Induction of aryl hydrocarbon (benzo[a]pyrene) hydroxylase and 2-acetylaminofluorene N-hydroxylase by polycyclic hydrocarbons in regenerating liver from inbred strains of mice

The effect of partial hepatectomy (67–75 per cent excised) on aryl hydrocarbon (benzo[a]pyrene) hydroxylase and 2-acetylaminofluorene N-hydroxylase activities in genetically responsive C57BL/6N and nonresponsive DBA/2N inbred strains of mice was studied. Basal aryl hydrocarbon hydroxylase activity in C57BL/6N mice is reduced by 70 per cent within 12 hr but returns to control values between 36 and 48 hr after the operation and remains at that level thereafter. Similar, but less pronounced, changes are observed in the DBA/2N mouse. Treatment with 3-methylcholanthrene (80 mg kg^?1 intraperitoneally) 1 hr after partial hepatectomy induces aryl hydrocarbon hydroxylase in C57BL/6N mice, by 48 hr, to the same extent as in mice with intact livers. In DBA/2N mice, even after partial hepatectomy, treatment with 3-methylcholanthrene has no effect on aryl hydrocarbon hydroxylase activity. The effect of partial hepatectomy and 3-methylcholanthrene treatment on 2-acetylaminofluorene N-hydroxylase activity in both strains of mice is similar to that observed on aryl hydrocarbon hydroxylase activity.     

Tissue and sex differences in the activation of aromatic hydrocarbon hydroxylases in rats

Betamethasone and α-naphthoflavone produced similar activation of biphenyl 2-hydroxylase and benzo[a]pyrene 3-hydroxylase in control male rat liver microsomes. In small intestinal epithelial microsomes, betamethasone had no effect whereas α-naphthoflavone caused a pronounced activation of benzo[a]pyrene hydroxylation and a lesser activation of biphenyl 2-hydroxylation. In lung microsomes, betamethasone had no effect on either enzyme activity whereas α-naphthoflavone had no effect on biphenyl 2-hydroxylase but inhibited benzo[a]pyrene hydroxylase. In kidney cortex microsomes from male rats both compounds caused inhibition or had no effect whereas in kidney cortex microsomes from female rats betamethasone activated whereas α-naphthoflavone had no effect.Activation also occurred in isolated viable hepatocytes from male rats. The response of biphenyl 2-hydroxylase was very similar to that found in male rat liver microsomes but benzo[a]pyrene hydroxylase was more sensitive to activation and less sensitive to inhibition than in microsomes. The findings are interpreted as demonstrating the presence of more than one ‘latent’ aromatic hydrocarbon hydroxylase in rodents.     

Metabolic activation of Trp-P-2, a mutagenic amine from tryptophan-pyrolysate,by liver microsomes from 3-methylcholanthrene-responsive and non-responsive mice

1. The metabolic activation of a tryptophan pyrolysate, Trp-P-2 (3-amino-1-methyl-5H-pyrido[4,3-b]indole), was studied using liver microsomes from mice of 3-methylcholanthrene-responsive, C57BL/6N (B6) strain and non-responsive, DBA/2N (D2) strain.

2. The formation of N-hydroxy-Trp-P-2 (3-hydroxylamino-1-methyl-5H-pyrido-[4,3-b]indole) by hepatic microsomes was markedly increased by the pretreatment with 3-methylcholanthrene in B6 mice, but not in D2 mice.

3. The same treatment increased the activity to convert Trp-P-2 to a mutagen(s) in the Salmonella/microsome test system in B6 mice, but not in D2 mice.

4. The formation of N-hydroxy-Trp-P-2 corresponded with the increase in the number of the revertants, and with the activities of aromatic hydrocarbon hydroxylase and biphenyl 2-hydroxylase.

5. Addition of α-naphthoflavone to microsomes from control and 3-methyl-cholanthrene-treated B6 mice effectively decreased the activities to convert Trp-P-2 to a mutagen(s) and to N-hydroxylate Trp-P-2.

