Insulin-stimulated glucose metabolism plays a key role in the regulation of leptin mRNA expression and protein secretion. However, it is not known whether stimulation of leptin production by glucose metabolism is regulated at the level of promoter activation or at a step distal to the promoter. Therefore, in order to investigate the transcriptional regulation of the leptin promoter by insulin-stimulated glucose metabolism, 3T3-L1 cells were transfected with a plasmid containing the leptin promoter driving a luciferase reporter gene. Leptin promoter activity was increased after 48 hours of treatment by 219 +/- 64 (p = 0.028) and 225 +/- 69% (p = 0.046) at insulin concentrations of 16 and 160 nM, respectively. The activation of the leptin promoter induced by insulin (16 nM) was markedly inhibited by 2-deoxy-D-glucose (2-DG, 50 mg/dl), a competitive inhibitor of glucose metabolism. The increment of insulin-stimulated leptin promoter activation was reduced by 52 +/- 11% (p = 0.028 vs insulin alone). The activity of a control plasmid (pGL2-Control) was unaffected by insulin or 2-DG. These results provide strong evidence that insulin-stimulated glucose metabolism, and not insulin per se, mediates the effects of insulin to increase the transcriptional activity of the leptin promoter.

Cardiotrophin (CT)-1 is a regulator of glucose and lipid homeostasis. In the present study, we analyzed whether CT-1 also acts to peripherally regulate metabolic rhythms and adipose tissue core clock genes in mice. Moreover, the circadian pattern of plasma CT-1 levels was evaluated in normal-weight and overweight subjects. The circadian rhythmicity of oxygen consumption rate (Vo₂) was disrupted in aged obese CT-1-deficient (CT-1^-/-) mice (12 mo). Although circadian rhythms of Vo₂ were conserved in young lean CT-1^-/- mice (2 mo), CT-1 deficiency caused a phase shift of the acrophase. Most of the clock genes studied (Clock, Bmal1, and Per2) displayed a circadian rhythm in adipose tissue of both wild-type (WT) and CT-1^-/- mice. However, the pattern was altered in CT-1^-/- mice toward a lower percentage of the rhythm or lower amplitude, especially for Bmal1 and Clock. Moreover, CT-1 mRNA levels in adipose tissue showed significant circadian fluctuations in young WT mice. In humans, CT-1 plasma profile exhibited a 24-h circadian rhythm in normal-weight but not in overweight subjects. The 24-h pattern of CT-1 was characterized by a pronounced increase during the night (from 02:00 to 08:00). These observations suggest a potential role for CT-1 in the regulation of metabolic circadian rhythms.-López-Yoldi, M., Stanhope, K. L., Garaulet, M., Chen, X. G., Marcos-Gómez, B., Carrasco-Benso, M. P., Santa Maria, E. M., Escoté, X., Lee, V., Nunez, M. V., Medici, V., Martínez-Ansó, E., Sáinz, N., Huerta, A. E., Laiglesia, L. M., Prieto, J., Martínez, J. A., Bustos, M., Havel, P. J., Moreno-Aliaga, M. J. Role of cardiotrophin-1 in the regulation of metabolic circadian rhythms and adipose core clock genes in mice and characterization of 24-h circulating CT-1 profiles in normal-weight and overweight/obese subjects.