* Molecular and Environmental Toxicology Center, Department of Nutritional Sciences, University of Wisconsin, 1415 Linden Drive, Madison, Wisconsin 53706;
Department of Mathematics and Statistics, MSC03 2150, 1 University of New Mexico, Albuquerque, New Mexico 87131;
Department of Pediatrics, University of New Mexico, 2211 Lomas Blvd NE, Albuquerque, New Mexico 87131;
Medical Lab Sciences, MSC09 5250, 1 University of New Mexico, Albuquerque, New Mexico 87131; and
¶ College of Pharmacy, MSC09 5360, 1 University of New Mexico, Albuquerque, New Mexico 87131
Received July 1, 2003; accepted August 19, 2003
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ABSTRACT |
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Key Words: insulin regulation; embryonic cardiac enlargement; neonatal macrosomia; AhR.
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INTRODUCTION |
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The AhR is a member of the basic-helix-loop-helix PAS (Per-ARNT-SIM) transcription family, which also includes its dimerization partner ARNT, hypoxia-inducible factor 1-alpha (HIF1), PER1-3, and others. Many members of this protein family, including AhR, act as environmental sensors, regulating downstream responses to environmental cues (Gu et al., 2000
). For example, HIF1
is involved in the sensing and mediating the response to hypoxia (Wang et al., 1995
), while the PER proteins as well as CLOCK and NPAS are involved in the control of circadian rhythmicity in response to day/night cycles (King et al., 1997
; Reick et al., 2001
; Zheng et al., 1999
). Studies using AhR-null mice have identified new roles for the AhR in normal physiology and development, distinct from its role as an environmental sensor of toxicants (Abbott et al., 1999
; Fernandez-Salguero et al., 1997
; Lahvis et al., 2000
).
The AhR and its dimerization partner ARNT have been shown to be expressed in the developing mouse and avian heart (Abbott and Probst, 1995; Abbott et al., 1995
; Walker et al., 2000
). Treatment of chick embryos with TCDD has been demonstrated to result in cardiac malformations, and the expression of AhR and ARNT in the chick cardiovascular system is consistent with a potential role in this toxicity (Walker and Catron, 2000
; Walker et al., 1997
). Indeed, TCDD has been demonstrated to be a cardiovascular teratogen in all species tested (Harris et al., 1973
; Hassoun et al., 1984
; Hornung et al., 1999
; Walker and Catron, 2000
). Interestingly, mice lacking the AhR have been demonstrated to develop cardiac hypertrophy and hypertension, suggesting that the AhR is required for cardiovascular homeostasis in the adult mouse (Lund et al., in press; Thackaberry et al., 2002
). Given these data, we investigated the role of the AhR in cardiac development using AhR-null mice.
Here we show that AhR-null embryos develop increased heart weight, and that pregnant mice lacking the AhR develop altered insulin regulation and responsiveness as seen by decreased fasting plasma insulin levels and insulin resistance. However, hyperglycemia and altered glucose tolerance, characteristics of gestational diabetes, were not observed. These results suggest that, although the embryonic cardiac enlargement in mice born to AhR-null females was not associated with an overt diabetic condition, alterations in insulin regulation and tissue responsiveness cannot be eliminated as potential causative factors. In addition, we also report that 23% of nonpregnant seven-month-old female AhR-/- mice develop glucose intolerance, and that all AhR-null mice of this age show significantly reduced fasting plasma insulin and tend to exhibit insulin resistance. Despite these alterations in insulin regulation and responsiveness, nonpregnant seven-month-old AhR-/- mice secrete normal amounts of insulin in response to a bolus dose of glucose and do not exhibit hyperglycemia. These data demonstrate that AhR-null females develop altered insulin regulation during pregnancy or as they age, but that these effects are subtle and do not result in overt diabetes.
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MATERIALS AND METHODS |
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mRNA analysis.
mRNA was isolated from pooled (46 hearts) d14.5, d17.5, or neonatal hearts using Trizol (Life Technologies, Rockville, MD). Atrial natriuretic factor (ANF), beta-myosin heavy chain (ß-MHC), myosin light chain 2V (MLC-2V), and 28S ribosomal RNA (28S), were measured by ribonuclease protection assay (RPA), using an RPA II kit from Ambion (Austin, TX). The probe for MLC-2V was supplied by Dr. Gary Lyons (University of Wisconsin-Madison). Probes for murine ANF, ß-MHC, and 28S were generated by polymerase chain reaction (PCR), as previously described (Thackaberry et al., 2002). Four litters per genotype were used for each transcript. Blots were quantified using a Cyclone Storage Phosphor System phosphoimager (Packard) with OptiQuant software. Values were normalized against 28S RNA.
