Transcriptional regulation of leptin by hCG, cAMP and estradiol in placenta

MAYMÓ J; GAMBINO Y; PÉREZ PÉREZ A; SÁNCHEZ MARGALET V; CALVO JC; VARONE C

Seggau Castle

Congreso; International Federation of Placenta Associations Meeting; 2008

IFPA

Leptin, the 16000 MW protein product of the obese gene, was originally considered as an adipocyte-derived signaling molecule for the central control of metabolism. However, leptin has been suggested to be involved in other functions during pregnancy, particularly in placenta, where it was found to be expressed. Recently, we have demonstrated that leptin promotes proliferation and survival of trophoblastic cells (1). In the present work we aimed to study the regulation of leptin expression in placenta. Methods: BeWo cells, cultured under standard conditions, as well as human placenta explants were used. Western blot analyses were carried out to detect leptin expression as well as the phosphorylated form of proteins involved in mayor signalling pathways. Transfection assays with reporter constructs and expression vectors for some intermediates of signaling pathways were used to determine the transcriptional regulation of leptin. Results: We have found that leptin was maximally induced by 100 IU hCG/ml (12.9 times), 1 uM cAMP (4.4 times) and 100 nM estradiol (8.8 times). These effects were time and dose-dependent. Estradiol effect was partially blocked with 10 nM ICI 182780. Moreover, treatment with hCG. cAMP or estradiol enhanced leptin promoter activity evaluated by transient transfection with a plasmid construction containing different promoter regions and the reporter gene luciferase. Similar results were obtained with placental explants evidencing physiological relevance. Since hCG signal transduction usually involves cAMP signaling, this pathway was analyzed. We found that dibutyril cAMP counteracted hCG effect on leptin expression. Thereafter we determined that hCG effect could be partially blocked by pharmacologic inhibition of MAP kinase pathway with 50 mM PD98059 but not by the inhibition of the PI3K pathway with 0.1 mM Wortmannin. On the other way hCG, cAMP and estradiol effects promoted MAPK pathway as determined by ERK1/ERK2 phosphorylation in placental cells. Moreover cotransfection with a dominant negative mutant of MAPK blocked the hCG-mediated activation of leptin expression.These signalling pathways were confirmed in explants obtained from placenta of healthy donors. Conclusions: All these findings suggest that leptin expression is regulated by multiple placental effectors. However, further studies are needed to explain the molecular mechanisms underlying these effects, our results support the importance of leptin in the biology of reproduction.