Sex hormone-binding globulin (SHBG) is a glycoprotein whose production has been demonstrated to be regulated by both sex steroids, as well as by thyroid hormone and peptide hormones such as insulin. However, none of these regulatory factors would explain the marked decrease in serum SHBG seen throughout the prepubertal and pubertal time period in both boys and girls. Furthermore, current in vitro data show that both androgens and estrogens can stimulate SHBG production by the human hepatoblastoma cell line Hep G2; yet, in vivo androgens appear to suppress SHBG levels, while estrogens are associated with elevated levels. This study was undertaken to determine possible mechanisms to explain this phenomenon. Hep G2 cell cultures were incubated with insulin-like growth factor I (IGF-I), epidermal growth factor (EGF), transforming growth factor alpha (TGF-alpha), or dehydroepiandrosterone (DHEA). Significant decreases in the level of SHBG in the culture medium relative to control cultures occurred for each of the growth factors (P less than .01), whereas an increase in SHBG levels was observed in the medium of DHEA-treated cells. When cells were coincubated with IGF-I and thyroxine (T4), which alone stimulates SHBG production both in vivo and in vitro, the SHBG response to T4 was blunted. These results suggest that growth factors, as well as insulin, may be important determinants in SHBG production.