Glucose measurements.
All mice were maintained on a 12-h dark/light cycle, with lights off at 1800 h. Mice were deprived of food for 15 h prior to blood collection (fasted), or fed ad libitum. All samples collected from timed-pregnant mice were taken at d14.5 of gestation, and correct timing of the pregnancies was confirmed at parturition. For ad libitum fed glucose measurements, blood samples were taken at 2400 h (n = 4 for both genotypes), 0400 h (n = 4 for both genotypes), 1000 h (n = 19 for AhR+/+, 11 for AhR-/-), 1500 h (n = 19 for AhR+/+, 11 for AhR-/-), and 2000 h (n = 4 for both genotypes). For nonpregnant mice, plasma glucose was measured from seven-month-old female mice following 15 h of fasting (n = 5 for AhR+/+, 8 for AhR+/-, 9 for AhR-/-) or when fed ad libitum at or around 1200 (n = 4 for all genotypes). Plasma was collected by centrifugation at 2, 750 x g for 10 min at 4°C, and frozen at -20°C until analyzed. Glucose was measured using a Vitros Systems GLU DT enzymatic assay and analyzer (Endocrine Sciences Products, Calabasas Hills, CA). The upper detection limit was 450 mg/dl, and all values that exceeded this limit were reported as 450 mg/dl.
For glucose tolerance tests, two- to three-month-old pregnant mice (n = 4 per genotype), and three- (n = 6 for all genotypes) and seven-month-old (n = 14 for AhR+/+, 13 for AhR+/-, 17 for AhR-/-) nonpregnant mice were fasted for 15 h. An initial blood sample was collected, and mice were then given 2 g/kg glucose by oral gavage. Blood samples were taken at 15, 30, 60, 90, 120, and 150 min after gavage and analyzed for glucose. For insulin tolerance tests, randomly fed two- to three-month-old pregnant females or seven-month-old nonpregnant females were given 0.75 units of porcine insulin (Sigma, St. Louis, MO) per kg of body weight in phosphate buffered saline via ip injection, and blood samples were taken at 15, 30, and 60 min, analyzed for glucose, and compared to preinjection values.
C-Peptide measurements.
Two- to three-month-old pregnant (d14.5 of gestation, four mice per genotype), nonpregnant three- month-old (n = 11 for AhR+/+, 6 for AhR+/-, 14 for AhR-/-) and nonpregnant seven-month-old (n = 12 for AhR+/+, 8 for AhR+/-, 15 for AhR-/-) female mice were fasted for 15 h, anesthetized, and blood was collected via cardiac puncture using a heparinized syringe. Correct timing of the pregnancies was confirmed by morphological analysis of the embryos at the time of sacrifice. Plasma was collected by centrifugation at 2, 750 x g for 10 min at 4°C, and blood was then frozen at -20°C until analyzed. C-Peptide was measured using a radioimmunoassay (RIA, Linco Research, St. Charles, MO). For secretory insulin response to glucose, mice were fasted for 15 h, given 2 g/kg glucose via oral gavage, and blood was collected and C-peptide measured 15 min after glucose administration.
Hemoglobin A1c quantification.
Whole blood from nonpregnant female AhR-/- and wild-type mice at 3 (n = 5 for AhR+/+, 6 for AhR-/-), 4.5 (n = 3 for both genotypes) and 6.5 (n = 6 for AhR+/+, 4 for AhR-/-) months of age was collected via tail clip. Hemoglobin A1c was isolated using a boronate affinity column (Endocrine Sciences) and quantified using a spectrophotometer at 414 nm. Results are expressed as a percent of total hemoglobin concentration.
Statistics.
For analysis of the nonpregnant glucose tolerance tests, a Bonferroni adjustment of a mean-shift outlier test, accounting for censoring at the upper detection limit of 450 mg/dl, was used to segregate the outlying "glucose intolerant" AhR-/- mice from the "normal" AhR-/- mice (Weisberg, 1985). All other data sets and comparisons of individual time points from the glucose tolerance tests were compared using Students t test. A Kolmogorov-Smirnov (KS-distance) test was used to confirm normal distribution of the data for all other data sets. Statistical significance was set at p < 0.05 for all comparisons.
