Abstract
Mechanism controlling myo-adipogenic balance in skeletal muscle is of great significance for human skeletal muscle dysfunction and myopathies as well as livestock meat quality. In the present study, two cell subpopulations with particular potency of adipogenic or myogenic differentiation were isolated from neonatal porcine longissimus dorsi using the preplate method to detect mechanisms underlying distinct fate commitment of myogenic and adipogenic cells in skeletal muscle. Both cells share a common surface expression profile of CD29+CD31−CD34−CD90+CD105+, verifying their mesenchymal origin. A total of 448 differentially expressed genes (DEGs) (FDR < 0.05 and |log2 FC| ≥ 1) between two distinct cells were identified via RNA-seq, including 358 up-regulated and 90 down-regulated genes in myogenic cells compared with adipogenic cells. The results of functional annotation and enrichment showed that 42 DEGs were implicated in cell differentiation, among them PDGFRα, ITGA3, ITGB6, MLCK and MLC acted as hubs between environment information processing and cellular process, indicating that the interaction of the two categories exerts an important role in distinct fate commitment of myogenic and adipogenic cells. Particularly, we are first to show that up-regulation of intracellular Ca2+-MLCK and Rho-DMPK, and subsequently elevated MLC, may contribute to the distinct commitment of myogenic and adipogenic lineages via mediating cytoskeleton dynamics.
Highlights
Based on functional annotation and enrichment analysis of DEGs, and the elevated intracellular Ca2+ concentration in myogenic cells, we are first to identified that different mediation of Rho-dystrophia myotonica protein kinase (DMPK) and Ca2+-Myosin light chain kinase (MLCK) by extracellular signal molecules PDGFs and Extracellular matrix (ECM), and subsequently MLC expression, might contributed to distinct fate commitment to myogenic or adipogenic lineage via remodeling the cytoskeleton dynamics
Myogenic cells committed to multi-nuclei myotubes and myogenic-specific genes such as myoblast determination protein 1 (MyoD1) and myogenic factor 5 (Myf5) were highly expressed
In agreement with the above results, RNA-seq data showed that adipogenic cells had a higher level of mRNA expression of PPARγ, a key transcription factor for adipogenesis, compared with myogenic cells (FDR < 0.05), and a lesser level of MyoD1, a key transcription factor for myogenesis (FDR < 0.05) (Fig. 2f)
Summary
All procedures performed in this experiment were approved by China Agriculture. All methods were performed in accordance with the relevant guidelines and regulations. Three 3-day-old newborn Yorkshire pigs (BW = 1.4 ± 0.2 kg) were acquired from the Beijing Pig Breeding Center and euthanized. Longissimus dorsi muscles were harvest from the last lib of pigs after slaughtered, and immediately rinsed in 75% ethanol for 3 s, transferred to PBS (Hyclone, Thermo Scientific, Marietta, OH, USA), containing antibiotics (100 U/ml of penicillin and 100 mg/ml of streptomycin) (Gibco-BRL, Grand Island, NY, USA), and immediately transferred to a biological safety cabinet (HF-1100, HK, China) for cell isolation
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