60 % of peak force independent of phenotype, while LGMD2B myobundles exhibited significantly greater force loss than HLT controls. Conclusions: We present the first in vitro tissue-engineered model of human LGMD2B cardiac muscle and show that, unlike engineered skeletal muscle and similar to in vivo findings, dysferlin deficiency does not compromise engineered cardiac tissue structure or function. Ongoing transcriptional analysis of HLT and LGMD2B cardiobundles vs. myobundles will probe putative cardioprotective pathways, with subsequent loss- and gain-of-function studies planned to validate novel therapeutic targets transferable to skeletal muscle.