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RESULTS |
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Increased Expression of Cardiac Hypertrophy Genes in the Developing AhR-/- Heart
To further characterize the cardiac phenotype of embryonic and neonatal AhR-/- mice, we measured the expression of ß-MHC, ANF, and MLC-2V transcripts in the hearts of d14.5 and d17.5 embryos, as well as in neonates (Fig. 3). ß-MHC was upregulated 1030-fold at all developmental time points studied (Fig. 3A
), while ANF was upregulated significantly on d14.5 and in neonates (Fig. 3B
). MLC-2V transcript levels were not significantly increased in null mutant hearts at any time point examined (Fig. 3C
).
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AhR-/- Females Develop Glucose Intolerance with Age
To determine whether a diabetic phenotype developed with age, we performed glucose tolerance tests on three- and seven-month-old females of all three AhR genotypes. Although the peak levels of glucose were higher on average in AhR-null females at 15 and 30 min following an oral glucose load, no statistical difference was seen in glucose clearance among the three genotypes at three months of age (Fig. 7A). At seven months of age, however, AhR-null mice exhibited elevated but highly variable glucose levels at 15, 30, and 60 min after glucose administration (Fig. 7B
). Using a Bonferroni outlier test, we determined that this variability resulted from 23% (4/17) of the seven- month-old AhR-/- females exhibiting an impairment of glucose clearance. When these four AhR-null females were analyzed separately, their ability to clear glucose was significantly impaired compared to wild types, AhR+/-, and the remaining AhR-/- female mice (Fig. 7C
).
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AhR-/- Females Do Not Develop Increased Glycosylated Hemoglobin Concentrations
We next measured glycosylated hemoglobin A1c concentrations as a measurement of chronic hyperglycemia. Hemoglobin is glycosylated at higher rates in animals experiencing chronic hyperglycemia, and can be quantified as an indicator of hyperglycemia over time (Dan et al., 1997). Our results showed that AhR-/- female mice did not experience increased hemoglobin A1c levels at three, four-and-a-half, or six-and-a-half months of age (Fig. 10
). Interestingly, AhR-null females actually have significantly lower hemoglobin A1c levels at six-and-a-half months of age compared to wild types.
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DISCUSSION |
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AhR-/- mice born to AhR-/- females exhibited significantly enlarged hearts. This cardiac enlargement was associated with thicker ventricular walls and upregulation of the cardiac hypertrophy markers ß-MHC and ANF. ANF and ß-MHC are highly expressed during normal fetal cardiac development and are subsequently downregulated early postnatally (Mercadier et al., 1989). Both ß-MHC and ANF are markers of cardiac hypertrophy in adult animals (Sugden and Clerk, 1998
), and increased expression of ANF has been reported in neonatal hearts undergoing hypertrophy (Bruneau et al., 2001
; Walker and Catron, 2000
). To our knowledge, these findings are the first to report upregulation of ß-MHC and ANF associated with both embryonic and neonatal cardiac hypertrophy. In contrast, in normal animals MLC-2V is not highly expressed in the fetal heart, but is upregulated in the adult hypertrophied heart (Lee et al., 1988
). Thus, it is not inconsistent that MLC-2V was not increased in embryonic and neonatal AhR-/- hypertrophic hearts. The increased expression of ß-MHC and ANF, taken together with the thickening of the ventricular walls on d14.5 and d17.5 embryos indicate that AhR-/- mice experience cardiac hypertrophy prior to birth.
The increase in heart size in AhR-null neonates can also be explained by an increased in proliferation in the hearts of AhR-null embryos. At d14.5, AhR-null embryos had a significantly increased proliferation index, as measured by PCNA staining. This suggests that the cardiac enlargement seen in AhR-null mice may be a result of hyperplasia. While the induction of the cardiac hypertrophy markers ß-MHC and ANF suggests that this is a hypertrophic phenotype, it seems likely that both hypertrophy and hyperplasia are involved, since embryonic cardiomyocytes, unlike adult cardiomyocytes, are capable of cell division.
