Leptin induces endothelin-1 in endothelial cells in vitro

Leptin, a protein encoded by the obese gene, is produced by adipocytes and released into the bloodstream. In obese humans, serum leptin levels are increased and correlate with the individual's body mass index and blood pressure. Elevated serum concentrations of endothelin-1 (ET-1), a potent vasoconstrictor and mitogen, were also observed in obese subjects. The pathomechanisms underlying this ET-1 increase in obesity are poorly understood. In the present study, we investigated the influence of the ob gene product leptin on the expression of ET-I in human umbilical vein endothelial cells (HUVECs). Binding studies using (125)I-radiolabeled leptin revealed high- and low-affinity leptin binding sites on HUVECs (Kd1=13.1+/-3.1 nmol/L and Kd2=1390+/-198 nmol/L, respectively), mediating a time- and dose-dependent increase of ET-1 mRNA expression and protein secretion after incubation of HUVECs with leptin. This leptin-induced ET-1 expression was inhibited by preincubation of HUVECs with 0.75 mumol/L antisense phosphorothioate oligonucleotides directed against the leptin receptor Ob-Rb. Furthermore, after incubation with leptin, increased nuclear staining of c-fos and c-jun, the major components of the transcription factor AP-1, and increased AP-1 DNA binding were observed. Transient transfection studies with ET-1 promoter constructs showed that leptin-induced promoter activity was abolished in the absence of AP-1 binding sites or by cotransfection with a plasmid overexpressing a mutated jun, which is able to bind c-fos but not DNA. Thus, leptin upregulates ET-1 production in HUVECs via a mechanism potentially involving jun binding members of the bZIP family.