Transgenic mouse lines expressing rat AH receptor variants — A new animal model for research on AH receptor function and dioxin toxicity mechanisms

Han/Wistar (Kuopio; H/W) rats are exceptionally resistant to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) toxicity mainly because of their mutated aryl hydrocarbon receptor (AHR) gene. In H/W rats, altered splicing of the AHR mRNA generates two AHR proteins: deletion (DEL) and insertion (INS) variants, with the INS isoform being predominantly expressed. To gain further insight into their functional properties, cDNAs of these and rat wild-type (rWT) isoform were transferred into C57BL/6J-derived mice by microinjection. The endogenous mouse AHR was eliminated by selective crossing with Ahr-null mice. A single mouse line was obtained for each of the three constructs. The AHR mRNA levels in tissues were generally close to those of C57BL/6 mice in INS and DEL mice and somewhat higher in rWT mice; in testis, however, all 3 constructs exhibited marked overexpression. The transgenic mouse lines were phenotypically normal except for increased testis weight. Induction of drug-metabolizing enzymes by TCDD occurred similarly to that in C57BL/6 mice, but there tended to be a correlation with AHR concentrations, especially in testis. In contrast to C57BL/6 mice, the transgenics did not display any major gender difference in susceptibility to the acute lethality and hepatotoxicity of TCDD; rWT mice were highly sensitive, DEL mice moderately resistant and INS mice highly resistant. Co-expression of mouse AHR and rWT resulted in augmented sensitivity to TCDD and abolished the natural resistance of female C57BL/6 mice, whereas mice co-expressing mouse AHR and INS were resistant. Thus, these transgenic mouse lines provide a novel promising tool for molecular studies on dioxin toxicity and AHR function.     

The AH receptor and a novel gene determine acute toxic responses to TCDD: segregation of the resistant alleles to different rat lines

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD),12 the most toxic congener of dioxins, exhibits wide sensitivity differences between a sensitive Long-Evans (L-E) rat and a resistant Han/Wistar (H/W) rat. The sensitivity is determined probably by two autosomal genes and it is highly end point dependent. The difference is more than 1000-fold for acute toxicity and negligible for CYP1A1 induction. The rat strains were recently shown to have differences in the size of AH receptor (AHR), which mediates most effects of TCDD. In the present study, the rat strains were crossed and the resistant alleles of genes determining TCDD sensitivity were segregated to new rat lines. Selection was based on AHR phenotype determined by Western blot and resistance to TCDD lethality. Two genes determining resistance were found: the Ahr and a novel gene designated "B." In homozygous rats, the H/W type Ahrhw allele prevented TCDD lethality up to 2000 microg/kg or more, and the H/W type "Bhw" allele also increased resistance to TCDD lethality but to a lesser extent. Heterozygous rats were only slightly more resistant to acute lethality than the respective sensitive homozygous rats. CYP1A1 induction was similar irrespective of the Ahr and "B" genotypes, but a substantial increase in serum bilirubin seen after low doses in sensitive rats occurred only after large doses in "Bhw/hw" and not at all in Ahrhw/hw rats. In conclusion, the Ahrhw allele is a major determinant of the exceptional resistance of H/W rats to TCDD lethality. There is also an additional gene, whose function remains to be characterized, conferring limited resistance to TCDD toxicity. These two H/W rat-derived alleles are separately expressed in the new rat lines created.     

2,3,7,8-Tetrachlorodibenzo-p-dioxin-induced change in intestinal function and pathology: evidence for the involvement of arylhydrocarbon receptor-mediated alteration of glucose transportation