A progressive cardiac hypertrophy occurs in adult male AhR-null mice (Fernandez-Salguero et al., 1997; Thackaberry et al., 2002
). This adult cardiac hypertrophy appears to be mechanistically distinct from the neonatal hypertrophy, since the adult hypertrophy is manifested beginning at five months of age, while three-month-old AhR-null males have normal heart weights. Thus, embryonic and neonatal cardiac hypertrophy in AhR-/- mice may resolve following birth, when the fetus is removed from the maternal environment, and results in normal heart weights by three months of age.
A series of mating experiments revealed that the neonatal cardiac enlargement was dependent, at least in part, on the maternal AhR genotype. Mice born to AhR-/- females, including both AhR-/- and AhR+/- neonates, exhibited significantly increased heart weights compared to mice born to AhR+/+ females. In contrast, mice born to AhR+/- females, including AhR+/+, AhR+/-, and AhR-/- neonates, fail to exhibit cardiac enlargement, compared to mice born to AhR+/+ females. The effects of maternal AhR genotype on neonatal body weight and heart-to-body weight ratio were more complex. Heterozygous neonates born to AhR-/- females had significantly increased body weights, while their AhR-/- littermates did not. However, only AhR-/- neonates born to AhR-/- females showed an increase in heart/body weight ratio. The increase in heart-to-body weight ratio in these mice suggests that the cardiac enlargement seen in AhR-/- mice is dependent on a combination of both maternal and neonatal loss of AhR. The AhR+/- mice born to AhR-/- females show cardiac hypertrophy that is increased in proportion to body weight. In contrast, AhR-/- neonates show cardiac hypertrophy that is increased to a larger degree than their body weights. Neonates born to AhR+/- dams show no increases in heart weight, body weight, or heart-to-body weight ratio, further suggesting that the cause of the neonatal macrosomia is dependent on both maternal and embryonic AhR status.
Since diabetic pregnancy has been shown to increase neonatal heart and body weight in a manner dependent on maternal genotype alone (Rizzo et al., 1992; Spellacy et al., 1985
), we investigated glucose metabolism in pregnant AhR-/- females. Two hallmarks of gestational diabetes are glucose intolerance, or the inability to effectively clear a bolus dose of glucose, and insulin resistance, a reduced ability of peripheral tissues to uptake glucose following insulin release. To determine whether pregnant AhR-null females experienced gestational diabetes, we performed tests for glucose tolerance and insulin resistance. Pregnant AhR-/- mice are able to clear a bolus dose of plasma glucose normally; however, they exhibited a substantial delay in their ability to clear glucose in response to insulin. This suggests that the peripheral tissues in pregnant AhR-/- mice do not uptake plasma glucose in response to insulin as efficiently as controls, but that this insulin resistance was not sufficient to retard glucose clearance following a bolus dose of glucose. Thus, it appears that pregnant AhR-null mice exhibit a partial insulin resistance.
Impaired glucose uptake in response to insulin could be overcome by significantly increased insulin production, or hyperinsulinemia. Indeed, hyperinsulinemia is often seen in insulin resistant gestational diabetes (Buchanan et al., 1990). However, fasting C-peptide levels were significantly reduced, rather than elevated, in pregnant AhR-null mice. This result is contradictory to what would be expected if AhR-null mice were overcoming insulin resistance with increased insulin production. Furthermore, when challenged with a bolus dose of glucose, pregnant AhR-/- mice secreted normal levels of insulin. Thus, while the pregnant AhR-null mice had an apparent insulin resistance, this was not compensated for by increased insulin production.
While the decreased fasting insulin levels and insulin resistance seen in pregnant AhR-/- mice were not associated with abnormal glucose tolerance, we hypothesized that these abnormalities may be correlated with hyperglycemia under ad libitum feeding conditions. To investigate this possibility, we measured plasma glucose concentrations throughout a 24-h period. Our results demonstrate that pregnant AhR-null mice do not experience hyperglycemia at any time, and thus, the altered insulin regulation does not lead to hyperglycemia under any conditions studied.
One form of gestational diabetes, mature onset diabetes of the young-2 (MODY-2) shares many similarities with the phenotype seen in pregnant AhR-null mice. MODY-2 results when a single copy of the glucokinase gene is disrupted, leading to altered glucose sensing and reduced insulin secretion (Froguel et al., 1993). Thus, like pregnant AhR-/- mice, MODY-2 mice have altered basal insulin levels and normal ad libitum fed glucose levels. However, unlike AhR-null mice, MODY-2 mice develop mild hyperglycemia under fasting conditions (Bali et al., 1995
), exhibit abnormal glucose tolerance, and do not develop insulin resistance. While the similarities between the MODY-2 mouse model and pregnant AhR-null mice are intriguing, further studies are needed to characterize the defect leading to reduced insulin production in AhR-null animals.
