Bladder organoid-conditioned media as a serum-free booster for myoblast proliferation

In recent years, the growing attention to sustainable food production and animal welfare has positioned cultured meat technologies. A key technical requirement for implementing this approach is the development of culture media independent of fetal bovine serum (FBS). 3D organoids are composed of diverse stem and differentiated cells that replicate the structure and function of living tissues. Therefore, we hypothesized that the growth factors secreted from 3D organoids could replace the FBS. In this study, we evaluated the effects of conditioned medium prepared from organoid supernatants derived from various mouse organs, including the lung, gallbladder, spleen, kidney, and bladder, on myoblast proliferation. Among these, mouse bladder organoid-derived supernatant (MBOS) showed the most pronounced proliferative effect on C2C12 cells, a myoblast cell line. RNA sequencing and flow cytometry analyses revealed that MBOS-treated C2C12 cells upregulated cell cycle-related genes ( CCNB1 and CDK1 ) and increased the proportion of cells in the G2/M phase. Although MBOS contained a higher concentration of insulin-like growth factor (IGF)-1 than the control medium, neutralization with IGF-1 antibody did not reduce the proliferative effect. MBOS significantly promoted the proliferation of primary bovine myoblasts, suggesting its cross-species applicability. These findings suggest that MBOS is a promising candidate for FBS-free culture media with the potential to produce cultured meat.