Chronic Exposure to Low Doses of Dioxin Promotes Liver Fibrosis Development in the C57BL/6J Diet-Induced Obesity Mouse Model

Abstract

Background: Exposure to persistent organic pollutants (POPs) has been associated with the progression of chronic liver diseases, yet the contribution of POPs to the development of fibrosis in non-alcoholic fatty liver disease (NAFLD), a condition closely linked to obesity, remains poorly documented.

Objectives: We investigated the effects of subchronic exposure to low doses of the POP 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), an aryl hydrocarbon receptor ligand, on NAFLD progression in diet-induced obese C57BL/6J mice.

Methods: Male C57BL/6J mice were fed either a 10% low-fat (LFD) or a 45% high-fat (HFD) purified diet for 14 weeks and TCDD-exposed groups were injected once a week with 5 μg/kg TCDD or the vehicle for the last 6 weeks of the diet.

Results: Liver histology and triglyceride levels showed that exposure of HFD fed mice to TCDD worsened hepatic steatosis, as compared to either HFD alone or LFD plus TCDD and the mRNA levels of key genes of hepatic lipid metabolism were strongly altered in co-treated mice. Further, increased liver collagen staining and serum transaminase levels showed that TCDD induced liver fibrosis in the HFD fed mice. TCDD in LFD fed mice increased the expression of several inflammation and fibrosis marker genes with no additional effect from a HFD.

Conclusions: Exposure to TCDD amplifies the impairment of liver functions observed in mice fed an enriched fat diet as compared to a low fat diet. The results provide new evidence that environmental pollutants promote the development of liver fibrosis in obesity-related NAFLD in C57BL/6J mice. Citation: Duval C, Teixeira-Clerc F, Leblanc AF, Touch S, Emond C, Guerre-Millo M, Lotersztajn S, Barouki R, Aggerbeck M, Coumoul X. 2017. Chronic exposure to low doses of dioxin promotes liver fibrosis development in the C57BL/6J diet-induced obesity mouse model. Environ Health Perspect 125:428-436; http://dx.doi.org/10.1289/EHP316.

Figure 1

Effect of TCDD on HFD-induced obesity and hepatic steatosis. Mice fed either a LFD or a HFD for a total of 14 weeks were injected with 5 μg/kg of TCDD (LF-tcdd and HF-tcdd, respectively) or the vehicle (LF-ctrl and HF-ctrl, respectively) during the last 6 weeks. (A) Body weight (BW) gain, inguinal and epididymal white adipose tissue (WAT) weight. (B) Leptin mRNA levels in epididymal WAT (eWAT) measured at 14 weeks and plasma fasted-leptin concentrations at 13 weeks. (C) Liver weight. (D) Hematoxylin-eosin staining (H&E) of representative liver sections of the different groups, black arrows indicate the islets of infiltrated inflammatory cells (bar = 150 μm). (E) Hepatic triglyceride content measured at 14 weeks. Note: Data are expressed as mean ± SEM; a, versus LF-ctrl; b, versus LF-tcdd; c, versus HF-ctrl; p < 0.05.

Figure 2

Effect of the co-exposure to TCDD and HFD on the hepatic mRNA levels of markers of lipid and carbohydrate metabolism. Mice fed either a LFD or a HFD for a total of 14 weeks were injected with 5 μg/kg of TCDD (LF-tcdd and HF-tcdd, respectively) or the vehicle (LF-ctrl and HF-ctrl, respectively) during the last 6 weeks. The mRNA levels of hepatic genes were measured by qPCR. Mean expression in the LF-ctrl group is set at 100%. (A) Markers of lipid accumulation. (B) Markers of lipogenesis and FA oxidation. (C) Markers of carbohydrate metabolism. Note: Data are expressed as mean ± SEM; a, versus LF-ctrl; b, versus LF-tcdd; c, versus HF-ctrl; p < 0.05.

Figure 3

Effect of the combined exposure to TCDD and HFD on the development of hepatic fibrosis. Mice fed either a LFD or a HFD for a total of 14 weeks were injected with 5 μg/kg of TCDD (LF-tcdd and HF-tcdd, respectively) or the vehicle (LF-ctrl and HF-ctrl, respectively) during the last 6 weeks. The hepatic mRNA levels of markers of (A) inflammation and (B) fibrosis were measured by qPCR. Mean expression in the LF-ctrl group is set at 100%. (C) Picro-sirius red staining shows fibrotic scars of collagen I and III (large black arrows, bar = 150 μm). (D) Quantification of picro-sirius red staining. (E) Serum alanine (ALAT) and aspartate (ASAT) aminotransferase activities. Note: Data are expressed as mean ± SEM; a, versus LF-ctrl; b, versus LF-tcdd; c, versus HF-ctrl; p < 0.05.

Figure 4

Schema of the effects of TCDD, high fat diet and their combination on liver gene expression and various end points. The effects of TCDD are shown in A, for the high-fat diet in B, and for their combination in C as compared to untreated mice (LF-ctrl). Question marks indicate end points for which the possible modifications have not been measured in the present study.