Optimizing adipogenic cocktail composition to enhance beige adipogenesis and evaluate thermogenic potential in primary mouse subcutaneous fat cell cultures.

BACKGROUND: Subcutaneous white adipose tissue (WAT) is crucial for systemic metabolic homeostasis, with beige adipocytes in this depot contributing to energy metabolism through inducible thermogenesis. Differentiated adipocyte cultures derived from mouse inguinal WAT are a widely used system to study beige adipose biology and identify therapeutic targets, as they retain the genetic and epigenetic traits of native adipocytes while providing experimental flexibility. However, variability during the adipocyte induction and differentiation poses a challenge, impacting beige adipogenesis and experimental outcomes. METHODS: This study conducted an unbiased analysis of four distinct adipogenic cocktails to evaluate their effects on beige adipogenesis in inguinal stromal vascular fraction cells from wild-type and genetically modified mice, as well as on the thermogenic activation of differentiated adipocytes. RESULTS: Different combinations of adipogenic inducers, including dexamethasone, 3-isobutyl-1-methylxanthine, insulin, triiodothyronine, indomethacin, and rosiglitazone (Rosi), recruited beige adipocytes with varying levels of thermogenic characteristics. The peroxisome proliferator-activated receptor gamma agonist, Rosi, emerged as a key inducer, maximizing beige adipocyte biogenesis during the differentiation phase rather than the induction phase. However, Rosi-enhanced beige adipocyte differentiation exhibited limited thermogenic activation at the transcriptional level but not at the rapid signal transduction and real-time functional level in response to a β-adrenergic receptor agonist. CONCLUSIONS: These findings underscore the importance of optimizing adipogenic cocktails, as they significantly influence experimental outcomes. This study offers valuable guidance for selecting effective combinations of adipogenic inducers tailored to specific research objectives and relevant in vitro models of beige adipose biology.