Publication Date:
2012-06-22
Description:
Background: Current methods of isolation of muscle satellite cells from different animal species are highlyvariable making inter-species comparisons problematic. This variation mainly stems from theuse of different proteolytic enzymes to release the satellite cells from the muscle tissue(sometimes a single enzyme is used but often a combination of enzymes is preferred) and thedifferent extracellular matrix proteins used to coat culture ware. In addition, isolation ofsatellite cells is frequently laborious and requires pre-plating of the cell preparation onuncoated flasks or Percoll centrifugation to remove contaminating fibroblasts. Themethodology employed to isolate and culture satellite cells in vitro can critically determinethe fusion of myoblasts into multi-nucleated myotubes. These terminally differentiatedmyotubes resemble mature myofibres in the muscle tissue in vivo, therefore optimal fusion isa keystone of in vitro muscle culture. Hence, a simple method of muscle satellite cellisolation and culture of different vertebrate species that can result in a high fusion rate ishighly desirable. Results: We demonstrate here a relatively simple and rapid method of isolating highly enrichedmuscle satellite cells from different avian and mammalian species without the need of Percollcentrifugation or pre-plating. In brief, muscle tissue was mechanically dissociated, digestedwith a single enzyme (pronase), triturated with a 10-ml pipette, filtered and directly platedonto collagen coated flasks. Following this method and after optimization of the cell cultureconditions, excellent fusion rates were achieved in the duck, chicken, horse and cow (withmore than 50% cell fusion), and to a lesser extent pig, pointing to pronase as a highly suitableenzyme to release satellite cells from muscle tissue. Conclusions: Our simplified method presents a quick and simple alternative to isolating highly enrichedmuscle satellite cell cultures which can subsequently rapidly differentiate into well developedprimary myotubes. The use of the same isolation protocol allows better inter-speciescomparisons of muscle satellite cells. Of all the farm animal species investigated, harvestedchicken muscle cells showed the highest percentage of muscle satellite cells, and equinemuscle cells presented the highest fusion index, an impressive [almost equal to] 77%. Porcine cells displayedthe lowest amount of satellite cells but still achieved a modest fusion rate of [almost equal to] 41%.
Electronic ISSN:
1471-2121
Topics:
Biology
,
Medicine
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