6. These results indicate that N-hydroxy-Trp-P-2 is a proximate or ultimate mutagenic principle of Trp-P-2.     

Depression of hepatic microsomal enzyme systems by lentinan in mice

Studies were performed to determine the effects of an immunopotentiating agent, lentinan, on the hepatic drug-metabolizing enzymes in mice. Lentinan was injected twice a day for two days, and the enzyme activities were determined 12 hr after the last injection of lentinan. A lentinan dose of over 0.25 mg/kg was required to cause a significant decrease (20-40%) in the hepatic microsomal aminopyrine N-demethylase and aniline hydroxylase activities. The loss of drug-metabolizing activity by the treatment with lentinan agreed with the loss of cytochrome P-450 content in many cases. Strain and substrate differences concerning the effect of lentinan on the metabolism of drug were also observed. That is to say, the loss of cytochrome P-450 content by the treatment with lentinan was observed in the ddY, C57BL/6 and BDF1 strain mice, but was not observed in the DBA/2, C3H/He and C57BL/10 strain mice. The decrease in the activities of 7-ethoxycoumarin O-deethylase and biphenyl 2-hydroxylase by the treatment with lentinan was considerably less than that of aminopyrine N-demethylase and aniline hydroxylase in ddY mice.     

Effects of pregnancy on the cytochrome P-450 system in mice

The effects of pregnancy on the hepatic cytochrome P-450-dependent mixed-function mono-oxygenase system (P-450) from day 6 to day 18 of gestation were examined in the C57BL/6J mouse. Pregnancy induced an initial increase and then a decrease in total P-450 content, a decrease in microsomal aminopyrine-N-demethylase activity, and had no effect on nucrosomal ethylmorphine-N-demethylase activity. Pregnancy also induced in the C57BL/6J and the DBA/2J mice a new major isozyme of P-450 (P-450_gest) as determined by high performance liquid chromatography and gel electrophoresis.     

Strain independent elevation of hepatic mono-oxygenase enzymes in female mice

1. Hepatic microsomal drug metabolizing activity was compared in male and female AL/N, Balb/c, Balb/cJ, CD-1, C57BL/6J, C57BL/10J, DBA/2, and DBA/2J mice. 2. Cytochrome P-450-dependent hexobarbital hydroxylase and aminopyrine N-demethylase activities were sexually dimorphic with apparent maximal velocities consistently higher in females. Hexobarbital-induced sleeping times were greater in males, corresponding to their lower hepatic mono-oxygenase enzyme activities measured in vitro. 3. No murine sex differences were observed for the hydroxylation of aniline, while UDP-glucuronyltransferase activity was strain dependent with either no sexual dimorphism or higher activities in males. 4. Testectomy resulted in an elevation of hepatic hexobarbital metabolism to female levels in all strains examined. Thus, decreased hepatic cytochrome P-450-dependent xenobiotic metabolism in adult male mice results from the suppressive effects of gonadal androgens. 5. Sexually dimorphic patterns of hepatic hexobarbital and aminopyrine metabolism in adult mice are opposite in orientation and lower in magnitude than the well established relationship in rats in which these same substrates are metabolized at a rate 3 to 5 fold higher in intact males as compared to females or gonadectomized males.     

Some characteristics of hamster liver and lung microsomal aryl hydrocarbon (biphenyl and benzo(a)pyrene) hydroxylation reactions

Aryl hydrocarbon hydroxylation (AHH) reactions were compared using liver and lung microsomes of corn oil- and 3-methylcholanthrene (3-MC)-treated hamsters, employing benzo(a)pyrene (BAP) and biphenyl as substrates. The predominant AHH activity of liver and lung microsomes from corn oil- or 3-MC-treated hamsters was biphenyl 4-hydroxylase. Biphenyl 2-hydroxylase and BAP-hydroxylase activities were approximately 50 per cent as active as biphenyl 4-hydroxylase in liver and approximately 1–3 per cent as active as biphenyl 4-hydroxylase in lung microsomes. Biphenyl 4-hydroxylase activity was 70–80 per cent as active in lung as in liver microsomes. Treatment with 3-MC in vivo induced the biphenyl 4-hydroxylation reaction in liver but not in lung microsomes, the biphenyl 2-hydroxylation reaction both in lung and liver microsomes, and the BAP hydroxylation reaction in lung but not in liver microsomes. Biphenyl 2- and 4-hydroxylase activities of liver microsomes displayed similar sensitivities to inhibition by a number of chemical inhibitors in vitro. Inhibition of biphenyl hydroxylation reactions by metyrapone or carbon monoxide did not distinguish between lung or liver microsomal mono-oxygenases of corn oil- or 3-MC-treated hamsters. While small differences were expressed by inhibition with ethylmorphine, large differences became apparent through inhibition studies with BAP or α-naphthoflavone. It is concluded that the major aromatic hydroxylase activity of lung microsomes from corn oil- or 3-MC-treated hamsters resembles the constitutive (uninduced) AHH of the liver microsomes and that the minor aromatic hydroxylase activity of lung microsomes from corn oil- or 3-MC-treated hamsters resembles the induced AHH of the liver microsomes.     

