FSH receptor in vitro modulation by testosterone and hCG in human luteinized granulosa cells
Eur J Obstet Gynecol Reprod Biol. Dec.
2012 doi: 10.1016/j.ejogrb.2012.08.020
OBJECTIVE: To investigate the effect of testosterone and hCG on FSH receptor (FSHR) protein and mRNA expression in human granulosa cells (GC) in vitro. STUDY DESIGN: Experimental in vitro cell culture obtained from healthy women undergoing IVF/ICSI due to male factor infertility. Human follicular fluid samples were obtained and after cumulus-oocyte complexes were identified, fluids were pipetted onto Ficoll gradients and centrifuged for 15 min at 400 x g at room temperature. Cells at the interface were removed and plated in 24-well plates for 3 days in M-199 with 10% FBS. Cells were treated with different concentrations of testosterone and hCG. After purification, cells were labeled with specific antibodies and the protein expression of the FSHR was evaluated by flow cytometry in the GC population. Also, total RNA was extracted from confluent GC and the FSHR gene expression was evaluated by RT-PCR. RESULTS: FSHR expression was modulated by treating GC in vitro at different testosterone/hCG concentrations. When compared with untreated GC, we observed a significant effect of testosterone and hCG on the expression of the FSHR at the protein level. Time course experiments confirmed that the gene expression of the FSHR peaked at 12-24h when testosterone or hCG was used as a stimulus. CONCLUSIONS: Both testosterone and hCG are able to positively modulate FSHR expression at gene and protein level in human GC in vitro.