RANKL inhibition in the amelioration of sarcopenia: potential cross-tissue effects of musculoskeletal therapies

Osteoprotegerin (OPG) and denosumab, a human monoclonal antibody, bind and inhibit RANKL, a major regulator of osteoclastic bone resorption. As well as reducing fracture risk, use of denosumab has also significantly reduced fall incidence in placebo-controlled trials, suggesting RANKL inhibition may influence skeletal muscle function. In mouse models OPG has induced beneficial effects on muscle function. However, it is not understood how OPG or denosumab regulate muscle homeostasis, therefore investigations were made into skeletal muscle RANKL inhibition to determine its potential to help mitigate age-related skeletal muscle loss, also known as sarcopenia. Primary mouse and immortalised human myoblasts and myotubes were treated with human recombinant (h)OPG or denosumab at different stages of in vitro myogenesis to explore their effects on proliferation, myogenic capacity, NFκB signalling dynamics and atrophy/hypertrophy pathways. In an in vitro primary mouse myogenesis model, hOPG increased proliferation markers, improved myogenic phenotype by increasing myotube area, diameter, and fusion index, downregulated NFκB signalling and its downstream targets (e.g., MuRF1) and regulated hypertrophy/atrophy pathways to positively favour maintenance or growth of myotubes. In immortalised human muscle cells (KM670), expression of proteins of the OPG-RANKL-RANK axis increased as in vitro myogenesis progressed. Thus, the effects of hOPG and denosumab were more noticeable at later stages of myogenesis when RANK localised on myotubes. Both hOPG and denosumab improved human myotube phenotype, specifically, increasing myotube fusion index. hOPG may also regulate KM670 proliferation. The pro-myogenic effect of hOPG on in vitro mouse myogenesis may be caused by NFκB-mediated downregulation of atrogenes, accompanied with a synchronistic activation of muscle growth pathways. Initial investigations suggest that denosumab induces a pro-myogenic effect in vitro, a finding that warrants further exploration of underlying mechanisms and magnitude of benefit. Elucidating OPG-RANKL-RANK dynamics in skeletal muscle could help understand muscle physiology and/or help develop treatments to ameliorate sarcopenia.