We next investigated whether the AhR was also required for insulin regulation and responsiveness in nonpregnant mice. Our results demonstrate that nonpregnant female AhR-null mice exhibited decreased fasting plasma insulin levels and decreased sensitivity to insulin-mediated glucose uptake by seven months of age, and that 23% of seven-month-old AhR-/- female mice develop overt glucose intolerance. However, none of these effects were observed at three months of age.
While all three-month-old AhR-null females tested had normal glucose tolerances, 23% (4/17) of seven-month-old AhR-/- females showed significant glucose intolerance, suggesting that age increases the dysregulation of glucose in AhR-null females. The incomplete penetrance of this phenotype suggests that other factors may be involved, and that the percentage of animals exhibiting this phenotype could continue to increase with age. Glucose intolerance was not seen in pregnant 3- to 4-month-old AhR-null females, indicating that age, but not pregnancy, is one important risk factor for development of this abnormality. Unfortunately, due to the reduced reproductive capacity of AhR-null mice (Abbott et al., 1999), studies cannot be preformed on pregnant seven-month-old AhR-null females.
In addition to glucose intolerance, seven-month-old AhR heterozygous and null females also have decreased fasting plasma insulin levels that were not apparent at three months. This suggests that, like the development of glucose intolerance, the development of reduced fasting insulin production occurs with age. Interestingly, while three-month-old AhR-null females show no alteration in fasting insulin levels, pregnant AhR-null mice of similar age do exhibit decreased fasting insulin. This suggests that both pregnancy and increased age are contributing risk factors in the development of this phenotype. The reduction in fasting plasma insulin seen in AhR+/- mice is also significant. This suggests that loss of a single AhR allele is sufficient to alter insulin regulation in mice. The only other reported abnormality seen in AhR+/- mice is cardiac hypertrophy (Thackaberry et al., 2002), which, interestingly, is also seen at seven months of age.
Despite the reduction in fasting plasma insulin, impaired insulin secretion in response to glucose was not observed. These results indicate that AhR-/- mice are able to secrete normal amounts of insulin in response to a bolus dose of glucose. Therefore, it seems unlikely that the decreased fasting insulin plays a role in the impaired glucose tolerance in seven-month-old AhR-null females.
Seven-month-old AhR-null female mice also demonstrate a significantly reduced sensitivity to exogenous insulin. A similar insulin insensitivity is seen in pregnant AhR-/- mice, implicating age and pregnancy as contributing factors involved in this phenotype. Thus, it seems likely that reduced responsiveness to insulin plays a causative role in the impaired glucose tolerance.
The phenotype seen in AhR-null females resembles some characteristics of type II diabetes, including a reduced ability of peripheral tissues to absorb glucose in response to insulin (decreased insulin response) and impaired glucose tolerance. Furthermore, the reduced insulin sensitivity and glucose intolerant phenotypes in AhR-null females develop with age, while reduced insulin sensitivity also develops with pregnancy. Notably, age and pregnancy have been demonstrated to increase the risk of developing type II diabetes (Bertoni et al., 2002). This suggests that the AhR-null phenotype and type II diabetes share certain risk factors, and therefore may have some commonalities. Despite the similarities between type II diabetes and the female AhR-null phenotype, type II diabetes typically results in hyperglycemia, increased hemoglobin A1C, and hyperinsulinemia. Seven-month-old AhR-null females fail to exhibit any of these characteristics. In fact, they exhibit normal glycemic control, reduced hemoglobin A1C, and hypoinsulinemia.
It has been demonstrated that TCDD exposure and AhR activation are linked to alterations in insulin and glucose regulation, but these data fail to provide clear evidence of a mechanism by which the AhR regulates carbohydrate metabolism. TCDD causes hypoinsulinemia and hypoglycemia in animal models (Ebner et al., 1988; Gorski and Rozman, 1987
; Viluksela et al., 1999
), but is associated with hyperinsulinemia, hyperglycemia, insulin resistance, and increased risk for type II diabetes in humans (Bertazzi et al., 2001
; Cranmer et al., 2000
; Henriksen et al., 1997
; Michalek et al., 1999
). Thus, the role of the AhR in insulin regulation and tissue responsiveness is unclear.