Regulation of hepatic monooxygenases by phenobarbital, 3-methylcholanthrene, and polychlorinated biphenyls in rapid and slow acetylator mice

A/J and C57BL/6J inbred mouse strains have been previously used as models of slow and fast acetylators, respectively, of human acetylator polymorphism. Studies were carried out to characterize possible differences in basal activities of hepatic monooxygenases and the response of these mouse strains to microsomal enzyme inducers. No significant difference in cytochrome P-450 content and associated enzyme activities of ethylmorphine N-demethylase and benzo(a)pyrene hydroxylase were observed between the two strains. The administration of the inducers, phenobarbital or the polychlorinated biphenyl mixture Aroclor 1254, resulted in significant increases in cytochrome P-450 and ethylmorphine N-demethylase activity and minimal changes in benzo(a)pyrene hydroxylase activity in both strains. Pretreatment with 3-methylcholanthrene resulted in little or no increase in N-demethylase activity in both strains. The polycyclic hydrocarbon caused a CO difference spectral shift to a lower wavelength only in the C57BL/6J mice. Further, it increased benzo(a)pyrene hydroxylase activity in both strains, but to a greater extent in the C57BL/6J strain. Electrophoretic studies using solubilized microsomal preparations confirmed the findings that the fast acetylators were highly responsive to the inducing properties of the polycyclic aromatic hydrocarbon, whereas the slow acetylators were relatively much less responsive to its inducing properties. The latter strain appeared to be more responsive to the inducing properties of the phenobarbital class of inducers, as reflected in the inducibility of cytochrome P-450 and the associated enzymic activities in the liver.     

The biochemical and genetic characteristics of murine ovarian aryl hydrocarbon (Benzo[a]pyrene) hydroxylase activity and its relationship to primordial oocyte destruction by polycyclic aromatic hydrocarbons

Primordial oocyte destruction by polycyclic aromatic hydrocarbons requires metabolic activation of the polycyclic hydrocarbon to an ovotoxic metabolite. The first and perhaps subsequent step(s) in the metabolism of polycyclic aromatic hydrocarbons to the proximate ovotoxin occurs via a microsomal cytochrome P-450-dependent monooxygenase. The role of aryl hydrocarbon (benzo[a]pyrene) hydroxylase (AHH, EC 1.14.14.2) in activation of polycyclic hydrocarbons to ovotoxic products was studied in 11 inbred and two F₁ heterozygote murine strains. The polycyclic aromatic hydrocarbon responsive C57BL/6N and C57BL/6J mice had two- to threefold increases in ovarian AHH activity after polycyclic hydrocarbon treatment. The polycyclic aromatic hydrocarbon nonresponsive DBA/2N and DBA/2J mice had no change in ovarian AHH activity after similar polycyclic hydrocarbon treatment. The polycyclic aromatic hydrocarbon responsive C57BL/6N and C57BL/6J mice had more rapid primordial oocyte destruction after polycyclic hydrocarbon treatment than the nonresponsive DBA/2N and DBA/2J mice. The (DBA/2N)(C57BL/6N)F₁ and (DBA/2J)(C57BL/6J)F₁ mice were responsive to polycyclic hydrocarbons with two- to threefold increases in ovarian AHH activity after treatment, however, the rate of oocyte destruction was similar to that observed in the nonresponsive DBA/2N and DBA/2J mice. Survey of seven additional inbred strains of mice failed to demonstrate a strong correlation between either absolute ovarian AHH activity or fold induction of ovarian AHH activity and oocyte destruction. Similarly, treatment with 2,3,7,8-tetrachlorodibenzo-p-dioxin, a potent inducer of ovarian AHH activity in both polycyclic hydrocarbon responsive and nonresponsive strains, did not alter the rate of oocyte destruction after treatment with benzo[a]pyrene or 3-methylcholanthrene. Although metabolic activation of polycyclic aromatic hydrocarbons to an ovotoxin is necessary for primordial oocyte destruction, strain differences in sensitivity to oocyte destruction reflect the biological sum of activation, detoxification, and repair. Therefore, differences in ovarian AHH activity or inducibility may not reflect differences in oocyte destruction after polycyclic hydrocarbon treatment.     

