TY - JOUR T1 - Bioengineered human skeletal muscle with a Pax7+ satellite cell niche capable of functional regeneration JF - bioRxiv DO - 10.1101/2020.05.04.076828 SP - 2020.05.04.076828 AU - J.W. Fleming AU - A.J. Capel AU - R.P. Rimington AU - P. Wheeler AU - O.G. Davies AU - M.P. Lewis Y1 - 2020/01/01 UR - http://biorxiv.org/content/early/2020/05/07/2020.05.04.076828.abstract N2 - Skeletal muscle (SkM) regenerates following injury, replacing damaged tissue with high fidelity. However, in serious injuries non-regenerative defects leave patients with loss of function, increased re-injury risk and often chronic pain. Progress in treating these non-regenerative defects has been slow, with advances only occurring where a comprehensive understanding of regeneration has been gained. Tissue engineering has allowed the development of bioengineered models of SkM which regenerate following injury to support research in regenerative physiology. To date however, no studies have utilised human myogenic precursor cells (hMPCs) to closely mimic human physiology due to difficulties generating sufficient cell numbers and the relatively low myogenic potential of hMPCs. Here we address problems associated with cell number and hMPC mitogenicity using magnetic association cell sorting (MACS), for the marker CD56, and media supplementation with fibroblast growth factor 2 (FGF-2) and B-27 supplement. Cell sorting allowed extended expansion of myogenic cells and supplementation was shown to improve myogenesis within engineered tissues and force generation at maturity. In addition, these engineered human SkM contained a Pax7+ niche and regenerated following Barium Chloride (BaCl2) injury. Following injury, reductions in function (87.5%) and myotube number (33.3%) were observed, followed by a proliferative phase with increased MyoD+ cells and a subsequent recovery of function and myotube number. An expansion of the Pax7+ cell population was observed across recovery suggesting an ability to generate Pax7+ cells within the tissue, similar to the self-renewal of satellite cells seen in vivo. This work outlines an engineered human SkM capable of functional regeneration following injury, built upon an open source system adding to the pre-clinical testing toolbox to improve the understanding of basic regenerative physiology.Competing Interest StatementThe authors have declared no competing interest.GMGrowth mediumDMDifferentiation mediumECMExtracellular matrixMPCMyogenic precursor cellhMPCHuman myogenic precursor cellMyHCMyosin heavy chainSCSatellite cellVMLVolumetric muscle lossFAPFibro/adipogenic cell3DThree dimensionalPSCPluripotent stem cellDMEMDulbecco’s Modified Eagle’s MediumFBSFetal Bovine SerumP/SPenicillin/StreptomycinMEMMinimal essential mediumPBSPhosphate buffered salineKRHKrebs-Ringer-HEPESBaCl2Barium Chloride ER -