Influence of a novel hypophyseal factor on steroid metabolism in cultured hepatoma cells.

Abstract

A rat hepatoma cell line in tissue culture (HTC cells) was treated with hypophyseal extracts from adult male and female rats. Cell homogenates were then assayed for-steroid metabolizing enzymes using 4-androstene-3,17-dione as substrate. The major products were the 5a-reduced derivatives (5a-androstane-3,17-dione, androsterone, and epiandrosterone). When the cells were grown in the presence of female hypophyseal extract the apparent activity of the 5a-reductase increased markedly, whereas treatment with male hypophyseal extract was without effect. Treatment with female hypophyseal extract resulted in a marked decrease in the apparent Km for 5a-reductase from 667 & 120 to 99 + 4 jsM in addition to a decrease in the apparent Vmax from 67 i 12 to 46 + 2 pmol of product/min per mg of protein. A logarithmic dose-response was obtained with female hypophyseal extract. Treatment of the HTC cells with purified rat hypophyseal follicle-stimulating hormone, luteinizing hormone, growth hormone, thyrotropic hormone, and prolActin had only marginal effects on 5a-reductase activity. Crude female hypophyseal extracts were at least 6-fold more potent than any of the standard hormone preparations and at least 250-fold more potent than male hypophyseal extracts when based on activity per mg of pituitary tissue. Chromatography of crude female hypophyseal extracts on Sephadex G-25 indicated that the factor was of high molecular weight. The identity of this activity with a hypophyseal "feminizing" factor is postulated.Sexual differences in liver metabolism of steroid hormones in rats have been known for a long time (1-4). Recently, evidence has been presented that the sexual differentiation of liver steroid metabolism is controlled by the hypothalamicohypophyseal system (5). This control seems to be exerted via a hypophyseal "feminizing" factor, and most of the effects of estrogens and androgens on liver metabolism are probably mediated by changes in the secretion of this factor (5). Experiments where the effect of prolactin, follicle-stimulating hormone (FSH), and luteinizing hormone (LH) on liver enzyme activities has been studied give no indication that these hormones are involved in the feminization of the hepatic enzyme pattern (6, 7).The regulation of hepatic steroid metabolism by the pituitary is important since it indicates the possible existence of a new hypophyseal hormone. The present paper describes the effect of a factor (or factors), present in the adult female rat pituitary, on tissue cultured hepatoma cells, which is similar to that of the "feminizing" factor on rat liver in vivo (5). The HTC cells thus seem to provide a potential tool for the isolation and characterization of such a factor. MATERIALS AND METHODS Steroids. 4-[4-14C]Androstene-3,17-dione was obtained from the Radiochemical Centre (Amersham, England) and purified by thin-layer chromatography. Unlabeled 4-androstene-3,17-dione, 3a-hydroxy-5a-androstan-17-one (androsterone), and 3,3-hydroxy-5a-androstan-17-one (epiandrosterone) were kindly donated...

Hypothalamic‐pituitary‐gonadal Relationships in Man From Birth to Puberty

“…In contrast, however, the neonatal administration of a large dose of oestrogens produces masculinization of the hypothalamus and, in adult life, the development of the permanent oestrus syndrome (Gorski, 1971). A similar permanent 'programming' action of testosterone secreted at birth appears to occur in the hypothalamus, and will reveal itself in the skin and the liver enzyme activities in adulthood (De Moor et al, 1973;Gustafsson et al, 1975b).…”

Effect of the anterior pituitary gland and gonads on enzyme components of the hepatic microsomal cytochrome P-450 system of male and female rats

“…Both growth hormone (GH) and prolactin have been proposed as the feminizing hormone (Lax et al 1976;Rumbaugh & Colby, 1980;Mode, Norstedt, Simic, Eneroth & Gustafsson, 1981). It may, however, be neither of these but an as yet uncharacterized hormone (Gustafsson, Larsson, Skett & Stenberg, 1975;Skett, Eneroth, Gustafsson, Sonnenschein, Stenberg & Ahlen, 1978;). In this paper we have studied the role of the anterior pituitary gland in the maintenance of the sex difference in the concentration of cytochrome P-450 in rat liver microsomes.…”