Ingestion of soil contaminated with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) alters hepatic enzyme activities in rats.

Female rats were treated with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in either corn oil or contaminated soil from the Minker site in Missouri. Eight doses ranging from 0.015 to 5 micrograms TCDD/kg were used in the corn oil group; the range was 0.015 to 5.5 micrograms TCDD/kg in the TCDD-contaminated soil group. Rats in a third group were given equal amounts of soil uncontaminated with TCDD. No acute toxicity or effects on body weight gain were observed at these doses. In general, equivalent doses of TCDD in corn oil or TCDD in soil produced similar increases in hepatic aryl hydrocarbon hydroxylase activity (AHH) and UDP glucuronyltransferase activity although effects were slightly greater in the TCDD-corn oil groups. In the corn oil groups, the induction of AHH ranged from about 30-fold at the highest dose to twofold at the lowest dose studied. TCDD also caused an increase in cytochrome P-450 concentration and a shift in spectral peak from 450 to 448 nm. There was no effect of TCDD on ethylmorphine N-demethylase, consistent with previous reports. Liver concentrations of TCDD (mean +/- SD) in the 5-micrograms/kg groups were 40.8 +/- 6.3 ppb in the TCDD corn oil group and 20.3 +/- 12.9 ppb in the TCDD-contaminated soil group. Our results suggest that the bioavailability of TCDD in soil in rats is approximately 50%. Therefore, ingestional exposure to TCDD-contaminated soil may constitute a significant health hazard in view of its extremely high toxicity and relatively high bioavailability.

Correlation of hepatic cytosolic androgen binding proteins with androgen induction of hepatic microsomal ethylmorphine N-demethylase in the rat.

Previous reports have demonstrated the presence of moderate to high affinity binding for androgens in the cytosol of livers from male rats. This binding was significantly lower in female rats or in immature rats of either sex. The hepatic androgen binding protein, which sedimented at approx. 4 S on sucrose density gradients, has been called a receptor which mediates the actions of androgens in the liver. The experiments in the present study were designed to evaluate the hepatic androgen binding protein for characteristics which have been attributed to receptors in other tissues and to correlate the presence of androgen binding with androgen induction of hepatic drug metabolism. In the current studies, we have shown that cytosol from the livers of male rats bound [3H]dihydrotestosterone [( 3H]DHT) and translocated this steroid ligand to the nucleus in a time and temperature dependent manner. Cytosol prelabeled with [3H]DHT, when passed over a column of denatured DNA cellulose, eluted in three radioactive peaks. Two of these peaks were absent when cytosol from livers of female or hypophysectomized males was used. In addition, the presence of high concentrations of hepatic androgen binding correlated well with the ability of androgen to induce ethylmorphine N-demethylase, a marker of microsomal cytochrome P-450-dependent drug metabolism. Values for both parameters were higher in males than in either females or hypophysectomized males. Testosterone treatment induced both parameters in ovariectomized females and 17 beta-estradiol repressed both in males. However, testosterone treatment failed to induce hepatic androgen binding in hypophysectomized males and immature males, both of which are also unresponsive to androgen induction of drug metabolism. The results suggest that one or more hepatic cytosolic androgen binding proteins possess several characteristics associated with steroid receptors in reproductive tract tissue. Furthermore, this binding may be implicated as a mediator for the androgen induction of at least one component of hepatic drug metabolism.

Dioxin in soil: bioavailability after ingestion by rats and guinea pigs.

Soil environmentally contaminated with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) was given by gavage to guinea pigs and rats. The development of a characteristic clinicopathologic syndrome in guinea pigs, the induction of aryl hydrocarbon hydroxylase in rats, and the presence of TCDD in the livers of both species show that TCDD in soil exhibits high biological availability after ingestion.

Imprinting of hepatic estrogen-binding proteins by neonatal androgens.

