title: Regulation of the steroidogenic response of cultured human granulosa cells: effects of serum and 25-hydroxycholesterol.
abstract: OBJECTIVE
  To examine the effect of serum and 25-hydroxycholesterol on steroidogenesis in cultured human granulosa cells from women undergoing assisted fertilization.
  
  
  METHODS
  Retrospective.
  
  
  METHODS
  Private Fertility Clinic and National Research Institute.
  
  
  METHODS
  Women undergoing IVF-ET or GIFT programs.
  
  
  RESULTS
  In serum-free medium P production decreased significantly with culture time (2, 4, 6, and 8 days: 566 +/- 128, 161 +/- 50, 71 +/- 16, and 36 +/- 7 ng/mL P, respectively; conversion factor to SI unit, 3.180; mean +/- SEM). The addition of 25-hydroxycholesterol (10 micrograms/mL), a substrate for steroidogenesis, did not prevent the decrease in P levels. However, P production was greater in the presence of this substrate at all times. The presence of fetal bovine serum (10% FBS) in the cultures allowed the maintenance of 75% of P production with respect to the initial time considered (at which maximal P values are detected). Cultured granulosa cells treated with 10 ng/mL LH in the presence of FBS showed an increase in the percentage of stimulation with culture time (2, 4, and 7 days: 2.4%; 54.8%, and 55.1%, respectively). This effect was not observed when 25-hydroxycholesterol was added to the cultures. Similar results to that obtained by LH were attained when steroidogenesis was stimulated with 0.1 mM dibutyryl cyclic adenosine 3':5' monophosphate (cAMP). In addition, cAMP production in response to 100 ng/mL LH in the presence of 0.1 mM methyl-isobutyl-xanthine decreased with culture time, showing a time dependency similar to that observed for P.
  
  
  CONCLUSIONS
  Our results demonstrate that the decrease in granulosa cell steroidogenic activity with culture time is inhibited by serum but not by 25-hydroxycholesterol, suggesting that other factors despite LH and cholesterol are necessary to support the luteal function.
authors.affiliation: Instituto de Biología y Medicina Experimental, University of Buenos Aires, Argentina.
authors.forename: L
authors.initials: L
authors.lastname: Dain
authors.forename: H
authors.initials: H
authors.lastname: Guerrero
authors.forename: E
authors.initials: E
authors.lastname: Polak de Fried
authors.forename: M
authors.initials: M
authors.lastname: Tesone
mesh_terms: D001790:Blood Physiological Phenomena; D003994:Bucladesine / Q000494:pharmacology; D002478:Cells, Cultured; D005260:Female; D006107:Granulosa Cells / Q000378:metabolism*; D006801:Humans; D006888:Hydroxycholesterols / Q000494:pharmacology*; D007986:Luteinizing Hormone / Q000494:pharmacology; D011374:Progesterone / Q000096:biosynthesis*
chemical_list: D006888:Hydroxycholesterols; D011374:Progesterone; D003994:Bucladesine; C007997:25-hydroxycholesterol; D007986:Luteinizing Hormone