These data demonstrate that pregnant AhR-null mice have altered insulin regulation, but do not exhibit overt gestational diabetes, and that nonpregnant AhR-null females develop altered insulin regulation by seven months of age, and 23% of these mice develop glucose intolerance. The partial dependence of the neonatal phenotype on the maternal genotype suggests a metabolic abnormality in the mother that promotes the development of macrosomia and cardiac hypertrophy; however, the ultimate cause of the neonatal cardiac hypertrophy and macrosomia is uncertain. In addition, nonpregnant seven-month-old AhR-null female mice do not develop hyperglycemia or increased hemoglobin A1C, indicating that they do not exhibit overt diabetes. These data demonstrate that the AhR is required for normal cardiovascular development, as well as insulin regulation in pregnant and nonpregnant female mice.
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ACKNOWLEDGMENTS |
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NOTES |
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REFERENCES |
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Abbott, B. D., and Probst, M. R. (1995). Developmental expression of two members of a new class of transcription factors: II. Expression of aryl hydrocarbon receptor nuclear translocator in the C57BL/6N mouse embryo. Develop. Dyn. 204, 144155.[ISI][Medline]
Abbott, B. D., Schmid, J. E., Pitt, J. A., Buckalew, A. R., Wood, C. R., Held, G. A., and Diliberto, J. J. (1999). Adverse reproductive outcomes in the transgenic Ah receptor-deficient mouse. Toxicol. Appl. Pharmacol. 155, 6270.[CrossRef][ISI][Medline]
Bali, D., Svetlanov, A., Lee, H. W., Fusco-DeMane, D., Leiser, M., Li, B., Barzilai, N., Surana, M., Hou, H., and Fleischer, N. (1995). Animal model for maturity-onset diabetes of the young generated by disruption of the mouse glucokinase gene. J. Biol. Chem. 270, 2146421467.
Bertazzi, P. A., Consonni, D., Bachetti, S., Rubagotti, M., Baccarelli, A., Zocchetti, C., and Pesatori, A. C. (2001). Health effects of dioxin exposure: A 20-year mortality study. Am. J. Epidemiol. 153, 10311044.
Bertoni, A. G., Krop, J. S., Anderson, G. F., and Brancati, F. L. (2002). Diabetes-related morbidity and mortality in a national sample of U. S. elders. Diabetes Care. 25, 471475.
Bruneau, B. G., Bao, Z. Z., Fatkin, D., Xavier-Neto, J., Georgakopoulos, D., Maguire, C. T., Berul, C. I., Kass, D. A., Kuroski-de Bold, M. L., de Bold, A. J., et al. (2001). Cardiomyopathy in Irx4-deficient mice is preceded by abnormal ventricular gene expression. Mol. Cell. Biol. 21, 17301736.
Buchanan, T. A., Metzger, B. E., Freinkel, N., and Bergman, R. N. (1990). Insulin sensitivity and B-cell responsiveness to glucose during late pregnancy in lean and moderately obese women with normal glucose tolerance or mild gestational diabetes. Am. J. Obstet. Gynecol. 162, 10081014.[ISI][Medline]
Cranmer, M., Louie, S., Kennedy, R. H., Kern, P. A., and Fonseca, V. A. (2000). Exposure to 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD) is associated with hyperinsulinemia and insulin resistance. Toxicol. Sci. 56, 431436.
