Comparison of secreted miRNAs and proteins during proliferation and differentiation of bovine satellite cells in culture implies potential roles in regulating myogenesis

Abstract

Cultivated meat is an emerging new technology to produce sustainable meat for the future. The common approach for cultivated meat, is the isolation of satellite cells from donor animals, followed by in vitro proliferation and differentiation into primitive muscle fibers. The transformation of satellite cells into myofibers is tightly orchestrated by intra-cellular signaling, while the inter-cellular signaling is less well understood. Thus, the current study was conducted to map the secretion of potential signaling molecules (MicroRNAs and proteins) during proliferation and differentiation. Primary cultures of satellite cells were grown to 50% and 80% confluence, representing the proliferative phase or serum-starved for 1 and 3 days to induce differentiation. Post incubation in FBS-free media, the media were collected and analyzed for miRNA and protein content using gene-arrays and LC-MS/MS, respectively. When comparing the miRNA secretome at 50% and 80% confluence, we observed four differentially expressed miRNA, while only five were differentially expressed when comparing Day 1 to Day 3. A subsequent in silico analysis suggested that pathways of importance for myogenesis, e.g., MAPK and AMPK signaling, could be regulated by the secreted miRNAs. In addition, >300 proteins were secreted, including insulin-like growth factor 1 binding proteins 2, 3, 4, 5 and 6. In conclusion, this study demonstrated differential secretion of several miRNAs and proteins during both proliferation and differentiation of bovine satellite cells in vitro.

Keywords: Cultivated meat; Extracellular vesicles; In vitro; In vitro meat; Skeletal muscle; Tissue engineering.