Although numerous studies have been performed to clarify the mechanism(s) underlying the toxicological responses induced by dioxins, their effect on the intestine is less well understood. To address this issue, we examined the effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on the pathology and function of the intestine in arylhydrocarbon receptor (AhR)-sensitive (C57BL/6J) and -less-sensitive (DBA/2J) mice. A single oral administration of TCDD (100 mug/kg) to C57BL/6J mice produced changes in villous structure and nuclear/cytoplasm ratio in the epithelial cells of the intestine. Furthermore, in an oral glucose tolerance test, the serum glucose level was significantly increased in the C57BL/6J mouse but not in the DBA/2J mouse by TCDD treatment. In agreement with this, the expression of intestinal mRNAs coding sodium-glucose co-transporter 1 (SGLT1) and glucose transporter type 2 were increased only in C57BL/6J mice by TCDD. The increase in the former transporter was also confirmed from its protein level. The glucose level in the intestinal contents is thought to be one of the factors contributing to SGLT1 induction. Concerning with this, the intestinal activity of sucrase and lactase was significantly increased only in C57BL/6J mice by TCDD. These results suggest that while TCDD produces initial damage to the intestinal epithelium, the tissues induce SGLT1 to facilitate the absorption of glucose, which is expected, at least partially, to combat the wasting syndrome induced by TCDD. The data provided here also suggest that AhR is involved in the mechanism of SGLT1 induction.     

Acute Toxicity of Perfluorodecanoic Acid in C57BL/6 Mice Differs from 2,3,7,8-Tetrachlorodibenzo-p-dioxin

Perfluorodecanoic acid (PFDA) is a 10-carbon straight-chainfatty acid. Its toxicity in rats has been reported to resemblethat produced by exposure to 2,3,7,8-tetrachlorodibenzo-P-dioxin(TCDD). Mice which are "responsive" to TCDD toxicity carry theAh^b allele, while mice homozygous for the Ah^d gene are lesssensitive to TCDD toxicity. To characterize the toxicity ofPFDA and determine if it is under the control of the Ah locus,female responsive C57BL/6N (Ah^b/d) mice and congenic C57BL/6Jmice, differing only at the Ah locus (responsive, Ah^b/b; heterozygousresponsive, Ah^b/d and "nonresponsive," Ah^d/d), were administereda single oral dose of PFDA, at levels from 0 to 320 mg/kg bodyweight, observed daily for overt signs of toxicity, and weighedthree times weekly. In the wild-type congenic C57BL/6J (Ah^b/b)subline, mice were killed at 2, 7, 14, and 30 days followingexposure. All other mice were killed on Day 30. Serum was takenfrom the C57BL/6N mice for analysis of thyroid hormone levels.Selected organs from all mice were weighed and fixed for histopathologicalexamination. Dose-related mortality was observed as early as5 days postexposure and time-to-death was inversely relatedto dose. Dramatic decreases in body weight occurred shortlyfollowing treatment in all strains. Serum triiodothyronine (T₃)and thyroxine (T₄) levels increased with increasing dose. Therewas a marked increase (p < 0.05) in absolute and relativeliver weights and a significant decrease in thymus weights.Hepatocellular hypertrophy was observed in all treated miceother than controls. A marked increase in hepatocyte peroxisomeswas observed in all treatment groups. Thus, in contrast to TCDD,the acute toxicity of PFDA in the female C57BL/6 mouse doesnot vary with the Ah allele and is distinct from that reportedfor TCDD.     

Biphasic response for hepatic microsomal enzyme induction by 2,3,7,8-tetrachlorodibenzo-p-dioxin in C57BL/6J and DBA/2J mice

