Effect of type 1 transforming growth factor-β on the level of aromatase cytochrome P-450 in human fetal hepatocytes

Aromatase cytochrome P-450 (P-450(AROM)) is the enzyme in the steroidogenic pathway controlling the formation of oestrogens from 19 carbon steroid precursors. Aromatase is present in various tissues of the human fetus. The liver is second only to the placenta in the level of P-450(AROM) activity in the fetus. In this study we examined the effects of type 1 transforming growth factor-β (TGFβ) on P-450(AROM) expression in human fetal (HF) hepatocytes. The HF hepatocytes were dispersed into single cells which were placed into monolayer cell culture until confluent. Cells were then rinsed and treated in serum-free media with dibutyryl cyclic AMP (Bu₂cAMP) for 72 h. Treatment with Bu₂cAMP (2 mmol/l) caused a fivefold increase in aromatase activity in hepatocytes. The increase in aromatase activity apparently represented an increase in P-450(AROM) enzyme as determined by immunoblotting using an antibody directed against human placental aromatase. TGFβ blocked basal, as well as Bu₂cAMP increases, in aromatase activity by over 50%. The effect of TGFβ was dose-dependent with maximal inhibition observed using 2-5 ng TGFβ/ml. Immunodetectable P-450(AROM) decreased in parallel with activity following TGFβ treatment. The mechanism of TGFβ action was not through increasing phosphodiesterase (PDE) breakdown of cAMP since inhibition of PDE had no effect on TGFβ action. Finally we examined the level of P-450(AROM) mRNA using competitive polymerase chain reaction amplification. Bu₂cAMP increased mRNA levels of P-450(AROM) by 2.5-fold, while TGFβ inhibited this induction by 35%. The results of this investigation demonstrated that TGFβ is a potent regulator of P-450(AROM) expression in HF hepatocytes.