Glucocorticoid steroids are commonly prescribed for many inflammatory conditions, but chronic daily use produces adverse effects including muscle wasting and weakness. In contrast, shorter glucocorticoid pulses may improve athletic performance, although the mechanisms remain unclear. Muscle is sexually dimorphic and comparatively little is known about how male and female muscles respond to glucocorticoid steroids. We investigated the impact of onceweekly glucocorticoid exposure on skeletal muscle performance comparing male and female mice. One month of once-weekly glucocorticoid dosing improved muscle specific force in both males and females. Transcriptomic profiling of isolated myofibers identified a striking sexually dimorphic response to weekly glucocorticoids. Male myofibers had increased expression of genes in the IGF1/PI3K pathway and calcium handling, while female myofibers had profound upregulation of lipid metabolism genes. Muscles from weekly prednisone-treated males had improved calcium handling, while comparably treated female muscles had reduced intramuscular triglycerides. Consistent with altered lipid metabolism, weekly prednisone-treated female mice had greater endurance relative to controls. Using chromatin immunoprecipitation, we defined a sexually dimorphic chromatin landscape after weekly prednisone. These results demonstrate that weekly glucocorticoid exposure elicits distinct pathways in males versus females resulting in enhanced performance.

Muscular dystrophies are disorders characterized by progressive muscle loss and weakness that are both genotypically and phenotypically heterogenous. Progression of muscle disease arises from impaired regeneration, plasma membrane instability, defective membrane repair, and calcium mishandling. The ferlin protein family, including dysferlin and myoferlin, are calcium-binding, membrane-associated proteins that regulate membrane fusion, trafficking, and tubule formation. Mice lacking dysferlin (Dysf), myoferlin (Myof), and both dysferlin and myoferlin (Fer) on an isogenic inbred 129 background were previously demonstrated that loss of both dysferlin and myoferlin resulted in more severe muscle disease than loss of either gene alone. Furthermore, Fer mice had disordered triad organization with visibly malformed transverse tubules and sarcoplasmic reticulum, suggesting distinct roles of dysferlin and myoferlin. To assess the physiological role of disorganized triads, we now assessed excitation contraction (EC) coupling in these models. We identified differential abnormalities in EC coupling and ryanodine receptor disruption in flexor digitorum brevis myofibers isolated from ferlin mutant mice. We found that loss of dysferlin alone preserved sensitivity for EC coupling and was associated with larger ryanodine receptor clusters compared to wildtype myofibers. Loss of myoferlin alone or together with a loss of dysferlin reduced sensitivity for EC coupling, and produced disorganized and smaller ryanodine receptor cluster size compared to wildtype myofibers. These data reveal impaired EC coupling in Myof and Fer myofibers and slightly potentiated EC coupling in Dysf myofibers. Despite high homology, dysferlin and myoferlin have differential roles in regulating sarcotubular formation and maintenance resulting in unique impairments in calcium handling properties.

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A premature truncation variant in MYBPHL in humans and a loss of Mybphl in mice associates with dilated cardiomyopathy (DCM), atrial and ventricular arrhythmias, and atrial enlargement. MYBPHL encodes myosin binding protein H-like (MyBP-HL) and is expressed highly in the atria and in foci throughout the ventricle. We hypothesize that MyBP-HL is required for proper formation and function of the conduction system. Surface telemetry found atrioventricular block and atrial bigeminy, while intracardiac pacing revealed a faster atrial relative refractory period and atrial tachycardia in Mybphl -null mice. Ca 2+ transient analysis revealed that isolated Mybphl -null atrial cardiomyocytes had an increased occurrence of triggered Ca 2+ waves and more heterogenous Ca 2+ release than wild-type (WT) controls. Super-resolution microscopy revealed ryanodine receptor disorganization in Mybphl het and null atrial cardiomyocytes compared to WT controls. Immunofluorescence microscopy in WT adult mouse ventricles identified MyBP-HL-positive ventricular cardiomyocytes that co-localized with the ventricular conduction system marker contactin-2 near the atrioventricular node and in a subset of Purkinje fibers. Mybphl heterozygous ventricles showed 10% as many MyBP-HL-positive cells compared to WT. Lightsheet microscopy of perinatal day 5 hearts showed enrichment of MyBP-HL-positive cells within and immediately adjacent to the Cntn2-positive ventricular conduction system in WT hearts, but this association was not apparent in Mybphl heterozygous null hearts. These data, abnormal Ca 2+ release, shorter atrial refractory period, and atrial dilation could account for the observed atrial arrhythmias, bigeminy, and atrial tachycardia, whereas the proximity of MyBP-HL-positive cells with the ventricular conduction system provides insight into how a predominantly atrial expressed gene could cause ventricular arrhythmias.

A premature truncation (R255X) in MYBPHL associates with human dilated cardiomyopathy (DCM) and arrhythmias. Loss of Mybphl in mice causes DCM and arrhythmia. MYBPHL encodes myosin binding protein H-like (MyBP-HL) and is expressed highly in the atria. We hypothesize that MyBP-HL is required for proper conduction system function. Immunofluorescence microscopy on normal human atria showed MyBP-HL staining in all atrial cardiomyocytes with a sarcomere A-band pattern. Atria from the heterozygous (het) MYBPHL R255X mutant carrier lacked MyBP-HL staining. Human induced pluripotent stem cell-derived cardiomyocytes from the het MYBPHL R255X carrier and control cell lines were also examined. MyBP-HL was found in a subset of control cardiomyocytes, whereas R255X cells showed no MyBP-HL, suggesting that the R255X allele exerts a dominant-negative effect on the normal MYBPHL allele. Immunofluorescence microscopy in wild-type (WT) mouse ventricles identified MyBP-HL-positive ventricular cardiomyocytes that co-localized with the ventricular conduction system marker contactin-2 near the atrioventricular node and in a subset of Purkinje fibers. Mybphl het ventricles have 10% as many MyBP-HL-positive cells compared to WT. Surface telemetry revealed atrioventricular block and atrial bigeminy and intracardiac pacing revealed a shorter atrial relative refractory period and inducible atrial tachycardia in Mybphl -null mice. Ca 2+ transients measured with confocal microscopy revealed that isolated Mybphl -null atrial cardiomyocytes had an increased occurrence of triggered Ca 2+ waves and more heterogenous Ca 2+ release than WT controls. Super-resolution microscopy revealed ryanodine receptor disorganization in Mybphl het and null atrial cardiomyocytes compared to WT controls. Abnormal Ca 2+ release, shorter atrial refractory period, and dilated atria could account for the observed atrial arrhythmias, bigeminy, and atrial tachycardia.