Sertoli cells isolated from testes of 20-dayold rats and maintained in primary culture synthesized estradiol-17i, [1,3,5(10)estratriene-3,17IB-diolI (measured by specific radioimmunoassay) when testosterone (17fl-hydroxy-4-androsten-3-one) 0.5 IMM, was added to the culture medium. No detectable estradiol synthesis occurred when cells were incubated in medium containing pregnenolone (3f,-hydroxypregn-5-en-20-one), 0.5 AM, or The secretion of estrogens is a well-recognized function of the mammalian testes (1, 2), although the cellular source of this class of steroids has not been established. Hunt and Budd (3) implicated the Leydig cell as the site of testicular estrogen synthesis on the basis of evidence of feminization in men with interstitial cell tumors. Support for this conclusion came from the observations of Maddock and Nelson (4) that prolonged administration of human chorionic gonadotropin (HCG) to normal and hypogonadal men resulted in increased urinary estrogen excretion. Upon histological examination the Leydig cells appeared to be stimulated, whereas the germinal epithelium regressed and the Sertoli cells underwent changes described as "degenerative" (4).On the other hand, feminization of male dogs (5) and humans (6) with Sertoli cell tumors indicated that the Sertoli cell may be involved in estrogen synthesis. Further evidence was obtained by the demonstration that canine Sertoli cell tumors contained substantial amounts of estrogenic material (7,8).Recently, methods have been reported for the isolation and maintenance in primary cultures of Sertoli cells obtained from immature rat testes (9, 10). These cultures have been shown to respond to added follicle-stimulating hormone (FSH) and dibutyryl-adenosine 3':5' cyclic monophosphate (Bt2cAMP) (11-13). The development of a method for the isolation of Sertoli cells provided us with a direct means of determining whether Sertoli cells are capable of estrogen biosynthesis. The present studies were undertaken with this goal in mind, as well as to examine possible gonadotropic requirements for estrogen biosynthesis by Sertoli cells.
MATERIALS AND METHODSMaterials and Animals. Ovine luteinizing hormone (LH) (NIH-LH-S18; potency 1.03 NIH-LH-S1 units/mg and containing less than 0.050 NIH-FSH-S1 units/mg) and ovine FSH (NIH-FSH-S1O; potency 1.10 NIH-FSH-S1 units/mg and containing less than 0.010 NIH-LH-S1 units/mg) were provided through the Hormone Distribution Office, National Institutes of Health, Bethesda, Md. Highly purified FSH (G4-150C; having an activity of 50 NIH-FSH-S1 units/mg in the Steelman-Pohley assay and 0.02 NIH-LH-S1 units/mg in the ovarian ascorbic depletion test) was a gift from Dr. H. Papkoff. N6,02'-Dibutyryl-adenosine 3':5' cyclic monophosphate (Bt2cAMP) was obtained from Sigma Chemical Co. Testosterone (17fl-hydroxy-4-androsten-3-one), pregnenolone (3f3-hydroxypregn-5-en-20-one), collagenase type 1 (125 units/mg of solid), deoxyribonuclease 1 (crude, from beef pancreas, 780 Kunitz units/mg), and soybean trypsin inhibitor (type IS...