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Engraftment potential of dermal fibroblasts following in vivo myogenic conversion in immunocompetent dystrophic skeletal muscle.

ABSTRACT: Autologous dermal fibroblasts are promising candidates for enhancing muscle regeneration in Duchenne muscular dystrophy (DMD) due to their ease of isolation, immunological compatibility, and greater proliferative potential than DMD satellite cells. We previously showed that mouse fibroblasts, after MyoD-mediated myogenic reprogramming in vivo, engraft in skeletal muscle and supply dystrophin. Assessing the therapeutic utility of this system requires optimization of conversion and transplantation conditions and quantitation of engraftment so that these parameters can be correlated with possible functional improvements. Here we derived dermal fibroblasts from transgenic mice carrying mini-dystrophin, transduced them by lentivirus carrying tamoxifen-inducible MyoD, and characterized their myogenic and engraftment potential. After cell transplantation into muscles of immunocompetent dystrophic mdx^4cv mice, tamoxifen treatment drove myogenic conversion and fusion into myofibers that expressed high levels of mini-dystrophin. Injecting 50,000 cells/microliter (1 × 10⁶ total cells) resulted in a peak of ~600 mini-dystrophin positive myofibers in TA muscle single cross-sections. However, EDL muscles with up to 30% regional engraftment showed no functional improvements; similar limitations were obtained with whole muscle mononuclear cells. Despite the current lack of physiological improvement, this study suggests a viable initial strategy for using a patient-accessible dermal cell population to enhance skeletal muscle regeneration in DMD.

SUBMITTER: Muir LA

PROVIDER: S-EPMC4280788 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Chikungunya virus (CHIKV) is an arthritogenic alphavirus that acutely causes fever as well as severe joint and muscle pain. Chronic musculoskeletal pain persists in a substantial fraction of patients for months to years after the initial infection, yet we still have a poor understanding of what promotes chronic disease. While replicating virus has not been detected in joint-associated tissues of patients with persistent arthritis nor in various animal models at convalescent time points, viral RNA is detected months after acute infection. To identify the cells that might contribute to pathogenesis during this chronic phase, we developed a recombinant CHIKV that expresses Cre recombinase (CHIKV-3'-Cre). CHIKV-3'-Cre replicated in myoblasts and fibroblasts, and it induced arthritis during the acute phase in mice. Importantly, it also induced chronic disease, including persistent viral RNA and chronic myositis and synovitis similar to wild-type virus. CHIKV-3'-Cre infection of tdTomato reporter mice resulted in a population of tdTomato+ cells that persisted for at least 112 days. Immunofluorescence and flow cytometric profiling revealed that these tdTomato+ cells predominantly were myofibers and dermal and muscle fibroblasts. Treatment with an antibody against Mxra8, a recently defined host receptor for CHIKV, reduced the number of tdTomato+ cells in the chronic phase and diminished the levels of chronic viral RNA, implicating these tdTomato+ cells as the reservoir of chronic viral RNA. Finally, isolation and flow cytometry-based sorting of the tdTomato+ fibroblasts from the skin and ankle and analysis for viral RNA revealed that the tdTomato+ cells harbor most of the persistent CHIKV RNA at chronic time points. Therefore, this CHIKV-3'-Cre and tdTomato reporter mouse system identifies the cells that survive CHIKV infection in vivo and are enriched for persistent CHIKV RNA. This model represents a useful tool for studying CHIKV pathogenesis in the acute and chronic stages of disease.

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Engraftment potential of dermal fibroblasts following in vivo myogenic conversion in immunocompetent dystrophic skeletal muscle.

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