Pig Leydig cell culture: a useful in vitro test for evaluating the testicular toxicity of compounds.
In vivo studies have shown that the testis is a target organ for drugs and chemicals. In order to evaluate the testicular toxicity of compounds and to identify the mechanisms of their toxicity, we have developed a miniaturized primary culture of immature pig Leydig cells. Five well-known drugs with differing mechanisms of toxicity on testicular functions were tested to validate the model. Testosterone and progesterone secretion were measured to evaluate testicular function. Cell viability was assessed quantitatively using a colorimetric assay based on the reduction of a tetrazolium salt (3-(4,5-dimethylthiazol)-2,5-diphenyltetrazolium bromide) which stains viable cells only, thus allowing discrimination between specific inhibitors of Leydig cell function and nonspecific cytotoxic drugs. Ketoconazole and aminoglutethimide inhibited both testosterone and progesterone secretion, but without modifying cell viability. Spironolactone specifically blocked testosterone secretion and increased progesterone concentration without inducing cell mortality. Cycloheximide altered testicular steroid secretion by another mechanism of action. Chlorpromazine, which interferes with the secretion of gonadotropins in vivo, produced a significant inhibition of progesterone and testosterone secretion as a result of the cytotoxic effects of the drug. In conclusion, this in vitro test enables one to discriminate accurately between specific and nonspecific inhibitors of steroidogenesis and could reduce the number of false positives when screening for potential testicular toxins.
Brun
H P
HP
Institut de Recherche sur la Sécurité du Médicament, Rhone-Poulenc Rorer, Centre de Recherche de Vitry-Alfortville, Vitry sur Seine, France.
Leonard
J F
JF
Moronvalle
V
V
Caillaud
J M
JM
Melcion
C
C
Cordier
A
A
D000616:Aminoglutethimide / Q000633:toxicity; D000818:Animals; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002746:Chlorpromazine / Q000633:toxicity; D006063:Chorionic Gonadotropin / Q000494:pharmacology; D000242:Cyclic AMP / Q000096:biosynthesis; D003513:Cycloheximide / Q000633:toxicity; D007654:Ketoconazole / Q000633:toxicity; D007700:Kinetics; D007985:Leydig Cells / Q000166:cytology / Q000187:drug effects*; D008297:Male; D011374:Progesterone / Q000096:biosynthesis; D013148:Spironolactone / Q000633:toxicity; D013256:Steroids / Q000096:biosynthesis; D013552:Swine; D013737:Testis / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D013739:Testosterone / Q000096:biosynthesis; D013997:Time Factors; D014116:Toxicology / Q000379:methods
D006063:Chorionic Gonadotropin; D013256:Steroids; D000616:Aminoglutethimide; D013148:Spironolactone; D013739:Testosterone; D011374:Progesterone; D003513:Cycloheximide; D000242:Cyclic AMP; D007654:Ketoconazole; D002746:Chlorpromazine