Ablation of aryl hydrocarbon receptor promotes angiotensin II-induced cardiac fibrosis through enhanced c-Jun/HIF-1α signaling

Abstract

Aryl hydrocarbon receptor (AHR) is a transcription factor that binds to DNA as a heterodimer with the AHR nuclear translocator (ARNT) after interaction with ligands, such as polycyclic and halogenated aromatic hydrocarbons and other xenobiotics. The endogenous ligands and functions of AHR have been the subject of many investigations. In the present study, the potential role of AHR signaling in the development of left ventricular hypertrophy and cardiac fibrosis by angiotensin II (Ang II) infusion was investigated in mice lacking the AHR gene (Ahr^-/-). We also assessed the hypothesis that fenofibrate, a peroxisome proliferator-activated receptor-α (PPARα) activator, reduces cardiac fibrosis through the c-Jun signaling. Male Ahr^-/- and age-matched wild-type mice (n = 8 per group) were infused with Ang II at 100 ng/kg/min daily for 2 weeks. Treatment with Ang II increased systolic blood pressure to comparable levels in Ahr^-/- and wild-type mice. However, Ahr^-/- mice developed severe cardiac fibrosis after Ang II infusion compared with wild-type mice. Ang II infusion also significantly increased the expression of endothelin in the left ventricles of Ahr^-/- mice, but not in wild-type mice, and significantly increased the c-Jun signaling in Ahr^-/- mice. Ang II infusion also significantly enhanced the expression of hypoxia-inducible factor-1α (HIF-1α) and the downstream target vascular endothelial growth factor (VEGF) in the left ventricles of Ahr^-/- mice. These results suggested pathogenic roles for the AHR signaling pathway in the development of cardiac fibrosis. Treatment with fenofibrate reduced cardiac fibrosis and abrogated the effects of Ang II on the expression of endothelin, HIF-1α, and VEGF. The inhibitory effect of fenofibrate on cardiac fibrosis was mediated by suppression of VEGF expression through modulation of c-Jun/HIF-1α signaling.

Keywords: AHR; Angiotensin II; Cardiac hypertrophy; Fibrosis; HIF-1α; PPARα; Vascular endothelial growth factor.

Fig. 1

Effects of administration of Ang II on a body weight, b blood pressure, c heart weight, and d LV weight in wild-type and Ahr^−/− mice treated with low- or high-dose of Ang II. Data are mean ± SEM of eight mice per group. *P < 0.05 vs. wild-type control mice; ^#P < 0.05 vs. wild-type Ang II mice

Fig. 2

Effects of administration of Ang II and fenofibrate on a body weight, b blood pressure, c heart weight, and d LV weight in wild-type and Ahr^−/− mice treated with Ang II (at 100 ng/ kg/min) and fenofibrate. Data are mean ± SEM of eight mice per group. *P < 0.05 vs. wild-type control mice; ^#P < 0.05 vs. wild-type Ang II mice; ^†P < 0.05 vs. Ahr^−/− Ang II mice

Fig. 3

Expression levels of a Anp, b Bnp, c Colla1, d Col3a1, and e Ahr mRNAs in the left ventricle of wild-type and Ahr^−/− mice treated with Ang II (at 100 ng/kg/min) and fenofibrate. The expression levels were determined by quantitative RT-PCR analysis and expressed relative to the level of Actb mRNA. Data are mean ± SEM of eight mice per group. *P < 0.05 vs. wild-type control mice; ^#P < 0.05 vs. wild-type Ang II mice; ^†P < 0.05 vs. Ahr^−/− Ang II mice

Fig. 4

Light micrographs of a myocytes in hematoxylin–eosinstained sections and c interstitial and e perivascular fibrosis in Sirius red-stained sections of the LV wall in representative wild-type and Ahr^−/− mice treated with Ang II and fenofibrate. Scale bars, 100 μm. Quantitative analysis of b myocyte cross-sectional area, c interstitial fibrosis, and f perivascular fibrosis and in the left ventricle of wild-type and Ahr^−/− mice treated with Ang II and fenofibrate. Data are mean ± SEM of six mice per group. *P < 0.05 vs. wild-type control mice; ^#P < 0.05 vs. wild-type Ang II mice; ^†P < 0.05 vs. Ahr^−/− Ang II mice

Fig. 5

Expression levels of a endothelin-1 and b c-Jun in the left ventricle of wild-type and Ahr^−/− mice treated with Ang II (100 ng/kg/min) and fenofibrate. The expression levels were determined by quantitative RT-PCR analysis and expressed relative to the level of Actb mRNA. The activities of c endothelin-1 and d c-Jun in the left ventricle were also determined by ELISA. Data are mean ± SEM of eight mice per group. *P < 0.05 vs. wild-type control mice; ^#P < 0.05 vs. wild-type Ang II mice; ^§P < 0.05 vs. Ahr^−/− control mice; ^†P < 0.05 vs. Ahr^−/− Ang II mice

Fig. 6

Representative blots of a immunoblot analysis of the amounts of HIF-1α in nuclear extracts from LV tissues and VEGF in protein extracts from homogenates of LV tissues of wild-type and Ahr^−/− mice treated with Ang II and fenofibrate. b Capillary detected by immunohistochemical staining with anti-CD31 antibody in the LV wall. Scale bars, 100 μm. c Quantitative analysis of the capillary density in the left ventricle of wild-type and Ahr^−/− mice treated with Ang II and fenofibrate. Data are mean ± SEM of six mice per group. *P < 0.05 vs. wild-type control mice; ^#P < 0.05 vs. wild-type Ang II mice; ^†P < 0.05 vs. Ahr^−/− Ang II mice