Superfund Research Program

TCDD-Elicited Steatosis: The Role of Aryl Hydrocarbon Receptor Regulation in Lipid Uptake, Metabolism, and Transport

Project Leader: Timothy R. Zacharewski
Grant Number: P42ES004911
Funding Period: 2006-2021

Project-Specific Links

Final Progress Reports

Year:   2020  2012 

TCDD and related compounds elicit a variety of species-specific responses. Dr. Zacharewski and his team previously demonstrated that TCDD induces hepatic steatosis in mice which is reversible following the gradula clearance of hepatic TCDD in 4 weeks. The team's efforts have focused on 1) extending previous assessments of hepatic lipidomics to include lipid distribution in the serum and adipose tissue as well as the liver of fasted mice(Toxicol Sci. 125, 41-55, 2012; Toxicol Sci. 128, 377-86, 2012; Toxicol Sci. 131, 108-15, 2012; Toxicol Appl Pharmacol. 267, 184-91, 2013), and 2) determine the dose-dependent impact of repeated exposure to TCDD (every 4 days) on hepatic gene expression and lipid metabolism at 28 days.

Characterization of liver, serum, and gonadal white adipose tissue (gWAT) lipid composition was examined in wild type B6.129-Scd1tm1Myz/J mice orally gavaged with a single dose of 30 μg/kg TCDD. Animals were fasted for 4 h prior to sacrifice at 24, 72, and 168h post-dose. TCDD increased hepatic total fatty acid (TFA) content 1.7-fold while no FA changes were observed in serum and gWAT. The molar percentage of specific fatty acids in TCDD-treated relative to vehicle controls (mol%) were consistent with the induction of stearoyl-CoA desaturase 1 activity index associated with decreases in palmitate (16:0) and stearate (18:0), and increases in palmitate (16:1n7) and oleate (18:1n9). Similarly, although TFA levels did not increase in the serum, a 1.4-fold increase in the serum desaturation index was observed. Total serum cholesterol (CHOL), HDL CHOL, and LDL CHOL decreased following TCDD exposure and serum Apob48 and Apob100 apolipoproteins, found primarily in intestinal chylomicrons and hepatic VLDL particles were also decreased suggesting an inhibition of lipid efflux by TCDD. Hepatic and gWAT gene expression analysis revealed repression of Fasn, Dgat2, Hmgca1, Hmgcr, and Apoa1 and increased expression of Lcat and Ldlr in the liver. In the gWAT, Pnpla3 expression was repressed while Dgat1, Slc27a1, Adipoq, and Scd2 were induced. These gene expression changes are consistent with hepatic lipid accumulation and reduced transport of CHOL, as well as increased desaturation of FAs. The absence of increased serum FAs following a 4h fast supports previous reports of dietary lipids as a primary source of lipids for TCDD elicited hepatic accumulation.

Female C57BL/6 mice were gavaged with 0.001, 0.01, 0.03, 0.1, 0.3, 10, and 30 µg/kg TCDD or sesame oil vehicle every 4 days for a total of 28 days (7 doses). Repeated dosing did not alter body weight gain, but increased relative liver weight at 3, 10, and 30 µg/kg while gWAT weight decreased 50% at 30 µg/kg suggesting possible mobilization of adipose lipids. Global hepatic gene expression assessment using whole-genome 4x44k Agilent microarrays identified a total of 1,232 distinct genes exhibiting dose-dependent differentially expressed. Functional enrichment analysis identified over-represented functions associated with sugar metabolism, defense and immune responses, lipid metabolism, and glycerolipid metabolism among others. Dose-response modeling identified a median ED50 of 3.16 µg/kg TCDD for 944 unique genes showing a sigmoidal dose-response curve. Interestingly, assessment of the frequency distribution of ED50s suggests four distinct peaks at ~0.3, ~1, ~3, and ~9.3 µg/kg suggesting regulation of these genes may involve recruitment of different coregulators. Functional annotation of ED50 distribution peaks also suggests four functionally distinct peaks reflecting an increase in the severity of responses with increasing doses. Genes previously examined in 24h single TCDD dose exposures, Cyp1a1, Cyp1a2, Cyp1b1, Dysf, Gstp2, Notch1, Nqo1, Pla2g12a, Serpinb6a, Srxn1, Tiparp, Tnfaip2, Got1, and Pck1, were assessed by QRTPCR and exhibited a similar ED50 between repeated and single dose exposures. Altogether, these data suggest that repeated exposure leads to sustained AhR activation and gene regulation. Differential gene expression is currently being phenotypically anchored to morphometric assessments including fat accumulation, immune cell infiltration and fibrosis.