Unloading, neural lesions, and hormonal disorders after acute motor-complete spinal cord injury (SCI) cause one of the most severe forms of bone loss, a condition that has been refractory to available interventions tested to date. Thus, these features related to acute SCI provide a unique opportunity to study complex bone problems, potential efficacious interventions, and mechanisms of action that are associated with these dramatic pathological changes. This study was designed to explore the therapeutic potential of sclerostin antibody (Scl-Ab) in a rat model of bone loss after motor-complete SCI, and to investigate mechanisms underlying bone loss and Scl-Ab action. SCI rats were administered Scl-Ab (25 mg/kg/week) or vehicle beginning 7 days after injury then weekly for 7 weeks. SCI resulted in significant decreases in bone mineral density (-25%) and trabecular bone volume (-67%) at the distal femur; Scl-Ab completely prevented these deteriorations of bone in SCI rats, concurrent with markedly increased bone formation. Scanning electron microscopy revealed that SCI reduced numbers of osteocytes and dendrites concomitant with a morphology change from a spindle to round shape; Scl-Ab corrected these abnormalities in osteocytes. In ex vivo cultures of bone marrow cells, Scl-Ab inhibited osteoclastogenesis, and promoted osteoblastogenesis accompanied by increases in mRNA levels of LRP5, osteoprotegerin (OPG), and the OPG/RANKL ratio, and a decrease in DKK1 mRNA. Our findings provide the first evidence that robust bone loss after acute motor-complete SCI can be blocked by Scl-Ab, at least in part, through the preservation of osteocyte morphology and structure and related bone remodeling. Our findings support the inhibition of sclerostin as a promising approach to mitigate the striking bone loss that ensues after acute motor-complete SCI, and perhaps other conditions associated with disuse osteoporosis as a consequence of neurological disorders.

Exosomes are vesicles released by many eukaryotic cells; their cargo includes proteins, mRNA and microRNA (miR) that can be transferred to recipient cells and regulate cellular processes in an autocrine or paracrine manner. While cells of the myoblast lineage secrete exosomes, it is not known whether skeletal muscle fibers (myofibers) release exosomes. In this study, we found that cultured myofibers release nanovesicles that have bilamellar membranes and an average size of 60–130 nm, contain typical exosomal proteins and miRNAs and are taken up by C2C12 cells. miR-133a was found to be the most abundant myomiR in these vesicles while miR-720 was most enriched in exosomes compared to parent myofibers. Treatment of NIH 3T3 cells with myofiber-derived exosomes downregulated the miR-133a targets proteins Smarcd1 and Runx2, confirming that these exosomes have biologically relevant effects on recipient cells. Denervation resulted in a marked increase in miR-206 and reduced expression of miRs 1, 133a, and 133b in myofiber-derived exosomes. These findings demonstrate that skeletal muscle fibers release exosomes which can exert biologically significant effects on recipient cells, and that pathological muscle conditions such as denervation induce alterations in exosomal miR profile which could influence responses to disease states through autocrine or paracrine mechanisms.

Background The role of Numb, a protein that is important for cell fate and development and that, in human muscle, is expressed at reduced levels with advanced age, was investigated; adult mice skeletal muscle and its localization and function within myofibres were determined. Methods Numb expression was evaluated by western blot. Numb localization was determined by confocal microscopy. The effects of conditional knock out (cKO) of Numb and the closely related gene Numb‐like in skeletal muscle fibres were evaluated by in situ physiology, transmission and focused ion beam scanning electron microscopy, three‐dimensional reconstruction of mitochondria, lipidomics, and bulk RNA sequencing. Additional studies using primary mouse myotubes investigated the effects of Numb knockdown on cell fusion, mitochondrial function, and calcium transients. Results Numb protein expression was reduced by ~70% (P < 0.01) at 24 as compared with 3 months of age in gastrocnemius and tibialis anterior muscle. Numb was localized within muscle fibres as bands traversing fibres at regularly spaced intervals in close proximity to dihydropyridine receptors. The cKO of Numb and Numb‐like reduced specific tetanic force by 36% (P < 0.01), altered mitochondrial spatial relationships to sarcomeric structures, increased Z‐line spacing by 30% (P < 0.0001), perturbed sarcoplasmic reticulum organization and reduced mitochondrial volume by over 80% (P < 0.01). Only six genes were differentially expressed in cKO mice: Itga4, Sema7a, Irgm2, Vezf1, Mib1, and Tmem132a. Several lipid mediators derived from polyunsaturated fatty acids through lipoxygenases were up‐regulated in Numb cKO skeletal muscle: 12‐HEPE was increased by ~250% (P < 0.05) and 17,18‐EpETE by ~240% (P < 0.05). In mouse primary myotubes, Numb knockdown reduced cell fusion (~20%, P < 0.01) and delayed the caffeine‐induced rise in cytosolic calcium concentrations by more than 100% (P < 0.01). Conclusions These findings implicate Numb as a critical factor in skeletal muscle structure and function and suggest that Numb is critical for calcium release. We therefore speculate that Numb plays critical roles in excitation–contraction coupling, one of the putative targets of aged skeletal muscles. These findings provide new insights into the molecular underpinnings of the loss of muscle function observed with sarcopenia.

Our findings demonstrate that SOST KO mice were protected from the major sublesional bone loss that invariably follows SCI. The evidence indicates that sclerostin is an important mediator of the marked sublesional bone loss after SCI, and that pharmacological inhibition of sclerostin may represent a promising novel approach to this challenging clinical problem.

In [1], the author reference erroneously listed, "Christopher C Cardozo." The author reference is now corrected to include "Christopher P Cardozo."This erratum corrects the error.