Benzo[a]pyrene metabolism in mouse liver. Association of both 7,8-epoxidation and covalent binding of a metabolite of the 7,8-diol with the Ah locus

The 7,8-epoxidation of benzo[a]pyrene, and the 9,10-epoxidation of benzo[a]-pyrene trans-7,8-dihydrodiol coupled with covalent binding of the highly reactive diol-epoxide, are two key P-450-mediated reactions believed to be important in cancer initiation, mutagenesis and teratogenesis. New assays for these two reactions were developed with mouse liver microsomes. These two activities have apparent Km values (approximately 6 microM) similar to that of aryl hydrocarbon hydroxylase activity. Twenty-six individual 3-methylcholanthrene-treated Ahb/Ahd and Ahd/Ahd progeny of the (C57BL/6N)(DBA/2N) F1 X DBA/2N backcross were studied. Both of the newly described activities appear to represent P-450 protein(s) that are responsible for aryl hydrocarbon hydroxylase activity and that are coordinately controlled by the Ahb allele.     

Induction of liver microsomal cytochrome P-450 And associated monooxygenases by octachlorostyrene in inbred strains of mice: Lack of correlation with the murinE Ah locus

Single intraperitoneal injections of octachlorostyrene (OCS) and hexachlorobenzene in genetically polycyclic aromatic hydrocarbon ‘responsive’ C57/BL/6 (B6) mice led to a time- and dose-dependent increase in the levels of liver microsomal cytochromes P-450 and b₅ as well as in the activities of NADPH cytochrome P-450 (cytochrome c) reductase, ethylmorphine (EM) N-demethylase, 4-nitroanisole (PNA) O-demethylase and acetanilide 4-hydroxylase (AcA hydroxylase). No, or only a very moderate, increase in the activity of aryl hydrocarbon hydroxylase was seen after OCS and HCB, respectively. Pretreatments with phenobarbital (PB) or 3-methylcholanthrene (MC) both increased AcA hydroxylase activity to a similar degree, whereas pretreatment with polychlorinated biphenyls (Aroclor 1254) had an effect equal to the sum of PB and MC. Judged from sodium dodecylsulfate polyacrylamide gel electrophoresis studies, OCS and HCB predominantly increased a microsomal polypeptide of apparent mol. wt 52,000, similar to PB. A reduced response was seen after OCS or HCB treatment of aromatic hydrocarbon ‘non-responsive’ DBA/2 (D2) mice compared to B6 mice, both with respect to AcA hydroxylase as well as EM demethylase and PNA demethylase activities. OCS treatment of B6D2F1 mice resulted in a doubling of AcA hydroxylase activity, but in mice of the (B6D2)D2 backcross no distinct subgroupings of individual AcA hydroxylase activities were apparent. These results demonstrate that OCS is an inducer of the PB-type in mice and that induction of AcA hydroxylase by OCS is not regulated by the Ah locus.     

Strain independent elevation of hepatic mono-oxygenase enzymes in female mice

1. Hepatic microsomal drug metabolizing activity was compared in male and female AL/N, Balb c, Balb/cJ, CD-1, C57BL/6J, C57BL/10J, DBA/2, and DBA/2J mice.