The induction of the murine hepatic microsomal cytochrome P-450 monooxygenase system by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) was studied over a wide range of doses, including those associated with acute toxicity. Studies were conducted in two inbred strains of mice which vary at the Ah receptor and at a number of other genetic loci. C57BL/6J mice possess a high-affinity Ah receptor and are responsive to enzyme inductive effects of TCDD, whereas DBA/2J mice do not possess a high-affinity receptor and are less responsive to TCDD. In a dose-response study, 7-ethoxyresorufin O-deethylase (EROD) activity appeared to be maximally induced in C57BL/6J and DBA/2J mice at 7 days following exposure to 3 and 30 micrograms of TCDD/kg respectively. Very similar results were reported previously for the induction of aryl hydrocarbon hydroxylase activity in these strains of mice. However, at higher doses of TCDD (at least 45 micrograms/kg for C57BL/6J and 300 micrograms/kg for DBA/2J), EROD activity was further increased (2-fold) from the apparent maximal (plateau) level, resulting in an unusual biphasic log dose-response relationship. EROD activity remained at these elevated rates in both strains for doses approaching and exceeding the respective LD50 values for each strain. To further characterize this biphasic induction phenomenon, cytochrome P-450 content, benzo[a]pyrene metabolism, and EROD and NADPH-cytochrome P-450 reductase activities were measured 1, 3 and 7 days after TCDD administration to C57BL/6J (3 and 150 micrograms/kg) and DBA/2J (30 and 600 micrograms/kg) mice. Maximal responses occurred in both strains at 3 days for all doses. In both strains, TCDD produced a dose-dependent increase in cytochrome P-450 content, EROD, and benzo[a]pyrene metabolism. Furthermore, a 2-fold induction of reductase activity was observed in each strain following exposure to the respective high doses. Induction of cytochrome P1-450 and P3-450 was also measured by Western immunoblot, using antisera raised against the homologous rat isozymes. In both strains, TCDD produced a dose-related increase in two protein-staining bands recognized by anti-P-450BNF-B (P1-450) and anti-P-450BNF/ISF-G (P3-450) respectively. The extended induction of hepatic microsomal monooxygenase activities at the respective high doses of TCDD appears to be due, in part, to increases in NADPH-cytochrome P-450 reductase activity and cytochromes P1-450 and P3-450 content. Significant alterations in the expression of the cytochrome P-450 monooxygenase system following exposure to high doses of TCDD may be associated, in part, with the delayed acute toxicity reported at this level of exposure.     

Ah receptor in mice genetically "nonresponsive" for cytochrome P4501A1 induction: cytosolic Ah receptor, transformation to the nuclear binding state, and induction of aryl hydrocarbon hydroxylase by halogenated and nonhalogenated aromatic hydrocarbons in embryonic tissues and cells.

The aromatic hydrocarbon (Ah) receptor mediates induction of cytochrome P4501A1 and associated aryl hydrocarbon hydroxylase (AHH) activity in tissues or cells exposed to polycyclic aromatic hydrocarbons. Strains of mice designated "nonresponsive" do not show increased hepatic AHH activity when exposed in vivo to nonhalogenated aromatic hydrocarbons such as 3-methylcholanthrene, benz[a]anthracene (BA), or benzo[a]pyrene and have reduced sensitivity to halogenated inducers such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Recently, with a modified assay, we detected Ah receptor in hepatic cytosols from adult nonresponsive mice [Mol. Pharmacol. 35:823-830 (1989)]; the receptor was present in reduced amount, and the apparent affinity for TCDD was lower than in hepatic cytosol from responsive C57BL/6J mice. Using the same assay procedure, we now report detection of Ah receptor in cytosols prepared from embryonic tissue and from cultured embryo cells of both responsive (C57BL/6J) and nonresponsive mice (DBA/2J, AKR/J, and SWR/J). Cytosolic receptor in embryonic cells from nonresponsive as well as responsive strains was detectable both with [3H]TCDD and with [3H]3-methylcholanthrene. In addition, the receptor-ligand complex could be extracted from nuclei of embryo cells exposed to [3H]TCDD in culture. AHH activity was induced in embryo cell cultures incubated with either TCDD or BA. The EC50 values for AHH induction were virtually identical in cell cultures from nonresponsive (DBA/2J) and responsive (C57BL/6J) strains, using either TCDD or BA as the inducer. Moreover, the affinity with which [3H]TCDD bound to cytosolic Ah receptor was much more similar in cytosols from cell cultures from the two strains than in cytosols prepared from adult liver. Thus, embryonic cell cultures differ in at least three respects from the adult liver, as follows: (i) Ah receptor can be detected with [3H]3-methylcholanthrene in embryonic cell cytosols but not in cytosols from adult liver; (ii) the degree of difference between nonresponsive and responsive strains in the affinity with which [3H]TCDD binds to receptor is only about 2-fold in cytosol from embryonic cells, whereas it is almost 10-fold in adult liver; and (iii) induction of AHH activity (by either TCDD or by the nonhalogenated inducer BA) shows no significant difference between strains in embryonic cell culture, whereas there is at least a 15-fold difference in responsiveness between C57BL/6J and DBA/2J mice in adult liver in vivo. The mechanistic reason for the diminished degree of difference between responsive and nonresponsive mice during embryonic cell culture (compared with adult tissues) is not yet known.     