Dan, K., Fujita, H., Seto, Y., and Kato, R. (1997). Relation between stable glycated hemoglobin A1C and plasma glucose levels in diabetes-model mice. Exp. Anim. 46, 135140.[ISI][Medline]
Denison, M. S., Fisher, J. M., and Whitlock, J. P., Jr. (1988). Inducible, receptor-dependent proteinDNA interactions at a dioxin-responsive transcriptional enhancer. Proc. Natl. Acad. Sci. U.S.A. 85, 25282532.[Abstract]
Ebner, K., Brewster, D. W., and Matsumura, F. (1988). Effects of 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin on serum insulin and glucose levels in the rabbit. J. Environ. Sci. Health Part B 23, 427438.[ISI]
Fernandez-Salguero, P. Pineau, T. Hilbert, D. M., McPhail, T., Lee, S. S., Kimura, S., Nebert, D. W., Rudikoff, S., Ward, J. M., and Gonzalez, F. J. (1995). Immune system impairment and hepatic fibrosis in mice lacking the dioxin-binding Ah receptor. Science 268, 722726.[ISI][Medline]
Fernandez-Salguero, P. M., Ward, J. M., Sundberg, J. P., and Gonzalez, F. J. (1997). Lesions of aryl-hydrocarbon receptor-deficient mice. Vet. Pathol. 34, 605614.[Abstract]
Fernandez-Salguero, P. M., Hilbert, D. M., Rudikoff, S., Ward, J. M., and Gonzalez, F. J. (1996). Aryl-hydrocarbon receptor-deficient mice are resistant to 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin-induced toxicity. Toxicol. Appl. Pharmacol. 140, 173179.[CrossRef][ISI][Medline]
Froguel, P., Zouali, H., Vionnet, N., Velho, G., Vaxillaire, M., Sun, F., Lesage, S., Stoffel, M., Takeda, J., and Passa, P. (1993). Familial hyperglycemia due to mutations in glucokinase. Definition of a subtype of diabetes mellitus. N. Engl. J. Med. 328, 697702.
Gorski, J. R., and Rozman, K. (1987). Dose-response and time course of hypothyroxinemia and hypoinsulinemia and characterization of insulin hypersensitivity in 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD)-treated rats. Toxicology 44, 297307.[CrossRef][ISI][Medline]
Gu, Y. Z., Hogenesch, J. B., and Bradfield, C. A. (2000). The PAS superfamily: sensors of environmental and developmental signals. Annu. Rev. Pharmacol. Toxicol. 40, 519561.[CrossRef][ISI][Medline]
Harris, M. W., Moore, J. A., Vos, J. G., and Gupta, B. N. (1973). General biological effects of TCDD in laboratory animals. Environ. Health Perspect. 5, 101109.[Medline]
Hassoun, E., dArgy, R., and Dencker, L. (1984). Teratogenicity of 2, 3, 7, 8-tetrachlorodibenzofuran in the mouse. J. Toxicol. Environ. Health Part A 14, 337351.
Henriksen, G. L., Ketchum, N. S., Michalek, J. E., and Swaby, J. A. (1997). Serum dioxin and diabetes mellitus in veterans of Operation Ranch Hand. Epidemiology 8, 252258.[ISI][Medline]
Hornung, M. W., Spitsbergen, J. M., and Peterson, R. E. (1999). 2, 3, 7, 8-Tetrachlorodibenzo-p-dioxin alters cardiovascular and craniofacial development and function in sac fry of rainbow trout (Oncorhynchus mykiss). Toxicol. Sci. 47, 4051.[Abstract]
King, D. P., Zhao, Y., Sangoram, A. M., Wilsbacher, L. D., Tanaka, M., Antoch, M. P., Steeves, T. D., Vitaterna, M. H., Kornhauser, J. M., Lowrey, P. L., et al. (1997). Positional cloning of the mouse circadian clock gene. Cell 89, 641653.[ISI][Medline]
Lahvis, G. P., Lindell, S. L., Thomas, R. S., McCuskey, R. S., Murphy, C., Glover, E., Bentz, M., Southard, J., and Bradfield, C. A. (2000). Portosystemic shunting and persistent fetal vascular structures in aryl hydrocarbon receptor-deficient mice. Proc. Natl. Acad. Sci. U.S.A. 97, 1044210447.
Lee, H. R., Henderson, S. A., Reynolds, R., Dunnmon, P., Yuan, D., and Chien, K. R. (1988). Alpha 1-adrenergic stimulation of cardiac gene transcription in neonatal rat myocardial cells. Effects on myosin light chain-2 gene expression. J. Biol. Chem. 263, 73527358.
Lund, A. K., Goens, M. B., Kanagy, N. L., and Walker, M. K. (2003). Cardiac hypertrophy in aryl hydrocarbon receptor (AhR) null mice is correlated with elevated angiotensin II, endothelin-1 and mean arterial blood pressure. Toxicol. Appl. Pharmacol. (in press).
Mercadier, J. J., Samuel, J. L., Michel, J. B., Zongazo, M. A., de la, Bastie, D., Lompre, A. M., Wisnewsky, C., Rappaport, L., Levy, B., and Schwartz, K. (1989). Atrial natriuretic factor gene expression in rat ventricle during experimental hypertension. Am. J. Physiol. 257, H979987.[ISI][Medline]
Michalek, J. E., Akhtar, F. Z., and Kiel, J. L. (1999). Serum dioxin, insulin, fasting glucose, and sex hormone-binding globulin in veterans of Operation Ranch Hand. J. Clin. Endocrinol. Metab. 84, 15401543.