2. Cytochrome P-450-dependent hexobarbital hydroxylase and aminopyrine N-demethylase activities were sexually dimorphic with apparent maximal velocities consistently higher in females. Hexobarbital-induced sleeping times were greater in males, corresponding to their lower hepatic mono-oxygenase enzyme activities measured in vitro.

3. No murine sex differences were observed for the hydroxylation of aniline, while UDP-glucuronyltransferase activity was strain dependent with either no sexual dimorphism or higher activities in males.

4. Testectomy resulted in an elevation of hepatic hexobarbital metabolism to female levels in all strains examined. Thus, decreased hepatic cytochrome P-450-dependent xenobiotic metabolism in adult male mice results from the suppressive effects of gonadal androgens.

5. Sexually dimorphic patterns of hepatic hexobarbital and aminopyrine metabolism in adult mice are opposite in orientation and lower in magnitude than the well established relationship in rats in which these same substrates are metabolized at a rate 3 to 5 fold higher in intact males as compared to females or gonadectomized males.     

Genetic differences in the induction of aryl hydrocarbon hydroxylase and its components by 3-methylcholanthrene in liver and lung microsomes among four strains of guinea pigs

Four strains of guinea pigs (Hartley, No. 2, No. 13 and JY-1) were examined for the effects of intraperitoneal treatment with 3-methylcholanthrene on aryl hydrocarbon hydroxylase activity, total cytochrome P-450 content in liver and lung microsomes, and NADPH-cytochrome c reductase activity in liver microsomes. Following treatment with 3-methylcholanthrene at a dose of 50 mg/kg body weight, aryl hydrocarbon hydroxylase activity and cytochrome P-450 content in liver were both increased in all the strains used, and the activity of NADPH-cytochrome c reductase in liver was also increased in all strains except No. 13. While the cytochrome P-450 content in lung was increased in all the strains except No. 13, there was no increase in the aryl hydrocarbon hydroxylase activity in lung from any strain of guinea pig examined. When the dose of 3-methylcholanthrene was increased to 250 mg/kg body weight, an apparent induction of aryl hydrocarbon hydroxylase was detected in the lung from the Hartley strain of guinea pigs, but not in the other three strains. In summary, marked differences were seen in sensitivity to 3-methylcholanthrene between liver and lung, and apparent strain differences were observed among the guinea pigs used in this experiment.     

Genetic differences between C57BL/6 and DBA/2 mice in the inductions of UDP-glucuronyl transferases for 3-hydroxybenzo(a)pyrene,p-nitrophenol,and bilirubin by 3-methylcholanthrene

In C57BL/6 mice, aryl hydrocarbon hydroxylase (AHH) increased 1 day after treatment with 3-methylcholanthrene, and induction of UDP-glucuronyl transferases for 3-hydroxybenzo(a)pyrene, p-nitrophenol and bilirubin in the liver microsomes was observed 2 to 5 days later. In DBA/2 mice, neither AHH nor transferase activities were influenced by 3-methylcholanthrene. These results suggest that induction of activating and detoxicating enzymes is genetically linked.     

Induction of hepatic mono-oxygenase systems in fetal and neonatal rats with phenobarbital, polycylic hydrocarbons and other xenobiotics.

The induction of hepatic P-450 hemoprotein-dependent mono-oxygenase systems was studied in fetal and neonatal rats. The fetal liver was refractive to phenobarbital induction of aminopyrine and ethylmorphine N-demethylase. which are cytochrome P-450-dependent mono-oxygenases, but was not refractive to the 3-methylcholanthrene induction of benzo[a]pyrene hydroxylase. a cytochrome P₁-450-dependent mono-oxygenase. After parturition, all three enzyme activities were inducible. These and other observations suggest that a control mechanism operates in the fetal rat which selectively suppresses the induction of cytochrome P-450. but allows induction of cytochrome P₁-450. This selective suppression of phenobarbital induction in the fetus was reversed in part by the simultaneous administration of 3-methylcholanthrene. Several other inducing agents also partially reversed the suppression of phonobarbital induction in Fetal livers: dibenz-[a,c]anthracene, 2-diethylaminoethyl-2.2-diphenyl-valerate (SKF 525-A), and 3β-hydroxy-20-oxopregn-5-ene-16α-carbonitrile (PCN). Other inducing agents were inactive: α-naphthoflavone, β-naphthoflavone. 1,1-bis[p-chlorophenyl]2,2,2-trichloroethane (DDT). 2,4,5-trichlorophenoxyacetic acid (2,4,5-T). dieldrin. chlordane and chlorpromazine.     

