REGULATION OF OBESE MESSENGER-RNA EXPRESSION BY HORMONAL FACTORS IN PRIMARY CULTURES OF RAT ADIPOCYTES

The obese (ob) gene has been cloned recently and its protein product i s called ''leptin''. Leptin is an adipocyte-derived satiety factor tha t regulates body weight homeostasis. Several hormonal factors have bee n reported to regulate ob mRNA expression. To determine which factors are most important for regulation of ob mRNA expression, we examined t he effects of insulin, dexamethasone, a beta(3)-adrenergic agonist (CG P12177A), 8-bromo-cAMP, 8-bromo-cGMP and 1-methyl-3-isobutylxanthine ( MIX) on primary cultured adipocytes. Rat adipocytes obtained from epid idymal fat were cultured using the ceiling method. Total RNA was extra cted and the expression of ob mRNA was measured by quantitative revers e transcription-polymerase chain reaction. After 24 h of incubation, 1 00 nmol/l insulin significantly increased the expression of ob mRNA (2 1.4-fold compared to control). Moreover, insulin increased ob mRNA exp ression in a dose-dependent manner over a range of 1-100 nmol/l. The e ffect of 100 nmol/l insulin was similar to that seen with 20% newborn calf serum. Dexamethasone (25-1000 nmol/l) also increased ob mRNA expr ession (2.5-2.9-fold). The effect of dexamethasone occurred more rapid ly than insulin. CGP12177A (1-10 mu mol/l) and 0.5 mmol/l 8-bromo-cAMP had no effects, whereas 0.5 mmol/l 8-bromo-cGMP and 0.5 mmol/l MIX ha d stimulatory effects (2.8- and 2.4-fold increase in ob mRNA, respecti vely). The combination of 250 nmol/l dexamethasone and 0.5 mmol/l MM d id not have an additive effect on ob mRNA levels. Our present data sug gest that, of these agents, insulin is the most important factor regul ating ob mRNA expression.