Dose-response relationship of cytochrome P4501b1 mRNA induction by 2,3,7,8-tetrachlorodibenzo-p-dioxin in livers of C57BL/6J and DBA/2J mice

 The dose-response relationship of cytochrome P4501b1 (Cyp1b1) and Cyp1a1 induction in livers of TCDD-treated female C57BL/6J and DBA/2J mice are described. The animals were treated i.p. with 0.001, 0.01, 0.1, 1, 10 and 50 μg TCDD/kg for 24 h, and Cyp1b1 and Cyp1a1 mRNA expression was analyzed by RT-PCR. In the livers of both mouse strains, the Cyp1b1 and Cyp1a1 mRNA content was increased after TCDD exposure in a dose-dependent manner. These effects were more pronounced in TCDD-responsive C57BL/6J mice than in the less responsive DBA/2J mice, although Cyp1a1 was more responsive to TCDD than Cyp1b1 in both strains. The calculated ED₅₀ values for Cyp1b1 and Cyp1a1 induction in livers of TCDD-treated C57BL/6J mice were 1.3 and 0.08 μg TCDD/kg, respectively. The corresponding values for half-maximal induction response in livers of DBA/2J mice were 3.4 μg TCDD/kg for Cyp1b1 and 1.5 μg TCDD/kg for Cyp1a1. These results show that Cyp1b1 mRNA expression is less inducible by TCDD than Cyp1a1. Both genes are highly inducible in TCDD-responsive C57BL/6J mice expressing the high affinity arylhydrocarbon receptor (Ah receptor), suggesting that Cyp1b1, like Cyp1a1, is a potential Ah receptor-regulated gene. Received: 8 December 1995/Accepted: 6 February 1996     

Characterization of a specific binding protein for 2,3,7,8-tetrachlorodibenzo-p-dioxin in human thymic epithelial cells

Specific toxic and biochemical responses elicited by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in human thymic epithelial (TE) cells in culture are mediated by the TCDD receptor protein. Characterization of the physicochemical properties of the TCDD receptor in cytosol fractions from cultured human TE cells indicates that this protein is similar, but not identical, to the extensively studied receptor species present in mouse and rat liver. The human TCDD receptor sediments at 9.1 S on sucrose density gradients at 0 degrees, undergoes a temperature-dependent conversion at 20 degrees to a species sedimenting at 10.7 S, and partially dissociates in the presence of 0.4 M KCl, as judged by the appearance of a peak sedimenting at 5 S. Both the temperature- and salt-dependent changes in the observed physical properties of the human TCDD receptor are inhibited by sodium molybdate. Two molybdate-stabilized binding species can be resolved from TCDD specific binding isotherms to human TE cytosol. Under identical conditions, only a single TCDD specific binding component was detected in cytosol fractions from both mouse and rat liver. Mixing cytosol prepared from human TE cells with hepatic cytosol fractions from C57BL/6 mice revealed the presence of a heat-labile, trypsin-sensitive factor in human TE cells that inhibits TCDD specific binding to the murine hepatic receptor. Molybdate stabilized the mouse receptor against the actions of this human inhibitory factor. Molybdate also may stabilize the human TCDD receptor, as indicated by the 2- to 3-fold increase in total TCDD specific binding measured in human TE cytosol fractions in the presence of this metallo-oxyanion.     

