Technical note: Bovine adipocyte and preadipocyte co-culture as an efficient adipogenic model

Abstract

Reductionist studies of adipose tissue biology require reliable in vitro adipocyte culturing models. Current protocols for adipogenesis induction in stromal vascular fraction-derived preadipocytes require extended culturing periods and have low adipogenic rates. We compared the adipogenic efficiency of a 7-d co-culture model of visceral (VIS) and subcutaneous (SC) stromal vascular fraction-derived preadipocytes with mature adipocytes with a 14-d standard adipocyte differentiation protocol. We obtained preadipocytes and mature adipocytes from SC and VIS adipose tissue of nonlactating, nongestating Holstein cows (n = 6). Adipogenesis induction was performed using a standard protocol for 7 (SD7; control) or 14 d (SD14), and a co-culture model for 7 d (CC7). Culture conditions, including medium composition, were the same for all treatments. For CC7, 900 primary adipocytes/cm2 were placed in 0.4-μm transwell inserts and co-cultured with preadipocytes for adipogenesis induction. Both CC7 and SD14 similarly stimulated gene expression of adipogenic genes such as ADIPOQ, CEBPA, and CEBPB in VIS and SC. The CC7 increased triacylglycerol accumulation compared with SD14 and SD7. CC7 augmented triacylglycerol accumulation by 40- and 16-fold in SC and VIS compared with 22- and 4-fold increment in SD14, respectively. Lipolytic responses to 2-h β-adrenergic stimulation with 1 µM isoproterenol were higher in CC7 and SD14 than SD7 in SC; CC7 increased glycerol release compared with SD7 in VIS but SD7 and SD14 had similar responses. Overall, CC7 was more efficient in inducing adipogenesis in preadipocytes from VIS and SC than SD14. Furthermore, CC7 stimulated similar lipolysis and lipogenic responses than SD14 but in a shorter time. The adipogenic approach of co-culturing preadipocytes with mature adipocytes will improve the use of reductionist models to study adipocyte physiology in dairy cows and the assessment of pharmacological or nutritional interventions for enhancing dairy cow health and production.