Previous studies discovered a second class of estrogen-binding proteins distinct from estrogen receptor which exhibited higher capacity, lower affinity (HCLA) binding properties. HCLA sites underwent postpubertal sex differentiation, such that adult male levels were at least 10-fold higher than adult female levels. Neonatal castration of male rats prevented the subsequent sex differentiation of HCLA binding sites; adult male rats that were castrated neonatally exhibited typically female concentrations of these binding sites. If male rats were castrated at 19 days of age or later, postpubertal sex differentiation of HCLA binding sites proceeded as observed for intact males. Administration of testosterone propionate (TP) to castrated (neonatally) males during a critical period (days 6-13) imprinted for the subsequent sex differentiation of HCLA sites, whereas TP administration at other times did not. The expression of these imprinted sites was not manifested until puberty, as neonatal manipulation or TP treatment had no effect on HCLA sites in immature rats. The imprinted effect on HCLA binding sites appeared to be permanent and irreversible. Treatment of castrate males with diethylstilbestrol (DES) or zearalenol (P-1496) during the critical period was incapable of restoring development of normal male levels of HCLA sites. Competitive binding studies using several steroid hormones revealed similar data for intact males and castrate (neonatal) male rats that had received TP during the critical period. These studies demonstrate an early imprinting period during which androgen exposure programs for postpubertal development of sex differentiation of HCLA binding sites.

Influences of sex steroids on experimental carcinogenesis.

Sex steroids can apparently modify hepatic function in at least two ways (Figure 1). First, the liver contains specific receptor proteins for either androgens or oestrogens. Several studies, using effects on the monooxygenase system as an indicator of hormone action, have shown good correlations between the presence of sex-steroid receptors and hepatic response to these hormones. In general, androgens induce the activities of drug biotransformation reactions, whereas oestrogens depress them. Second, the endocrine milieu, during a critical period of early development, apparently programmes the hypothalamic-pituitary axis to release factors which cause certain aspects of liver biochemistry to undergo sex differentiation. Selected components of the cytochrome P-450 system exhibit sex differences, and these differences are imprinted by neonatal androgens and are initiated and maintained by the pituitary gland. Corresponding to sex differentiation of the hepatic monooxygenase system are significant sex differences in responses to hepatocarcinogens such as N-2-fluorenylacetamide and N-hydroxy-N-2-fluorenylacetamide. Higher incidences of hepatocellular carcinomas in males appear to result from exposure to neonatal androgens.

Changes in adrenal microsomal cytochrome(s) P-450 with aging in the guinea pig.

Studies were carried out to investigate the mechanism(s) responsible for the changes in adrenal microsomal mixed function oxidase activity which occur with aging (30-200 days) in guinea pigs. With aging, the rate os metabolism of xenobiotics [ethylmorphine and benzo(a)pyrene] by adrenal microsomes increased 3- to 5-fold. Steroid 17 alpha- and 21-hydroxylations, when expressed per mg protein, were similar in immature (30 days old) and mature (200 days old) animals. Adrenal microsomal NADPH- and NADH-cytochrome c reductase activities and cytochrome b5 concentrations increased wih aging, but cytochrome P-450 concentrations were not significantly different in young and old guinea pigs. Maximal type I difference spectra produced by steroids were the same in adrenal microsomes from 30- and 200-day-old guinea pigs, but the ethylmorphine-induced spectrum was far greater in the older animals. Progesterone enhanced NADPH-cytochrome P-450 reductase activity to about the same extent in adrenal microsomes from 30- and 200-day-old guinea pigs. Ethylmorphine had no effect on the rate of reduction of cytochrome P-450 in adrenals from young animals but produced a 4-fold increase in activity in adrenals from older animals. The results demonstrate selective changes in adrenal xenobiotic metabolism with aging and suggest that changes in the composition and/or reactivity of adrenal cytochromes P-450 are responsible for the effects of aging.

Is growth hormone the pituitary feminizing factor mediating the actions of estradiol on hepatic drug and steroid metabolism?

