Immortalization of Adipose-Derived Stromal Cells From Japanese Black Cattle

Adipose tissue plays a critical role in determining the characteristic palatability of Japanese Black cattle beef, including its distinctive aroma, tenderness, and flavor. These quality traits are closely associated with adipogenic differentiation and lipid metabolism; however, the underlying molecular mechanisms remain poorly understood. Adipose-derived stromal cells (ASCs) residing within adipose tissue possess the capacity to differentiate into adipocytes and other mesenchymal lineages and are, therefore, considered valuable experimental resources for adipose tissue research. Nevertheless, primary cells are subject to several limitations, including a restricted proliferative lifespan caused by culture-induced stress and reduced experimental reproducibility due to inter-individual variability, which hamper their use in long-term in vitro analyses. In this study, we immortalized ASCs derived from Japanese Black cattle using the K4DT method, which involves the introduction of the R24C mutant form of cyclin-dependent kinase 4 (CDK4^R24C), Cyclin D1, and telomerase reverse transcriptase. The established bovine ASCs (bASCs) [bASCs-K4DT] exhibited a stable proliferative capacity during long-term culture while retaining their ability to undergo adipogenic differentiation. These immortalized bASCs may serve as a useful in vitro model for elucidating the molecular mechanisms underlying adipocyte differentiation and formation of breed-specific aroma and flavor characteristics.