ICEECE2012 Oral Communications Female Reproductioin Basic (6 abstracts)
LH and hCG produce different responses in human granulosa lutein cells in vitro
L. Casarini 1 , E. Pignatti 1 , A. La Marca 2 , T. Marsella 2 , D. Tagliasacchi 2 , F. Adversi 2 , M. Lispi 3 , S. Longobardi 3 & M. Simoni 1
1University of Modena and Reggio Emilia, Modena, Italy; 2University Hospital of Modena, Modena, Italy; 3Merck KGaA, Rome, Italy.
Introduction: LH and hCG act on the same receptor (LHCGR), but it is not known whether they elicit the same cellular and molecular response. This study aims at comparing the activation of cell-signalling pathways and gene expression in response to LH and hCG.
Design: We evaluated the activation of cAMP, ERK and AKT-pathways and progesterone production by ELISA and Western blotting in human primary granulosa cells (hGLC) stimulated for up to 36 hours with equipotent doses of LH or hCG. The expression of target genes was measured by real-time PCR after 12-hours of stimulation with either gonadotropin, using specific ERK or AKT-pathways inhibitors (U0126 and LY294002) to differentiate their effects on gene expression.
Results: Continuous exposure of hGLC to equipotent doses of LH or hCG for 36 hours revealed that the intracellular cAMP production is pulsatile with a frequency of about 3–4 hours, with significantly higher stimulation by hCG vs LH (t-test; P<0.05; n=3), despite no differences in progesterone production, which increased progressively. Conversely, phospho-ERK and -AKT activation was more potent and sustained by LH vs hCG over 1 hour (t-test; P<0.05; n=4). hCG was almost inactive on AKT. Finally, LH significantly increased the expression of NRG-1 and CYP19A1 genes after ERK inhibition, while hCG decreased the expression of AREG after AKT inhibition (t-test; P<0.05; n=3).
Conclusions: In terms of cAMP production, hGLC respond to equipotent, constant LH or hCG stimulation in a pulsatile fashion. Acutely, hCG is more potent on cAMP production, while LH is more potent on the ERK and AKT activation. The early inhibition of ERK and AKT results in the differential induction of expression of target genes depending on ligand, indicating that the LHCGR is able to differentiate the activity of LH and hCG.
Declaration of interest: The authors declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research project.
Funding: This research did not receive any specific grant from any funding agency in the public, commercial or not-for-profit sector.
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