Evidence of heterogeneity within bovine satellite cells isolated from young and adult animals

Abstract

Satellite cells are a heterogeneous population of myogenic precursors responsible for muscle growth and repair in mammals. The objectives of the experiment were to examine the growth rates and degree of heterogeneity within bovine satellite cells (BSC) isolated from young and adult animals. The BSC were harvested from the semimembranosus of young (4.3 ± 0.5 d) and adult (estimated 24 to 27 mo) cattle and cultured en masse. Young animal BSC re-enter the cell cycle sooner and reach maximal 5-ethynyl-2'-deoxyuridine (EdU) incorporation earlier (P < 0.05) than adult contemporaries. Adult BSC contain fewer (P < 0.05) MyoD and myogenin immunopositive nuclei than BSC isolated from young animals after 3, 4, and 5 d in culture. These results indicate that BSC from young animals activate, proliferate, and differentiate sooner than isolates from adult animals. Lineage heterogeneity within BSC was examined using antibodies specific for Pax7 and Myf5, lineage markers of satellite cells, and myoblasts. Immunocytochemistry revealed the majority of Pax7-expressing BSC also express Myf5; a minor population (~5%) fails to exhibit Myf5 immunoreactivity. The percentage of Pax7:Myf5 BSC from young animals decreases sooner (P < 0.05) in culture than adult BSC, indicating a more rapid rate of muscle fiber formation. A subpopulation immunopositive for Myf5 only was identified in both ages of BSC isolates. The growth kinetics and heterogeneity of young BSC was further evaluated by clonal analysis. Single cell clones were established and analyzed after 10 d. Colonies segregated into 2 groups based upon population doubling time. Immunostaining of the slow-growing colonies (population doubling time ≥ 3 d) revealed that a portion exhibited asymmetric distribution of the lineage markers Pax7 and Myf5, similar to self-renewable mouse muscle stem cells. In summary, these results offer insight into the heterogeneity of BSC and provide evidence for subtle differences between rodent and bovine myogenic precursors.

Temporal correlation between differentiation factor expression and microRNAs in Holstein bovine skeletal muscle

“…The proliferative ability and temporal expression of mRNA by satellite cells seems to be dependent on the host and the muscle from which they are collected (Powell et al, 2014). Additional levels of heterogeneity are known to exist between satellite cells populations, as well as different extracellular matrix protein compositions, growth kinetics, myoblast fusion capacities, transcription factor profiles, motility kinetics and migratory properties (Biressi and Rando, 2010;Boldrin et al, 2010;Li et al, 2011;Li and Johnson, 2013). There are increasing evidence through gene expression profiling and cell-surface marker analysis that satellite cells are a heterogeneous population (Motohashi and Asakura, 2014).…”

In vitro supplementation with the porcine plasma product, betaGRO®, stimulates activity of porcine fetal myoblasts and neonatal satellite cells in a divergent manner

“…Satellite cell activity is modulated through the transition of transcriptional signals, that begins with elevated Pax7 expression levels in quiescent satellite cells and progresses through the expression of the myogenic regulator factors as the satellite cells begin to proliferate and commit to terminal differentiation (Zammit et al, 2006;Biressi and Rando, 2010). Li et al (2011) categorized in vitro proliferative satellite cells into three sub-classes of cells that express Pax7, Myf-5, and co-express Pax7 and Myf-5. Cells that solely express Pax7 are activated and are less proliferative than cells that co-express Pax7 and Myf-5, which are the most proliferative of the three populations of myogenic cells.…”

“…For detection of cells traversing S-phase, BrdU, a thymidine analog, was added to media 2 h before immunostaining (Allen et al, 1979). Temporal characterization of the muscle lineage markers was conducted as previously described (Li et al, 2011). Briefly, cells were fixed with 4% (wt/vol) formaldehyde (Polysciences, Warrington, PA, USA) in PBS.…”

“…Within 24 h of birth, whole left LM were collected from male piglets nearest to the average weight of the litter for PSC isolation (n = 3). The methods of Li et al (2011) were be followed for myoblast isolation with slight modifications. Briefly, muscles were excised of all visible connective tissue and minced with sterilized surgical scissors.…”

“…The transition from paired box transcription factor-7 (Pax7) to myogenic factor-5 (Myf-5) characterizes the developmental maturity of proliferative myogenic cells (Zammit et al, 2006). In addition, there are a subclass of daughter cells that co-express Pax7 and Myf-5, which exhibit advanced proliferative characteristics (Kuang et al, 2007;Li et al, 2011). This transcriptional transition is influenced by a variety of factors, including endocrine signaling factors.…”