Dioxin-induced retardation of development through a reduction in the expression of pituitary hormones and possible involvement of an aryl hydrocarbon receptor in this defect: A comparative study using two strains of mice with different sensitivities to dioxin

We have previously revealed that treating pregnant rats with 2,3,7,8-tetracholorodibenzo-p-dioxin (TCDD) reduces the expression of gonadotropins and growth hormone (GH) in the fetal and neonatal pituitary. A change in gonadotropin expression impairs the testicular expression of steroidogenic proteins in perinatal pups, and imprint defects in sexual behavior after reaching maturity. In this study, we examined whether TCDD also affects the expression of gonadotropin and GH in mice using C57BL/6J and DBA/2J strains which express the aryl hydrocarbon receptor (Ahr) exhibiting a different affinity for TCDD. When pregnant C57BL/6J mice at gestational day (GD) 12 were given oral TCDD (0.2–20 μg/kg), all doses significantly attenuated the pituitary expression of gonadotropin mRNAs in fetuses at GD18. On the other hand, in DBA/2J mice, a much higher dose of TCDD (20 μg/kg) was needed to produce a significant attenuation. Such reduction in the C57BL/6J strain continued until at least postnatal day (PND) 4. In agreement with this, TCDD reduced the testicular expression of steroidogenic proteins in C57BL/6J neonates at PND2 and 4, although the same did not occur in the fetal testis and ovary. Furthermore, TCDD reduced the perinatal expression of GH, litter size and the body weight of newborn pups only in the C57BL/6J strain. These results suggest that 1) also in mice, maternal exposure to TCDD attenuates gonadotropin-regulated steroidogenesis and GH expression leading to the impairment of pup development and sexual immaturity; and 2) Ahr activation during the late fetal and early postnatal stages is required for these defects.     

Comparative responsiveness of hypothyroxinemia and hepatic enzyme induction in Long-Evans rats versus C57BL/6J mice exposed to TCDD-like and phenobarbital-like polychlorinated biphenyl congeners.

Numerous mechanisms have been postulated to explain how polyhalogenated aromatic hydrocarbons alter thyroid homeostasis with almost all data derived from studies using the rat. This study compared the sensitivity of rats and mice to polychlorinated biphenyl (PCB)-induced hypothyroxemia. Male and female C57BL/6J mice and Long-Evans rats were dosed orally for 4 consecutive days with either PCB126 (0.03-300.0 microg/kg/day) or PCB153 (0.3-300.0 mg/kg/day). Trunk blood and livers were collected 24 h after the last dose and used to determine total serum thyroxine (T(4)) and hepatic microsomal T(4) glucuronidation activity. Hepatic microsomal ethoxyresorufin-O-deethylase (EROD) and pentoxyresorufin-O-deethylase (PROD) activities were also determined as markers for Ah receptor or phenobarbital response unit activation, respectively. PCB126 did not affect T(4) in the mouse but decreased T(4) (up to 50%) in the rat. PCB153 decreased T(4) (up to 80%) in both the rat and the mouse. PCB126 increased EROD in both rats (12- to 22-fold) and mice (15- to 20-fold). PCB153 induced hepatic PROD activity in both rats (30-fold) and mice (4-fold). T(4) glucuronidation was increased approximately 2- to 3-fold in both rats and mice treated with PCB153. PCB126 increased T(4) glucuronidation 13-fold in rats but only marginally (20%) in mice at the highest doses. Western blot analysis confirmed the PCB126-induced changes in expression of UGT1A in rats and the minimal increase in mice. These data suggest that species differences in response to chemicals that induce hypothyroxinemia are due to differential induction of hepatic UGT enzymes.     

Adrenocorticotropin (ACTH) and corticosterone secretion by perifused pituitary and adrenal glands from rodents exposed to 2,3,7, 8-tetrachlorodibenzo-p-dioxin (TCDD)