Peters, J. M., Narotsky, M. G., Elizondo, G., Fernandez-Salguero, P. M., Gonzalez, F. J., and Abbott, B. D. (1999). Amelioration of TCDD-induced teratogenesis in aryl hydrocarbon receptor (AhR)-null mice. Toxicol. Sci. 47, 8692.[Abstract]
Reick, M., Garcia, J. A., Dudley, C., and McKnight, S. L. (2001). NPAS2: An analog of clock operative in the mammalian forebrain. Science 293, 506509.
Rizzo, G., Arduini, D., and Romanini, C. (1992). Accelerated cardiac growth and abnormal cardiac flow in fetuses of type I diabetic mothers. Obstet. Gynecol. 80, t-76.
Schmidt, J. V., and Bradfield, C. A. (1996). Ah receptor signaling pathways. Annu. Rev. Cell Dev. Biol. 12, 5589.[CrossRef][ISI][Medline]
Shimizu, Y., Nakatsuru, Y., Ichinose, M., Takahashi, Y., Kume, H., Mimura, J., Fujii-Kuriyama, Y., and Ishikawa, T. (2000). Benzo[a]pyrene carcinogenicity is lost in mice lacking the aryl hydrocarbon receptor. Proc. Natl. Acad. Sci. U.S.A. 97, 779782.
Spellacy, W. N., Miller, S., Winegar, A., and Peterson, P. Q. (1985). Macrosomiamaternal characteristics and infant complications. Obstet. Gynecol. 66, 158161.[Abstract]
Sugden, P. H., and Clerk, A. (1998). Cellular mechanisms of cardiac hypertrophy. J. Mol. Med. 76, 725746.[CrossRef][ISI][Medline]
Thackaberry, E. A., Galbaldon, D. M., Walker, M. K., and Smith, S. M. (2002). Aryl hydrocarbon receptor null mice develop cardiac hypertrophy and increased hypoxia-inducible factor 1-alpha in the absence of cardiac hypoxia. Cardiovasc. Toxicol. 2, 263273.[Medline]
Viluksela, M., Unkila, M., Pohjanvirta, R., Tuomisto, J. T., Stahl, B. U., Rozman, K. K., and Tuomisto, J. (1999). Effects of 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD) on liver phosphoenolpyruvate carboxykinase (PEPCK) activity, glucose homeostasis and plasma amino acid concentrations in the most TCDD-susceptible and the most TCDD-resistant rat strains. Arch. Toxicol. 73, 323336.[CrossRef][ISI][Medline]
Walker, M. K. and Catron, T. F. (2000). Characterization of cardiotoxicity induced by 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin and related chemicals during early chick embryo development. Toxicol. Appl. Pharmacol. 167, 210221.[CrossRef][ISI][Medline]
Walker, M. K., Heid, S. E., Smith, S. M., and Swanson, H. I. (2000). Molecular characterization and developmental expression of the aryl hydrocarbon receptor from the chick embryo. Comp. Biochem. Physiol. C. Toxicol. Pharmacol. 126, 305319.[ISI][Medline]
Walker, M. K., Pollenz, R. S., and Smith, S. M. (1997). Expression of the aryl hydrocarbon receptor (AhR) and AhR nuclear translocator during chick cardiogenesis is consistent with 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin-induced heart defects. Toxicol. Appl. Pharmacol. 143, 407419.[CrossRef][ISI][Medline]
Wang, G. L., Jiang, B. H., Rue, E. A., and Semenza, G. L. (1995). Hypoxia-inducible factor 1 is a basic-helixloophelix-PAS heterodimer regulated by cellular O2 tension. Proc. Natl. Acad. Sci. U.S.A. 92, 55105514.[Abstract]
Weisberg, S. (1985). Applied Linear Regression, 2nd ed. John Wiley, New York.
Zheng, B., Larkin, D. W., Albrecht, U., Sun, Z. S., Sage, M., Eichele, G., Lee, C. C., and Bradley, A. (1999). The mPer2 gene encodes a functional component of the mammalian circadian clock. Nature 400, 169173.[CrossRef][ISI][Medline]