Hemopexin and aryl hydrocarbon hydroxylase induction in the mouse and in cell culture

Plasma hemopexin levels in mice treated with polycyclic aromatic compounds such as 3-methylcholanthrene or β-naphthoflavone are increased 71–160 per cent in three inbred strains known to be “responsive” at the Ah locus (C57BL/6N. C3H/HeN and Balb/cAnN) and ranged from 14 to 60 per cent in three inbred strains known to be “nonresponsive” at the Ah locus (DBA/2N, NZB/BLN and NZW/BLN). Phenobarbital treatment causes increases in plasma hemopexin of similar magnitude in both aromatic hydrocarbon responsive and nonresponsive mice. In F₂ progeny from crosses between the C57BL/6N and NZW/BLN progenitor strains, a correlation between inducible hepatic hydroxylase and plasma hemopexin concentrations could not be demonstrated, principally because of the small magnitude in hemopexin induction. Data in this study show that plasma hemopexin rises later than the initiation of the aryl hydrocarbon hydroxylase induction process and, therefore, presumably in response to increased heme synthesis—whether it is the genetically regulated hemoprotein P₁-450 induction by polycyclic aromatic compounds or hemoprotein P450 induction by pheno-barbital. An association between hemopexin and hydroxylase induction is not seen in H-4-II-E Reuber rat hepatoma cultures. Hemopexin synthesis and secretion into the growth medium occurs in H-4-II-E cells but is not detectable in HTC or Hepa-1 cultures. The rate of hemopexin appearance in the culture medium is the same in bcnz[a]anthracene-treated as in control H-4-II-E cells, whereas the hydroxylase induction occurs primarily in benz[a]anthracene-treated and not in control cultures. Daily changes in growth medium result in greater intracellular hemopexin levels and a greater rate of hemopexin secretion into the medium, compared with cells whose medium was changed less frequently or not at all.     

Effects of vitamin A deficiency on hepatic and extrahepatic mixed-function oxidase and epoxide-metabolizing enzymes in guinea pig and rabbit.

Male guinea pigs and male rabbits were fed a vitamin A deficient diet for 9 weeks and for 12 weeks respectively. Hepatic levels of vitamin A were significantly reduced in the vitamin A deficient animals. The activities of some xenobiotic-metabolizing enzymes were measured in the liver, lung and small intestine. Aryl hydrocarbon hydroxylase, aniline hydroxylase, and 7-ethoxycoumarin deethylase activities were decreased in the vitamin A deficient guinea pig liver. However, in the guinea pig small intestine, aniline hydroxylase, 7-ethoxycoumarin deethylase, aminopyrine demethylase, and aryl hydrocarbon hydroxylase specific activities were increased. In rabbits, vitamin A deficiency decreased hepatic aniline hydroxylase and 7-ethoxycoumarin deethylase activities but increased intestinal aminopyrine demethylase activity. Enzyme activities in lung were not altered by vitamin A deficiency in guinea pig or rabbit. Microsomal epoxide hydrase and microsomal supernatant glutathione S-transferase activities in the three tissues of both species were not altered by vitamin A deficiency.     

Effect of hexachlorobenzene on microsomal enzyme systems

Male rats were fed for 10 days on a diet containing 333 ppm hexachlorobenzene. Increased microsomal protein levels were noted compared to control rats. On a per g liver basis, the levels of aniline hydroxylase, biphenyl 4-hydroxylase, biphenyl 2-hydroxylase, 4-nitroanisole O-demethylase, esterase, cytochrome P-450 and cytochrome b₅ all increased compared with the control values. On a per mg microsomal protein basis, biphenyl 2-hydroxylase, 4-nitroanisole O-demethylase and cytochrome P-450 levels increased several-fold compared with the control values. It is suggested that, by inducing the 2-hydroxylation reaction, hexachlorobenzene might cause preferential ortho-hydroxylation, as do some carcinogenic polycyclic hydrocarbons, and that in some circumstances this could lead to the formation of carcinogens.     

