Regulation of placental leptin expression by cAMP involves crosstalk between PKA and MAPK signaling pathways

JULIETA L. MAYMÓ; ANTONIO PÉREZ-PÉREZ; JOSÉ DUEÑAS; JUAN CARLOS CALVO; VÍCTOR SÁNCHEZ-MARGALET; CECILIA L. VARONE

ENDOCRINOLOGY

The Endocrine Society

Lugar: MD EEUU; Año: 2010 vol. 151 p. 3738 - 3751

0013-7227

Leptin, a 16 kDa protein mainly produced by adipose tissue, has been involved in the control of energy balance through its hypothalamic receptor. However, pleiotropic effects of leptin have been identified in reproduction and pregnancy, particularly in placenta, where it was found to be expressed. In the current study, we examined the effect of cAMP, in the regulation of leptin expression in trophoblastic cells. We found that dibutyryl cAMP ((Bu)2cAMP), a cAMP analog, showed a inducing effect on endogenous leptin expression in BeWo and JEG-3 cell lines when analysed by Western blot and qRT-PCR. Maximal effect was achieved at 100 ìM. Leptin promoter activity was also stimulated, evaluated by transient transfection with a reporter plasmid construction. Similar results were obtained with human term placental explants, thus indicating physiological relevance. Since cAMP usually exerts its actions through activation of PKA signaling, this pathway was analyzed. We found that CREB phosphorylation was significantly increased with (Bu)2cAMP treatment. Furthermore, cotransfection with the catalytic subunit of PKA and/or the transcription factor CREB caused a significant stimulation on leptin promoter activity. On the other hand, the cotransfection with a dominant negative mutant of the regulatory subunit of PKA inhibited leptin promoter activity. We determined that cAMP effect could be blocked by pharmacologic inhibition of PKA or adenylyl ciclase in BeWo cells and in human placental explants. Thereafter, we decided to investigate the involvement of the MAPK/ERK signaling pathway in the cAMP effect on leptin induction. We found that 50 ìM PD98059, a MEK inhibitor, partially blocked leptin induction by cAMP, measured both by Western blot and reporter transient transfection assay. Moreover, ERK 1/2 phosphorylation was significantly increased with (Bu)2cAMP treatment and this effect was dose dependent. Finally, we observed that 50 ìM PD98059 inhibited cAMP dependent phosphorylation of CREB in placental explants. In summary, we provide some evidence suggesting that cAMP induces leptin expression in placental cells and that this effect seems to be mediated by a crosstalk between PKA and MAPK signaling pathways.