Six weeks of N-acetylcysteine antioxidant in drinking water decreases pathological fiber branching in MDX mouse dystrophic fast-twitch skeletal muscle.

Introduction: It has been proposed that an increased susceptivity to oxidative stress caused by the absence of the protein dystrophin from the inner surface of the sarcolemma is a trigger of skeletal muscle necrosis in the destructive dystrophin deficient muscular dystrophies. Here we use the mdx mouse model of human Duchenne Muscular Dystrophy to test the hypothesis that adding the antioxidant NAC at 2% to drinking water for six weeks will treat the inflammatory phase of the dystrophic process and reduce pathological muscle fiber branching and splitting resulting in a reduction of mass in mdx fast-twitch EDL muscles. Methods: Animal weight and water intake was recorded during the six weeks when 2% NAC was added to the drinking water. Post NAC treatment animals were euthanised and the EDL muscles dissected out and placed in an organ bath where the muscle was attached to a force transducer to measure contractile properties and susceptibility to force loss from eccentric contractions. After the contractile measurements had been made the EDL muscle was blotted and weighed. In order to assess the degree of pathological fiber branching mdx EDL muscles were treated with collagenase to release single fibers. For counting and morphological analysis single EDL mdx skeletal muscle fibers were viewed under high magnification on an inverted microscope. Results: During the six-week treatment phase NAC reduced body weight gain in three- to nine-week-old mdx and littermate control mice without effecting fluid intake. NAC treatment also significantly reduced the mdx EDL muscle mass and abnormal fiber branching and splitting. Discussion: We propose chronic NAC treatment reduces the inflammatory response and degenerative cycles in the mdx dystrophic EDL muscles resulting in a reduction in the number of complexed branched fibers reported to be responsible for the dystrophic EDL muscle hypertrophy.

In Vitro and In Vivo Anti-Clostridioides difficile Effect of a Probiotic Bacillus amyloliquefaciens Strain.

Clostridioides difficile infection (CDI) is a significant cause of hospital-acquired and antibiotic-mediated intestinal diseases and is a growing global public health concern. Overuse of antibiotics and their effect on normal intestinal flora has increased the incidence and severity of infections. Thus, the development of new, effective, and safe treatment options is a high priority. Here, we report a new probiotic strain, Bacillus amyloliquefaciens (BA PMC-80), and its in vitro/in vivo anti-C. difficile effect as a prospective novel candidate for replacing conventional antibiotics. BA PMC-80 showed a significant anti-C. difficile effect in coculture assay, and its cell-free supernatant (CFS) also exhibited a considerable anti-C. difficile effect with an 89.06 µg/ml 50% minimal inhibitory concentration (MIC) in broth microdilution assay. The CFS was stable and equally functional under different pHs, heat, and proteinase treatments. It also exhibited a high sensitivity against current antibiotics and no toxicity in subchronic toxicity testing in hamsters. Finally, BA PMC-80 showed a moderate effect in a hamster CDI model with reduced infection severity and delayed death. However, further studies are required to optimize the treatment condition of the hamster CDI model for better efficacy and identify the antimicrobial compound produced by BA PMC-80.

Minocycline Treatment Reduces Mass and Force Output From Fast-Twitch Mouse Muscles and Inhibits Myosin Production in C2C12 Myotubes.

Minocycline, a tetracycline-class of antibiotic, has been tested with mixed effectiveness on neuromuscular disorders such as amyotrophic lateral sclerosis, autoimmune neuritis and muscular dystrophy. The independent effect of minocycline on skeletal muscle force production and signalling remain poorly understood. Our aim here is to investigate the effects of minocycline on muscle mass, force production, myosin heavy chain abundance and protein synthesis. Mice were injected with minocycline (40 mg/kg i.p.) daily for 5 days and sacrificed at day six. Fast-twitch EDL, TA muscles and slow-twitch soleus muscles were dissected out, the TA muscle was snap-frozen and the remaining muscles were attached to force transducer whilst maintained in an organ bath. In C2C12 myotubes, minocycline was applied to the media at a final concentration of 10 µg/mL for 48 h. In minocycline treated mice absolute maximal force was lower in fast-twitch EDL while in slow-twitch soleus there was an increase in the time to peak and relaxation of the twitch. There was no effect of minocycline treatment on the other contractile parameters measured in isolated fast- and slow-twitch muscles. In C2C12 cultured cells, minocycline treatment significantly reduced both myosin heavy chain content and protein synthesis without visible changes to myotube morphology. In the TA muscle there was no significant changes in myosin heavy chain content. These results indicate that high dose minocycline treatment can cause a reduction in maximal isometric force production and mass in fast-twitch EDL and impair protein synthesis during myogenesis in C2C12 cultured cells. These findings have important implications for future studies investigating the efficacy of minocycline treatment in neuromuscular or other muscle-atrophy inducing conditions.