title: Gossypol inhibits human chorionic gonadotropin-stimulated testosterone production by cultured canine testicular interstitial cells.
abstract: Gossypol (GP) is a natural polyphenolic compound that possesses antifertility and antisteroidogenic activities in both males and females. The dog is highly sensitive to GP toxicity, yet GP's effect on canine testicular steroidogenesis has never been reported. Thus, the present study examines GP's effects on human chorionic gonadotropin (hCG)-induced testosterone (T) production by primary cultured canine testicular interstitial cells. After decapsulation and enzymatic dissociation of canine testes in Dulbecco's Modified Eagle Medium with Ham's Nutrient Mixture F-12 (1:1; DME/F-12) containing 0.1% collagenase, 0.1% BSA, and 10 micrograms/ml DNase 1 (37 degrees C, 20 min), interstitial cells were isolated by sedimentation and filtration (140 microns) and then cultured in supplemented DME/F-12 medium (5 micrograms/ml insulin, 5 micrograms/ml transferrin, 5 ng/ml sodium selenite; DME/F-12/S) containing 0.1% fetal bovine serum (FBS). FBS was used to enhance cell attachment during the first 24 hours of culture. After 24 hours, the medium was replaced with serum-free DME/F-12/S and the cells were cultured for an additional 24 hours. Thereafter, cells were treated with hCG (0.1 IU/ml) alone and in combination with GP (0.05, 0.5, 2.5 and 5.0 microM). Media were collected for T radioimmunoassay and cells for protein estimation after 8, 16 and 24 hours of treatment. Treatment with hCG significantly (p < 0.05) stimulated T production over that of controls at all treatment times examined. At 8, 16 and 24 hours, T secretion was elevated from 0.91 +/- 0.25, 1.32 +/- 0.42, and 1.41 +/- 0.40 pg/microgram protein to 2.36 +/- 0.50, 2.84 +/- 0.60, and 2.82 +/- 0.43 pg/microgram protein, respectively. At 0.5, 2.5 and 5.0 microM, GP significantly (p < 0.05) reduced hCG-induced T secretion at 16 and 24 hours of treatment to 1.79 +/- 0.50, 1.62 +/- 0.12, 1.34 +/- 0.16 (16 hr), and 1.53 +/- 0.38, 1.43 +/- 0.11, 1.42 +/- 0.32 (24 hr) pg/microgram protein, respectively. At 8 hours, T production was reduced by 2.5 and 5.0 microM GP to 1.08 +/- 0.55 and 0.93 +/- 0.61 pg/microgram protein, respectively. GP, however, did not reduce T production to below basal levels. These results demonstrate the inhibition of hCG-induced T production by GP in cultured canine testicular interstitial cells.
authors.affiliation: Department of Veterinary Physiology and Pharmacology, College of Veterinary Medicine, Ohio State University, Columbus 43210-1092, USA.
authors.forename: M
authors.initials: M
authors.lastname: Mushtaq
authors.forename: S
authors.initials: S
authors.lastname: Kulp
authors.forename: W
authors.initials: W
authors.lastname: Chang
authors.forename: Y C
authors.initials: YC
authors.lastname: Lin
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D006063:Chorionic Gonadotropin / Q000037:antagonists & inhibitors* / Q000494:pharmacology; D004285:Dogs; D006072:Gossypol / Q000494:pharmacology*; D006801:Humans; D007985:Leydig Cells / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D014176:Protein Biosynthesis; D011863:Radioimmunoassay; D013739:Testosterone / Q000096:biosynthesis*
chemical_list: D006063:Chorionic Gonadotropin; D013739:Testosterone; D006072:Gossypol