Thyroid hormone inhibits aromatase activity in porcine thecal cells cultured alone and in coculture with granulosa cells

We cultured porcine thecal and granulosa cells alone or in coculture to def ine whether thyroid hormone affects aromatase activity in porcine ovarian c ells. Dispersed cells were cultured with 10(-9) M triiodothyronine (T-3) fo r 24 hours. Testosterone (final concentration 10(-7) M) was added as aromat ase substrate for granulosa cells (Gc) cultured alone. Thecal (Th) androgen s serve as a substrate for estradiol secretion by Th cells cultured alone a nd in coculture with Gc. At the end of the preincubation time, the culture media was removed and replaced with fresh media containing 100 ng follicle stimulating hormone (FSH) or 10(-3) M 8-bromo-cyclic adenosine monophosphat e (8-Br-cAMP). After overnight incubation, the culture media was analyzed f or estradiol production by radioimmunoassay (RIA). T-3 inhibited basal, FSH -stimulated, and 8brcAMP-stimulated estradiol production in all culture con ditions. T-3 inhibited cAMP analogue 8-Br-cAMP and FSH-induced aromatase ac tivity to a similar extent, thus suggesting that the inhibitory effect of T -3 is downstream of cAMP formation. In the second part of the experiment a rabbit polyclonal antibody against human placental cytochrome P-450(arom) w as used to confirm the effect of T-3 on aromatase protein in Th and Gc. Pre treatment of Th and Cc with T-3 markedly decreased immunostaining for aroma tase in both types of cells, suggesting a direct effect of T-3 on this enzy me.