Hum. Reprod. Advance Access originally published online on August 11, 2009
Human Reproduction 2009 24(11):2890-2901; doi:10.1093/humrep/dep292
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Role of adiponectin receptors, AdipoR1 and AdipoR2, in the steroidogenesis of the human granulosa tumor cell line, KGN
1 INRA, UMR 85, Unité de Physiologie de la Reproduction et des Comportements F-37 380, Nouzilly, France 2 Unité d'Endocrinologie, de Diabétologie et des Maladies Métaboliques, CHRU Bretonneau 37000, Tours, France 3 Université de Lyon, (UCB-lyon1), IFR128, Lyon F-69007, France 4 INSERM, U758, Lyon F-69007, France 5 Ecole Normale Supérieure de Lyon, Lyon F-69007, France
6 Correspondence address. Unité de Physiologie de la Reproduction et des Comportements, Institut National de la Recherche Agronomique, 37380 Nouzilly, France. Tel: +33-2-47-42-77-89; Fax: +33-2-47-42-77-43; E-mail: jdupont{at}tours.fr
BACKGROUND: Adiponectin is involved in the regulation of energy homeostasis and more recently in the reproductive functions. We have previously shown that adiponectin receptors (AdipoR1 and AdipoR2) are expressed in human granulosa cells. However, it remains to be investigated whether both AdipoR1 and AdipoR2 or only one of these receptors serve as the major receptor(s) for adiponectin in human granulosa cells.
METHODS: The RNA interference (RNAi) technology was used to specifically knockdown the expression of either AdipoR1 or AdipoR2. Progesterone and estradiol levels in the conditioned media were measured by radioimmunoassay, and determination of cell proliferation by tritiated thymidine incorporation. The levels of adiponectin receptors and proteins involved in the steroidogenesis and in the signalling pathways were examined by western blot.
RESULTS: We generated AdipoR1 (R1) and AdipoR2 (R2) knockdown KGN cell lines. R1 cells were apoptotic and had increased expression levels of cleaved caspase 3 and decreased levels of BAD phosphorylation and PCNA as compared with control or parental KGN cells. R2 cells had similar morphology to control or KGN cells. However, they produced less progesterone and estradiol and expressed lower levels of StAR protein in response to FSH or IGF-1 stimulation compared with control cells. Furthermore, the increase of MAPK ERK1/2 phosphorylation in response to human recombinant adiponectin and FSH was lower in R2 than control cells.
CONCLUSIONS: In the human granulosa KGN cell-line, AdipoR1 seems to be involved in the cell survival whereas AdipoR2, through MAPK ERK1/2 activation, may be implicated in the regulation of steroid production.
Key words: adiponectin/human granulosa cells/steroidogenesis/signal transduction/cell survival
Submitted on March 8, 2009; resubmitted on July 10, 2009; accepted on July 20, 2009.