No abstract

Myoblast proliferation and differentiation are essential for normal skeletal muscle growth and repair. Muscle recovery is dependent on the quiescent population of muscle stem cells - satellite cells. During muscle injury, satellite cells become mitotically active and begin the repair process by fusing with each other and/or with myofibers. Aging, prolonged inactivity, obesity, cachexia and other muscle wasting diseases are associated with a decreased number of quiescent and proliferating satellite cells, which impedes the repair process.A high-content/high-throughput platform was developed and utilized for robust phenotypic evaluation of human primary satellite cells in vitro for the discovery of chemical probes that may improve muscle recovery. A 1600 compound pilot screen was developed using two highly annotated small molecule libraries. This screen yielded 15 dose responsive compounds that increased proliferation rate in satellite cells derived from a single obese human donor. Two of these compounds remained dose responsive when counter-screened in 3-donor obese superlot. The Alk-5 inhibitor LY364947, was used as a positive control for assessing satellite cell proliferation/delayed differentiation. A multivariate approach was utilized for exploratory data analysis to discover proliferation vs. differentiation-dependent changes in cellular phenotype. Initial screening efforts successfully identified a number of phenotypic outcomes that are associated with desired effect of stimulation of proliferation and delayed differentiation.

Phenotypical differences between muscle fibers are associated with a source of cellular energy. Coenzyme Q₁₀ (CoQ₁₀) is a major component of the mitochondrial oxidative phosphorylation process, and it significantly contributes to the production of cellular energy in the form of ATP. The objective of this study was to determine the relationship between whole-tissue CoQ₁₀ content, mitochondrial CoQ₁₀ content, mitochondrial protein, and muscle phenotype in turkeys. Four specialized muscles (anterior latissimus dorsi, ALD; posterior latissimus dorsi, PLD; pectoralis major, PM, and biceps femoris, BF) were evaluated in 9- and 20-week-old turkey toms. The amount of muscle mitochondrial protein was determined using the Bradford assay and CoQ₁₀ content was measured using HPLC-UV. The amount of mitochondrial protein relative to total protein was significantly lower (p < 0.05) at 9 compared to 20 weeks of age. All ALD fibers stained positive for anti-slow (S35) MyHC antibody. The PLD and PM muscle fibers revealed no staining for slow myosin heavy chain (S35 MyHC), whereas half of BF muscle fibers exhibited staining for S35 MyHC at 9 weeks and 70% at 20 weeks of age. The succinate dehydrogenase (SDH) staining data revealed that SDH significantly increases (p < 0.05) in ALD and BF muscles and significantly decreases (p < 0.05) in PLD and PM muscles with age. The study reveals age-related decreases in mitochondrial CoQ₁₀ content in muscles with fast/glycolytic profile, and demonstrates that muscles with a slow/oxidative phenotypic profile contain a higher proportion of CoQ₁₀ than muscles with a fast/glycolytic phenotypic profile.

No abstract

Obesity and insulin resistance are primary risk factors for Non-Alcoholic Fatty Liver Disease (NAFLD). NAFLD is generally exhibited by non-progressive simple steatosis. However, a significant subset of patient’s progress to nonalcoholic steatohepatitis (NASH) that is defined by the presence of steatosis, inflammation and hepatocyte injury with fibrosis. Unfortunately, there are no approved therapies for NAFLD or NASH and therefore therapeutic approaches are urgently needed. Niclosamide is an U.S. Food and Drug Administration (FDA)-approved anthelmintic drug that mediates its effect by uncoupling oxidative phosphorylation. Niclosamide and its salt forms, Niclosamide Ethanolamine (NEN), and Niclosamide Piperazine (NPP) have shown efficacy in murine models of diet induced obesity characterized by attenuation of the prominent fatty liver disease phenotype and improved glucose metabolism. While the exact mechanism(s) underlying these changes remains unclear, the ability to uncouple oxidative phosphorylation leading to increased energy expenditure and lipid metabolism or attenuation of PKA mediated glucagon signaling in the liver have been proposed. Unfortunately, niclosamide has very poor water solubility, leading to low oral bioavailability. This, in addition to mitochondrial uncoupling activity and potential genotoxicity have reduced enthusiasm for its clinical use. More recently, salt forms of niclosamide, NEN and NPP, have demonstrated improved oral bioavailability while retaining activity. This suggests that development of safer more effective niclosamide derivatives for the treatment of NAFLD and Type 2 Diabetes may be possible. Herein we explored the ability of a series of N-substituted phenylbenzamide derivatives of the niclosamide salicylanilide chemotype to attenuate hepatic steatosis using a novel phenotypic in vitro model of fatty liver and the high fat diet-fed mouse model of diet induced obesity. These studies identified novel compounds with improved pre-clinical properties that attenuate hepatic steatosis in vitro and in vivo. These compounds with improved drug properties may be useful in alleviating symptoms and protection against disease progression in patients with metabolic syndrome and NAFLD.