Platelet releasate normalises the compromised muscle regeneration in a mouse model of hyperlipidaemia.

NEW FINDINGS: What is the central question of this study? What is the impact of obesity-independent hyperlipidaemia on skeletal muscle stem cell function of ApoE-deficient (ApoE-/- ) mice? What is the main finding and its importance? Compromised muscle stem cell function accounts for the impaired muscle regeneration in hyperlipidaemic ApoE-/- mice. Importantly, impaired muscle regeneration is normalised by administration of platelet releasate. ABSTRACT: Muscle satellite cells are important stem cells for skeletal muscle regeneration and repair after injury. ApoE-deficient mice, an established mouse model of hyperlipidaemia and atherosclerosis, show evidence of oxidative stress-induced lesions and fat infiltration in skeletal muscle followed by impaired repair after injury. However, the mechanisms underpinning attenuated muscle regeneration remain to be fully defined. Key to addressing the latter is to understand the properties of muscle stem cells from ApoE-deficient mice and their myogenic potential. Muscle stem cells from ApoE-deficient mice were cultured both ex vivo (on single fibres) and in vitro (primary myoblasts) and their myogenic capacity was determined. Skeletal muscle regeneration was studied on days 5 and 10 after cardiotoxin injury. ApoE-deficient muscle stem cells showed delayed activation and differentiation on single muscle fibres ex vivo. Impaired proliferation and differentiation profiles were also evident on isolated primary muscle stem cells in culture. ApoE-deficient mice displayed impaired skeletal muscle regeneration after acute injury in vivo. Administration of platelet releasate in ApoE-deficient mice reversed the deficits of muscle regeneration after acute injury to wild-type levels. These findings indicate that muscle stem cell myogenic potential is perturbed in skeletal muscle of a mouse model of hyperlipidaemia. We propose that platelet releasate could be a therapeutic intervention for conditions with associated myopathy such as peripheral arterial disease.

The inhibitory subunit of cardiac troponin (cTnI) is modified by arginine methylation in the human heart.

BACKGROUND: The inhibitory subunit of cardiac troponin (cTnI) is a gold standard cardiac biomarker and also an essential protein in cardiomyocyte excitation-contraction coupling. The interactions of cTnI with other proteins are fine-tuned by post-translational modification of cTnI. Mutations in cTnI can lead to hypertrophic cardiomyopathy. METHODS AND RESULTS: Here we report, for the first time, that cTnI is modified by arginine methylation in human myocardium. Using Western blot, we observed reduced levels of cTnI arginine methylation in human hypertrophic cardiomyopathy compared to dilated cardiomyopathy biopsies. Similarly, using a rat model of cardiac hypertrophy we observed reduced levels of cTnI arginine methylation compared to sham controls. Using mass spectrometry, we identified cTnI methylation sites at R74/R79 and R146/R148 in human cardiac samples. R146 and R148 lie at the boundary between the critical cTnI inhibitory and switch peptides; PRMT1 methylated an extended inhibitory peptide at R146 and R148 in vitro. Mutations at R145 that have been associated with hypertrophic cardiomyopathy hampered R146/R148 methylation by PRMT1 in vitro. H9c2 cardiac-like cells transfected with plasmids encoding for a methylation-deficient R146A/R148A cTnI protein developed cell hypertrophy, with a 32% increase in cell size after 72 h, compared to control cells. DISCUSSION: Our results provide evidence for a novel and significant cTnI post-translational modification. Our work opens the door to translational investigations of cTnI arginine methylation as a biomarker of disease, which can include e.g. cardiomyopathies, myocardial infarction and heart failure, and offers a novel way to investigate the effect of cTnI mutations in the inhibitory/switch peptides.

Attenuation of oxidative stress-induced lesions in skeletal muscle in a mouse model of obesity-independent hyperlipidaemia and atherosclerosis through the inhibition of Nox2 activity.

Obesity leading to hyperlipidaemia and atherosclerosis is recognised to induce morphological and metabolic changes in many tissues. However, hyperlipidaemia can occur in the absence of obesity. The impact of the latter scenario on skeletal muscle and liver is not understood sufficiently. In this regard, we used the Apolipoprotein E-deficient (ApoE-/-) mouse model, an established model of hyperlipidaemia and atherosclerosis, that does not become obese when subjected to a high-fat diet, to determine the impact of Western-type diet (WD) and ApoE deficiency on skeletal muscle morphological, metabolic and biochemical properties. To establish the potential of therapeutic targets, we further examined the impact of Nox2 pharmacological inhibition on skeletal muscle redox biology. We found ectopic lipid accumulation in skeletal muscle and the liver, and altered skeletal muscle morphology and intramuscular triacylglycerol fatty acid composition. WD and ApoE deficiency had a detrimental impact in muscle metabolome, followed by perturbed gene expression for fatty acid uptake and oxidation. Importantly, there was enhanced oxidative stress in the skeletal muscle and development of liver steatosis, inflammation and oxidative protein modifications. Pharmacological inhibition of Nox2 decreased reactive oxygen species production and protein oxidative modifications in the muscle of ApoE-/- mice subjected to a Western-type diet. This study provides key evidence to better understand the pathophysiology of skeletal muscle in the context of hyperlipidaemia and atherosclerosis and identifies Nox2 as a potential target for attenuating oxidative stress in skeletal muscle in a mouse model of obesity-independent hyperlipidaemia.