Cholesterol absorption blocker ezetimibe prevents muscle wasting in severe dysferlin-deficient and mdx mice.

BACKGROUND: Muscular dystrophy (MD) causes muscle wasting and is often lethal in patients due to a lack of proven therapies. In contrast, mouse models of MD are notoriously mild. We have previously shown severe human-like muscle pathology in mdx [Duchenne MD (DMD)] and dysferlin-deficient limb-girdle MD type 2B (LGMD2B) mice by inactivating the gene encoding for apolipoprotein E (ApoE), a lipid transporter synthesized by the liver, brain and adipocytes to regulate lipid and fat metabolism. Having recently established that human DMD is a novel type of primary genetic dyslipidaemia with elevated cholesterol, we sought to determine whether cholesterol could exacerbate the muscle wasting process observed in severe rodent MD. METHODS: Severe mdx and dysferlin knock-out mice lacking ApoE were treated with ezetimibe (15 mg/kg/day), a clinically approved drug exhibiting few pleiotropic effects. In separate studies, dietary cholesterol was raised (from 0.2% to 2% cholesterol) in combination with experimental micro-injury and direct cholesterol injection assays. Muscles were assessed histologically for changes in collagen and adipocyte infiltration and both transcriptomic and cellular changes by RNA-seq and fluorescence-activated cell sorting analysis. RESULTS: Treatment of severe DMD and LGMD2B mice with ezetimibe completely prevented clinical signs of ambulatory dysfunction (0% incidence vs. 33% for vehicle treatment; P < 0.05). Histological analyses revealed that ezetimibe-reduced fibro-fatty infiltration up to 84% and 63% in severely affected triceps (P ≤ 0.0001) and gastrocnemius (P ≤ 0.003) muscles, resulting in a respective 1.9-fold and 2.2-fold retention of healthy myofibre area (P ≤ 0.0001). Additionally, raising dietary cholesterol and thus concentrations of plasma low-density lipoprotein-associated cholesterol (by 250%; P < 0.0001) reduced overall survivability (by 100%; P < 0.001) and worsened muscle damage in the LGMD2B triceps by 767% (P < 0.03). Micro-pin-induced mechanical injury in LGMD2B mice fed a high cholesterol diet exacerbated muscle damage by 425% (P < 0.03) and increased macrophage recruitment (by 98%; P = 0.03) compared with those injured on a chow diet. Parallel RNA-seq analyses revealed that injury in cholesterol-fed mice also modulated the expression of 3671 transcripts (1953 up-regulated), with fibrogenic, inflammatory and programmed cell death-associated pathways among the most enriched. Mice lacking dysferlin also displayed heightened muscle necrosis (by 123%; P < 0.0001) following a direct intramuscular injection of cholesterol compared with control mice. CONCLUSIONS: Cholesterol exacerbates rodent MD. Specific inhibition of cholesterol absorption with ezetimibe may safely attenuate human MD severity and delay death.

Angiotensin II receptor blocker losartan exacerbates muscle damage and exhibits weak blood pressure-lowering activity in a dysferlin-null model of Limb-Girdle muscular dystrophy type 2B.

There is no cure or beneficial management option for Limb-Girdle muscular dystrophy (MD) type 2B (LGMD2B). Losartan, a blood pressure (BP) lowering angiotensin II (AngII) receptor type 1 (ATR1) blocker (ARB) with unique anti-transforming growth factor-ß (TGF-ß) properties, can protect muscles in various types of MD such as Duchenne MD, suggesting a potential benefit for LGMD2B patients. Herein, we show in a mild, dysferlin-null mouse model of LGMD2B that losartan increased quadriceps muscle fibrosis (142%; P<0.0001). In a severe, atherogenic diet-fed model of LGMD2B recently described by our group, losartan further exacerbated dysferlin-null mouse muscle wasting in quadriceps and triceps brachii, two muscles typically affected by LGMD2B, by 40% and 51%, respectively (P<0.05). Lower TGF-ß signalling was not observed with losartan, therefore plasma levels of atherogenic lipids known to aggravate LGMD2B severity were investigated. We report that losartan increased both plasma triglycerides and cholesterol concentrations in dysferlin-null mice. Other protective properties of losartan, such as increased nitric oxide release and BP lowering, were also reduced in the absence of dysferlin expression. Our data suggest that LGMD2B patients may show some resistance to the primary BP-lowering effects of losartan along with accelerated muscle wasting and dyslipidemia. Hence, we urge caution on the use of ARBs in this population as their ATR1 pathway may be dysfunctional.