Although in utero maternal stress has been shown to have lasting effects on rodent offspring, fetal effects of chemically-induced alterations of the maternal hypothalamic-pituitary-adrenal axis (HPA) have not been well studied. This study examined the effects of in vivo 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure on pituitary-adrenal function in the male rat, pregnant female rat and pregnant female mouse. The secretion of adrenocorticotropin (ACTH) and corticosterone (CORT) in pituitary and adrenal glands, respectively, was assessed in ex vivo perifusion cultures. Male and pregnant female (gestation day 8) Sprague-Dawley rats were gavaged once with 10 microgram/kg TCDD, pregnant female mice once with 24 microgram/kg TCDD, and euthanized 10 days later. Hemi-pituitary (rat) or whole anterior pituitaries (mice) and right adrenal glands from the same animal were quartered, perifused under baseline and stimulated conditions. In both males and pregnant females, TCDD did not affect corticotropin releasing hormone (CRH)-stimulated ACTH secretion. Neither total pituitary ACTH nor plasma ACTH was altered in either sex or species by TCDD treatment. ACTH-stimulated CORT secretion was not affected by TCDD in either sex or species, and adrenal tissue and plasma CORT levels were unchanged in males and pregnant females by TCDD. However, the plasma ACTH:CORT ratio was decreased about 46% in male rats treated with TCDD. Plasma CORT levels were 23-fold higher and plasma ACTH levels were 1.5-fold higher in pregnant females than in male rats. In male versus female rats, adrenal CORT and anterior pituitary ACTH tissue levels were about 7.5- and 1.75-fold higher and ACTH, respectively. Female mouse adrenal tissue CORT was about 4-fold greater than female rat. The reduced plasma ACTH:CORT ratio in the male rat suggests that TCDD disturbs HPA function. Exposure of male rat to a 5-fold higher dose in earlier studies clearly demonstrated effects of TCDD on male rat HPA. The present study identified substantial HPA performance differences between male and pregnant female rats. The failure to detect a response to TCDD in pregnant female rat and mouse could be a function of both TCDD dose and the high level of secretion of both ACTH and CORT in pregnant animals. For the rat or mouse, a single exposure to TCDD during pregnancy does not appear sufficient to induce maternally-mediated developmental, reproductive and behavioral toxicity via the HPA axis.     

Synergistic interactions of 2,3,7,8-TCDD and 2,2',4,4',5,5'-hexachlorobiphenyl in C57BL/6J and DBA/2J mice: role of the Ah receptor

Treatment of C57BL/6J mice with 2,2',4,4',5,5'-hexachlorobiphenyl (HCBP, 500 mumol/kg) elevated hepatic cytosolic Ah receptor levels 82-107% for up to 14 days. Scatchard analysis of the [3H]2,3,7,8-TCDD (TCDD)-Ah receptor saturation binding curves from corn oil and HCBP treated rats gave KD values of 0.80 and 0.90 nM, respectively and confirmed that treatment with HCBP did not significantly alter receptor-radioligand affinities. Administration of HCBP to DBA/2J mice did not result in detectable hepatic cytosolic Ah receptor levels. Cotreatment of C57BL/6J mice with HCBP (500 mumol/kg) at a dose level of TCDD (1 nmol/kg) which elicited less than 10% of the maximum induction response resulted in significant synergistic induction of hepatic EROD and AHH [compared to animals treated only with TCDD (1 nmol/kg)]. In contrast, cotreatment of C57BL/6J mice with HCBP (500 mumol/kg) and maximally inducing dose levels of TCDD (100 or 500 nmol/kg) resulted in either a slight or no difference in the induction of AHH or EROD compared to the induction responses observed in mice treated only with TCDD. In contrast, cotreatment of DBA/2J mice with TCDD and HCBP (500 mumol/kg) resulted in significant synergistic induction of AHH and EROD at both submaximal (10-500 nmol/kg) and maximal (5000 nmol/kg) induction levels of TCDD. The only significant interactive effect of HCBP (500 mumol/kg) on the toxicity of TCDD in C57BL/6J and DBA/2J was protection from body weight loss observed after cotreatment of HCBP and TCDD in DBA/2J mice.     

