title: Effect of Anti-Müllerian hormone (AMH) and bone morphogenetic protein 15 (BMP-15) on steroidogenesis in primary-cultured human luteinizing granulosa cells through Smad5 signalling.
abstract: OBJECTIVE
  To determine if there is any effect of AMH and BMP-15 on estradiol and progesterone production from primary-cultured human luteinizing granulosa cells, to delineate what is the effect of FSH on their actions and which are the possible mechanisms involved.
  
  
  METHODS
  Luteinizing granulosa cells (GCs), obtained from follicular fluid of 30 women undergoing in vitro fertilization, were cultured, after a short 24-h preincubation period, in serum-free medium for 24 or/and 48 h in the presence/absence of various concentrations of AMH, BMP-15 and FSH alone or in combinations. Estradiol and progesterone production, SMAD5 phosphorylation and StAR expression were studied in parallel. Steroids were measured in culture-supernatant using enzyme-immunoassays, while Smad5-signaling pathway activation and StAR protein expression were assessed immunocytochemically.
  
  
  RESULTS
  We found that the treatment of AMH in GCs for 24/48 h attenuated FSH-induced estradiol production (p < 0.001), had no effect on basal estradiol levels, decreased basal progesterone production (p < 0.001) and FSH-induced StAR expression (p < 0.001). On the other hand, BMP-15 decreased basal estradiol levels (p < 0.001) and attenuated FSH-induced estradiol production (p < 0.001). Furthermore, BMP-15 reduced progesterone basal secretion (p < 0.001), an effect that was partially reversed by FSH (p < 0.01), probably via increasing StAR expression (p < 0.001). FSH-induced StAR expression was also attenuated by BMP-15 (p < 0.001). FSH, AMH and BMP-15 activated Smad-signaling pathway, as confirmed by the increase of phospo-Smad5 protein levels (p < 0.001 compared to control).
  
  
  CONCLUSIONS
  AMH and BMP-15 by interacting with FSH affect the production of estradiol and progesterone from cultured luteinizing-granulosa cells possibly via Smad5-protein phosphorylation.
authors.affiliation: Department of Obstetrics and Gynaecology, School of Health Sciences, Faculty of Medicine, University of Thessaly, 41110, Viopolis, Larissa, Greece.
authors.forename: Ermioni
authors.initials: E
authors.lastname: Prapa
authors.forename: Anna
authors.initials: A
authors.lastname: Vasilaki
authors.forename: Konstantinos
authors.initials: K
authors.lastname: Dafopoulos
authors.forename: Eleni
authors.initials: E
authors.lastname: Katsiani
authors.forename: Panagiotis
authors.initials: P
authors.lastname: Georgoulias
authors.forename: Christina I
authors.initials: CI
authors.lastname: Messini
authors.forename: George
authors.initials: G
authors.lastname: Anifandis
authors.forename: Ioannis E
authors.initials: IE
authors.lastname: Messinis
mesh_terms: D000328:Adult; D054304:Anti-Mullerian Hormone / Q000378:metabolism* / Q000494:pharmacology; D055430:Bone Morphogenetic Protein 15 / Q000378:metabolism* / Q000494:pharmacology; D002478:Cells, Cultured; D004958:Estradiol / Q000378:metabolism; D005260:Female; D005640:Follicle Stimulating Hormone / Q000378:metabolism / Q000494:pharmacology; D006107:Granulosa Cells / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D008875:Middle Aged; D011374:Progesterone / Q000378:metabolism; D015398:Signal Transduction; D051902:Smad5 Protein / Q000378:metabolism*; D055815:Young Adult
chemical_list: D055430:Bone Morphogenetic Protein 15; C497797:SMAD5 protein, human; D051902:Smad5 Protein; D011374:Progesterone; D004958:Estradiol; D054304:Anti-Mullerian Hormone; D005640:Follicle Stimulating Hormone