Cell, Vol. 7. 413-417, March 1976, Copyright:91976 by MIT Feminization of Hepatic S Metabolism in Male Rats with a Transplanted (MtT/F4) Peter Eneroth’, Jan-Ake Gustafsson+, Agne Larsson*, Paul Skett+, and Ake Stenberg+ Carlos Sonnenschein Tufts University School of Medicine Cancer Research Center Boston, Massachusetts 02111. Summary After transplantation of MtT/F4 pituitary tumor cells to male rats of the Fisher strain, the masculine type of hepatic steroid metabolism was changed into a feminine pattern of enzyme activities. Liver me- tabolism of steroid hormones in female rats was relatively unaffected following transplantation of pituitary tumor cells. Furthermore, extract from MtT/F4 tumors and “autonomous” pituitary tissue increased the Sa-reductase activity of hepatoma cells in the culture (HTC cells) at subsaturation concentrations of the substrate 4-androstene-3, 17-dione by decreasing the apparent K, of the en- zyme. It is concluded that the pituitary tumor (in accord with the secretion from an “autonomous” pituitary gland) secretes “feminotropin,” a novel hypophyseal principle that probably is an impor- tant regulator of hepatic steroid metabolism. It is suggested that pituitary tumor tissue of the MtT/F., type could be used as source of feminotropin in purification studies. Introduction Recent findings have indicated that the hypothala- mica-pituitary axis exerts a regulatory control over hepatic steroid metabolism in rats (Gustafsson and Stenberg, 1974a; 1975). At least part of this control is accomplished through the secretion in vivo of a factor (termed “feminotropin” or “feminizing fac- tor”) from the female rat pituitary that is responsible for the well known sexual differences characteristic of hepatic steroid metabolism in rats (Gustafsson and Stenberg, 1974a, 1975; Gustafsson et al., 1975). In the female liver, the apparent 5cu-reduc- tase activity is considerably greater than in the male liver (Einarsson, Gustafsson, and Stenberg, 1973; Gustafsson and Stenberg, 1974b), and extracts from rat pituitaries stimulate this activity in cultured hepatoma cells (Gustafsson et al., 1975). The inac- tivity of the male pituitary in vivo seems to be due to an inhibitory influence from the hypothalamus, since hypophysectomized male rats develop a femi- nine pattern of liver metabolism upon transplanta- “HarmonJaboratoriet. Karolinska Hospital: ‘Department of Chemistry, Karolinska Institutet; :Department of Pediatrics, St. Gdrans Children’s Hospital Stockholm, Sweden tion of a pituitary under the kidney capsule (Gustafsson and Stenberg, 1975). The concept of hypothalamico-pituitary regulation of hepatic steroid metabolism is supported by our investiga- tion, where we have studied the effects of a trans- planted pituitary tumor, MtT/F4 (Furth et al., 1956), on liver metabolism of steroids in rats of both sexes, and the effect of tumor extract and extract from “autonomous” pituitaries on the steroid metabo- lism of HTC cells. We conclude that this tumor con- tains and secretes feminotropin and suggest that the tumor may be an appropriate starting material for the purification of the factor. Results The metabolites formed after incubation of liver microsomal and supernatant fractions with 4-4- 14C-androstene-3, 17-dione and 5n-4-‘4C-andros- tane-3a, 17/I-diol have been identified in previous publications (Berg and Gustafsson, 1973; Einars- son et al., 1973; Gustafsson and Stenberg, 1974b). By measuring the formation of the products, it was possible to assay the following enzyme activities: the 5a- and 5/3-reduced metabolites, respectively, 3/3- and 17,&hydroxysteroid reductases, and 6p-, 7n-, and 16a-hydroxylases active on 4-andros- tene-3, 17-dione and the 2a-, 2p-, 7a-, 7p-, and 18-hydroxylases active on 5a-androstane-3a, 17p- dial. As can be seen from Figure 1 and Table 1, the presence of a pituitary tumor transplant had highly significant effects on the enzyme activities in male rat liver. The metabolism of 4-androstene-3, 17- dione underwent the following changes: the appar- ent 5a-reductase activity increased more than 3 times, whereas the 5p-, 17p-, and 3/3-reductase activities decreased by about 55, 60, and 80%, re- spectively; the S/3- and 7u-hydroxylase activities decreased by about 60 and 50%, respectively; and the 16a-hydroxylase activity decreased to a value below the detection level at the substrate concen- tration used. Even more pronounced changes were observed in the hepatic metabolism of Sn-andros- tane-3a, 17/3-diol in male tumor-bearing rats: the activities of 2~ and 18-hydroxylase were reduced by about 90%, and the activities of 2/3- and 7P-hy- droxylase were reduced to values below the detec- tion level. The only exception was the 7a-hydroxy- lase activity, which was not significantly changed. In contrast to these results, the hepatic steroid me- tabolism of female rats was only slightly affected by the presence of the tumor. 5a reduction of 4- androstene-3, 17-dione decreased by about 30%; 17/3-hydroxysteroid reduction increased by about 50%, and G,&hydroxylation increased by about 15%. No differences were seen in hydroxylation of