2,3,7,8-Tetrachlorodibenzo-p-dioxin-induced porphyria in genetically inbred mice: partial antagonism and mechanistic studies

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) (233 nmol/kg) causes a significant increase of hepatic uroporphyrin, heptacarboxyporphyrin, and total porphyrins in female C57BL/6 mice, ovariectomized C57BL/6 mice, male C57BL/10 mice, and male C57BL/6 mice 3 weeks after treatment. In contrast, 6-methyl-1,3,8-trichlorodibenzofuran (MCDF) was inactive at a dose of 750 mumol/kg. Cotreatment of the mice with TCDD (233 mol/kg) plus MCDF (750 mumol/kg) resulted in partial antagonism of TCDD-induced hepatic porphyrin accumulation only in the female mice. Parallel studies in female C57BL/6 mice showed that the TCDD-induced porphyria was accompanied by the induction of hepatic microsomal aryl hydrocarbon hydroxylase (AHH) and ethoxyresorufin O-deethylase (EROD) activities and the depression of uroporphyrinogen decarboxylase (UROD). MCDF (750 mumol/kg) did not significantly affect these enzymes. In the cotreatment studies (MCDF plus TCDD), MCDF partially antagonized TCDD-induced hepatic porphyrin accumulation but did not affect the levels of hepatic AHH, EROD, or UROD. These results indicate that other factors, in addition to the induction of cytochrome P450-dependent monooxygenases and depressed UROD activity, are important in TCDD-induced porphyria in C57BL/6 female mice.     

Rat and mouse differences in gender-predominant expression of organic anion transporter (Oat1-3; Slc22a6-8) mRNA levels.

Organic anion transporters (Oats) mediate the initial step of active renal excretion, specifically substrate uptake into proximal tubule cells. Despite extensive characterization of rat Oats, mouse Oat expression patterns are virtually unknown. This study was designed to identify basal expression patterns of mouse Oat1 (Slc22a6), Oat2 (Slc22a7), and Oat3 (Slc22a8) mRNA, compare these patterns with those in rat, and characterize postnatal development of mouse Oat mRNA. Tissues were collected from adult male and female 129J and C57BL/6 mice, and male and female C57BL/6 mice 0 to 40 days of age. Oat mRNA levels were determined by branched DNA signal amplification. Mouse Oat1 mRNA was primarily expressed in kidney of both strains, with male predominance. Mouse Oat2 mRNA levels were highest in kidney of both strains without gender predominance. In both strains, Oat3 mRNA was highest in kidney, and liver expression was male-predominant. However, only 129J mice had higher Oat3 mRNA levels in female kidney than in male kidney. During postnatal development, both Oat1 and Oat2 mRNA levels began to rise after 25 days of age. Oat3 mRNA levels rose gradually from birth through 40 days of age. Oat2 mRNA increased 30-fold during the first 40 days, whereas Oat1 and Oat3 increased about 2-fold. The most notable species differences in Oat mRNA expression were a lack of Oat2 female predominance in mouse kidney and a less dramatic Oat3 male predominance in mouse liver. With the exception of a significant species difference in Oat2 expression, many similarities were found between rat and mouse Oat mRNA levels.     

Prenatal TCDD in mice increases adult autoimmunity

Two immunologically different mouse strains, C57BL/6 and SNF₁, were exposed to a mid-gestation dose of TCDD. The C57BL/6 mouse has a high-affinity aryl hydrocarbon receptor (AhR) and is sensitive to TCDD. The SNF₁ mouse has a low-affinity AhR but spontaneously develops autoimmune nephritis. Autoreactive Vβ⁺CD4⁺17a and Vβ⁺CD3⁺ T cells were increased at 24-weeks-of-age in offspring of C57BL/6 mice, more so in females than males. The cytokine IFN-γ was elevated in the females, while IL-10 was elevated in males. Phenotypic changes in B-lineage cells were present in bone marrow and spleen, and circulating autoantibodies were increased after prenatal TCDD. Kidneys of males showed significant anti-IgG and anti-C3 deposition, suggesting early-stage autoimmune disease. The SNF₁ offspring similarly showed increased peripheral Vβ⁺ cells in the females, increased autoantibody production in both sexes, and increased IFN-γ production in females. Male SNF₁ mice had increased anti-IgG and anti-C3 deposition in kidneys. Both mouse models therefore showed clear signatures of enhanced autoimmunity after prenatal TCDD.