Previous studies have established that the effects of estradiol (E2) on hepatic steroid and drug metabolism are demonstrable only in the presence of the pituitary gland. Studies were carried out to test the hypothesis that GH is the pituitary feminizing factor mediating the actions of E2 on hepatic metabolism. E2 and GH administered to castrated male rats had similar effects on hepatic enzymes, decreasing the oxidataive metabolism of drugs [ethylmorphine demethylation, aniline, hydroxylation, and benzo(a)pyrene hydroxylation) and increasing steroid (corticosterone) delta 4-hydrogenase activity. None of these effects of E2 or GH could be demonstrated in hypophysectomized (hypox) rats. However, GH administration to T4- or ACTH-treated hypox rats resulted in some of the changes in drug and steroid metabolism seen in animals with intact pituitary glands. The actions of GH on hepatic microsomal enzymes were fully demonstrable in hypox rats receiving both T4 and ACTH. E2 had no effects in T4 plus ACTH-treated hypox rats. These and prior observations are consistent with the hypothesis that GH mediates the actions of E2 on hepatic microsomal drug- and steroid-metabolizing enzymes. The data also indicates that the cations of GH on hepatic metabolism are dependent upon the interactions with still other endocrine factors.

Strain differences in adrenal xenobiotic metabolism in guinea pigs.

Studies were carried out to compare adrenal and hepatic xenobiotic metabolism in various strains of guinea pigs. In all strains studied (Hartley, English Short Hair, NIH, Strain 2, Strain 13), microsomal protein and cytochrome P-450 levels and NADPH-cytochrome c reductase activities were greater in adrenals than livers. Neither adrenal values nor hepatic values for these parameters differed across strains. Ethylmorphine (EM) demethylase and benzo[a]pyrene (BP) hydroxylase activities were also greater in adrenals than livers in all strains. However, the rates of adrenal xenobiotic metabolism were far greater in the highly inbred Strain 2 and Strain 13 guinea pigs than in other strains. In contrast, hepatic metabolism of EM and BP was not strain-dependent. Adrenal steroid 21-hydroxylase activity was also similar in all strains. The results indicate that strain is an important determinant of adrenal but not hepatic xenobiotic metabolism in the guinea pig. In addition, genetic control of adrenal microsomal drug and steroid metabolism appear to be independent of one another.

Maturational changes in adrenal xenobiotic metabolism in male and female guinea pigs.

In young (25-day-old) guinea pigs, adrenal and hepatic benzo[a]pyrene (BP) hydroxylase activities were similar but the rates of ethylmorphine (EM) demethylation were greater in adrenals than liver. Between 25 and 50 days of age no sex differences in adrenal or hepatic enzyme activities were demonstrable. The rates of adrenal BP and EM metabolism increased with age in guinea pigs of both sexes; activities reached significantly higher levels in males than females. In contrast, hepatic metabolism of BP and EM declined with maturation and activities were similar in males and females at all ages. Neither microsomal cytochrome P-450 concentrations no NADPH-cytochrome c reductase activities correlated with the maturational changes or sex differences in xenobiotic metabolism. Adrenal microsomal steroid 21-hydroxylase activity did not change significantly with aging and was not sex-dependent. The results indicate that opposite changes occur in adrenal and hepatic xenobiotic metabolism as a function of aging, resulting in substantially greater adrenal than hepatic activity in sexually mature animals. The data also suggest that adrenal microsomal drug and steroid metabolism are independently regulated.

Requirement of the pituitary gland for gonadal hormone effects on hepatic drug metabolism in rats.

Previous studies have demonstrated an important role for the pituitary gland in the regulation of hepatic drug and steroid metabolism. The present studies were carried out to determine the relationship between the pituitary gland and the actions of gonadal hormones on hepatic drug metabolism in rats. Testosterone administration to castrated male rats increased hepatic microsomal cytochrome P-450 concentrations and enhanced the rates of ethylmorphine demethylation and benzo(a)pyrene hydroxylation. Estradiol treatment, on the other hand, lowered cytochrome P-450 levels and decreased the rates of aniline, ethylmorphine and benzo(a)pyrene metabolism in orchiectomized rats. However, when given to hypophysectomized male rats, neither testosterone nor estradiol affected cytochrome P-450 levels or enzyme activities. Similarly, 5alpha-dihydrotestosterone administration increased hepatic drug metabolism only in the presence of the pituitary gland. The effects of both testosterone and estradiol were fully demonstrable in the absence of the thyroid or adrenal glands, excluding the need for either organ. The results indicate an absolute dependence on the pituitary gland for gonadal hormone (testosterone and estradiol) actions on hepatic mixed-function oxidases.