Lack of correlation between formation of reactive metabolites and thymic atrophy caused by 3, 4, 3′, 4′-tetrachlorobiphenyl in C57BL/6N mice

The possible role of active metabolites of 3, 4, 3, 4-tetrachlorobiphenyl (TCB) in causing thymic atrophy was investigated using inbred strains of mice. The generation of reactive species which bind covalently to cellular proteins was used to monitor the formation of active TCB metabolites. The amount of in vitro covalent binding of TCB to proteins by liver microsomes was increased markedly by pretreatment of AHH-responsive C57BL/6N mice with either 3-methylcholanthrene (MC) or TCB itself, although these two inducers were not effective in AHH-nonresponsive DBA/2N mice. MC treatment also caused an induction of microsomal TCB-binding activity in all of the (C57BL/6N x DBA/2N) F₁ mice. Moreover, among 38 individuals of [(C57BL/6N) (DBA/2N) F₁ x DBA/2N] backcross, 23 mice responded to MC with respect to microsomal TCB-binding activity while others did not. These results suggest that the conversion of TCB to protein-bound metabolites is mediated by particular form(s) of cytochrome P-450 which is (are) induced by an Ah receptor mechanism. In order to ascertain wheter the active TCB metabolites play a role in causing thymic atrophy, ¹⁴C-labeled TCB was administered IP to C57BL/6N mice and the amount of covalent binding of radioactive metabolites to tissue proteins was determined. The in vivo binding was evident in the liver, particularly in the microsomal fraction, on the basis of protein content. In contrast, the thymic proteins contained no measurable amounts of bound radioactivity even when the mice showed marked thymic atrophy. These data suggest that thymic atrophy caused by TCB is not likely to result from the generation of reactive metabolites.Abbreviations Used TCB 3, 4, 3, 4-tetrachlorobiphenyl - PCB polychlorinated biphenyl(s) - TCDD 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin - MC 3-methylcholanthrene - PB phenobarbital - AHH aryl hydrocarbon hydroxylase     

Time and dose responses of the reduction in retinoid concentrations in C57BL/Rij and DBA/2 mice induced by 3,4,3',4'-tetrachlorobiphenyl

A single ip dose of 15 mg 3,4,3',4'-tetrachlorobiphenyl (TCB)/kg induced a 30 to 40% reduction of retinol and retinyl palmitate concentrations in hepatic tissue of C57BL/Rij mice within 2 to 4 days. This level of reduction was maintained for about 14 days. The ED50 was 32 mg TCB/kg for hepatic retinol and 17 mg TCB/kg for hepatic retinyl palmitate. In DBA/2 mice, however, no reduction in hepatic retinoids was observed even at doses up to 729 mg TCB/kg. The duration of the reduction in hepatic retinoids did not correlate with the induced aryl hydrocarbon hydroxylase (AHH) activity in C57BL/Rij mice. These data suggest that AHH and related enzymes are not directly involved in the TCB-induced reduction in retinoids in these mouse strains. No significant differences in the accumulation of TCB in hepatic tissues of C57BL/Rij and DBA/2 mice were observed; however, the estimated elimination rate in C57BL/Rij was two times faster than that in DBA/2 mice. In serum of DBA/2 mice, a rapid 50% decline in the concentration of retinol was observed after only 2 to 6 hr, remaining at the reduced value for about 14 days. The ED50 for serum retinol was 1 to 2 mg TCB/kg. Only a transient reduction in serum retinol, disappearing within 48 hr, was observed in the C57BL/Rij mouse. TCB accumulated to an almost 15-fold higher extent in serum of DBA/2 mice as compared with C57BL/Rij mice. The effect of TCB on retinoids in a few other strains of mice and in the Sprague-Dawley rat showed a reduction in serum retinol rather than in hepatic retinoids.