Abstract
BackgroundMaintaining stem cells in physiologically relevant states is necessary to understand cell and context-specific signalling paradigms and to understand complex interfaces between cells in situ. Understanding human stem cell function is largely based on tissue biopsies, cell culture, and transplantation into model organisms.MethodsHere, we describe a method to isolate post-mortem intact human muscle myofibers and culture muscle stem cells within the niche microenvironment to assay cellular dynamics, stem cell identity, stem cell hierarchy, and differentiation potential.ResultsWe show human myofiber culture maintains complex cell-cell contacts and extracellular niche composition during culture. Human satellite cells can be cultured at least 8 days, which represents a timepoint of activation, differentiation, and de novo human myofiber formation. We demonstrate that adult human muscle stem cells undergo apicobasal and planar cell divisions and express polarized dystrophin and EGFR. Furthermore, we validate that stimulation of the EGFR pathway stimulates the generation of myogenic progenitors and myogenic differentiation.ConclusionsThis method provides proof of principle evidence for the use of human muscle to evaluate satellite cell dynamics and has applications in pre-clinical evaluation of therapeutics targeting muscle repair.
Highlights
Maintaining stem cells in physiologically relevant states is necessary to understand cell and contextspecific signalling paradigms and to understand complex interfaces between cells in situ
Growth, and repair are facilitated by muscle resident stem cells which reside within skeletal muscle, resting within a specialized cleft underneath the basal lamina [1]
Human Psoas muscle is amenable for satellite cell culture in situ To evaluate our hypothesis that human myofibers could be cultured in a laboratory setting, we decided to isolate primary human tissue from neurological determination of death organ donors following ethics approval and informed consent
Summary
Maintaining stem cells in physiologically relevant states is necessary to understand cell and contextspecific signalling paradigms and to understand complex interfaces between cells in situ. Chemical and physical cues present in young and healthy muscle act to balance satellite cell quiescence, selfrenewal, and asymmetric division to maintain the satellite cell pool in a state amenable to rapid activation in response to injury [9, 10, 13]. Understanding if these processes are conserved or altered in human satellite cells is important to develop methods improving endogenous satellite cellmediated repair
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