COMPLETE ADIPOSE DIFFERENTIATION OF 3T3 L1-CELLS IN A CHEMICALLY DEFINED MEDIUM - COMPARISON TO SERUM-CONTAINING CULTURE CONDITIONS

The hormonal control of the adipose conversionof 3T3 Ll cells was studied in a chemically defined medium and compared to that under serum-containing conditions. Addition of insulin to a chemically defined medium (Dulbecco’s Modified Eagle’s Medium-Ham’s F-12, 1:1) resulted in a dose-dependent increase in glycero-3-phosphate dehydrogenase activity, a lipogenic marker enzyme. The maximally effective concentration of insulin was in the range of 10-8and 10-7mol/liter. Glucocorticoids were ineffective in the absence of insulin and exhibited an inhibitory action in serum-containing medium. In the presence of insulin, corticosterone and other glucocorticoids showed a potent adipogenic activity, with a maximally effective concentration at 10-8mol/1. 1-Methyl-3-isobutylxanthine (MIX), a cAMP-elevating agent, was also found to induce the adipose differentiation process in the presence of insulin. The effect of MIX could be partly replaced by (Bu)2cAMP. A combination of insulin and, for the first 2 days, glucocorticoids and MIX promoted a complete conversion of 3T3 cells within 6-8 days. The concentrations of the factors required for adipose conversion were substantially lower than those in serum-containing medium. The adipose conversion in serum-free medium was associated with an increase in cell number, since 0.3 µg/ml cytosin arabinoside not only inhibited cell replication, but also prevented the expression of glycero-3-phosphate dehydrogenase activity. Our results demonstrate a complete adipose differentiation of 3T3 Ll cells in a chemically defined culture system supplemented with insulin, glucocorticoids, and factors that increase the cAMP content of the cells. This system may provide an improved model to study adipogenic and antiadipogenic factors under defined conditions. © 1990 by The Endocrine Society.