Optimization of culture conditions of bovine adipose stem cells for cultivated fat development

Cultivated meat offers a sustainable alternative to meet the global demand for meat and to overcome the environmental problems of traditional livestock production. In addition to muscle, fat also plays an important role in influencing flavour, texture, and moisture. Cultured adipocytes represent a potential solution to improve the organoleptic properties of cultivated meat or meat alternatives. However, current strategies for the in vitro cultivation of ruminant adipogenic cells have significant limitations. The effects of culture medium composition (with or without serum) and culture type (2D vs. 3D) on the viability, proliferation, and adipogenic differentiation of bovine adipose stem cells (bASCs) were investigated. Cell viability and proliferation were assessed using live/dead and metabolic assays. After a 14-day induction period, adipogenic capacity was assessed by immunofluorescence staining, triglyceride determination, and semi-quantitative RT-PCR of peroxisome proliferator-activated receptor gamma (PPAR-g) and fatty acid binding protein 4 (FABP4). In the 2D culture system, the serum-free formulation improved the proliferation and adipogenic differentiation of bASC. The 3D spheroid culture provided an optimal environment for inducing adipocyte maturation, as evidenced by significant changes in the transcriptional activity of PPAR-g and FABP4 genes, lipid accumulation, and triglyceride content. We found that enhanced adipogenesis from bASCs can be achieved by serum-free media and 3D spheroid-based culture. Further optimization of parameters may be required to maximize the yield of adipocytes within a short culture time. These results provide valuable insights into differentiation strategies for bovine preadipocytes and may contribute to efficient and sustainable approaches to fat cultivation.