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The AhR: A regulator of liver fibrosis?
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Abstracts / Toxicology Letters 258S (2016) S39–S53
Workshop W09: Deciphering the role of the aryl hydrocarbon receptor in toxicity and its emerging functions in physiology
W09-2 The AhR: A regulator of liver fibrosis?
W09-1 Insights into novel functions of the dioxin receptor in cell differentiation and pluripotency
Xavier Coumoul 1,∗ , Stéphane Pierre 2 , Aline Chevallier 2 , Fatima Teixeira Clerc 3 , Ariane Ambolet Camoit 2 , Linh Chi Bui 2 , Anne Sophie Bats 4 , Jean Christophe Fournet 5 , Pedro Fernandez Salguero 6 , Robert Barouki 7 , Sophie Lotersztajan 3 , Martine Aggerbeck 1
Pedro M. Fernández Salguero 1,∗ , Nuria Moreno Marin 1 , Antonio Morales Hernandez 1 , Ana Nacarino Palma 1 , Beroe Paniagua 1 , Ascensión Infante Campos 2 , Aurea Gomez Duran 2 , Inmaculada Catalina Fernández 2 , Jaime M. Merino 1 1
Department of Biochemistry and Molecular Biology, Faculty of Sciences, University of Extremadura, Badajoz, Spain 2 Department of Pathology, Infanta Cristina University Medical Center, Badajoz, Spain The aryl hydrocarbon/dioxin receptor (AhR) is known for its critical role in the toxic and carcinogenic response to dioxin and, more recently, for its contribution to the homeostasis of the hepatic, vascular, reproductive and immune systems. AhR is required to maintain proper numbers of undifferentiated cancer stem-like cells in melanoma and human embryonic carcinoma. Here, we have used wild type and AhR-null mice to investigate how AhR affects the regenerative potential of the liver and lungs following acute exposure to carbon tetrachloride (Cl4C) or naphthalene (NPH), respectively. We also analyzed the long-term response of the liver to the carcinogen dimethylnitrosamine (DEN). In the lungs, NPH exhausted the bronchoalveolar epithelium in both genotypes. AhRnull mice restored basal conditions earlier than wild type mice and such process was associated with increased cell proliferation, higher numbers of CK14+ and Clara cells and enhanced expression of OCT4. In the liver, Cl4C damaged the parenchyma and induced necrosis in both AhR genotypes although lack of AhR accelerated regeneration by increasing proliferation and the number of liver stem-like Oval cells. These results suggest that, upon exposure to a toxic insult, AhR deficiency upregulates undifferentiated/stemlike cells in the liver and lung, likely accelerating regeneration. The fact that AhR-null mice treated with DEN developed massive liver tumors with respect to wild type mice, further supports that AhR deficiency promotes an undifferentiated status prone to tumor progression. We propose that AhR negatively regulates the generation and/or maintenance of undifferentiated cells presumed critical for tissue regeneration after toxin-induced damage. http://dx.doi.org/10.1016/j.toxlet.2016.06.1279
1
INSERM UMR-S 1124, Toxicologie Pharmacologie et Signalisation Cellulaire, Paris, France 2 Université Paris Descartes, Sorbonne Paris Cité, Paris, France 3 IRMB, INSERM U955, Hopital Henri Mondor, 94010 Creteil, France 4 AP-HP, Hôpital Européen Georges Pompidou, Service de Chirurgie Gynécologique Cancérologique, Paris, France 5 AP-HP, Hôpital Necker-Enfants Malades, Service d’Anatomo-Pathologie, Paris, France 6 Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias, Universidad de Extremadura, Badajoz, Spain 7 AP-HP, Hôpital Necker-Enfants Malades, Service de Biochimie Métabolique, Paris, France The contribution of pollutants to liver fibrosis is an important and poorly explored issue. In vitro studies suggest that 2,3,7,8tetrachlorodibenzo-p-dioxin (TCDD) and other aryl hydrocarbon receptor (AhR) ligands induce genes that are known to be upregulated during liver fibrosis. Our aim was to determine whether exposure to such pollutants can lead to liver fibrosis and to characterize the mechanisms of action. Mice were treated once a week with 25 g/kg of TCDD. Gene and protein expression as well as liver histology were investigated for each treatment. Treatment of mice with TCDD for 2 weeks modified the hepatic expression of markers of fibrosis (collagen-1A1 and ␣-smooth muscle actin). This is not observed in AhR knockout mice. Following 6 weeks of treatment, histological features of murine hepatic fibrosis became apparent. In parallel, the levels of inflammatory cytokines and markers of activated fibroblasts were found to be upregulated. Interestingly, we also found increased expression of genes of the TGF- pathway and a concomitant decrease of miR-200a levels. Because the transcription factors of the Snail family were shown to be involved in liver fibrosis, we studied their regulation by TCDD. Two members of the Snail family were increased, whereas their negative targets, the epithelial marker E-cadherin and Claudin 1, were decreased. Further, the expression of mesenchymal markers was increased. Finally, we confirmed that Snai2 is a direct transcriptional target of TCDD in the human hepatocarcinoma cell line, HepG2. http://dx.doi.org/10.1016/j.toxlet.2016.06.1280 W09-3 Small immune-modulating molecules interacting with the AhR system Dieter Schrenk Food Chemistry and Toxicology, University of Kaiserslautern, Germany The role of the arylhydrocarbon-receptor (AhR) as a modulator of immune and defense functions is an emerging issue. Kynurenine was identified as the first endogenous prototype modulator which can suppress certain immune functions via the AhR. The secretion of kynurenine by tumor cells was suggested to play a role in the resistance of certain tumors towards the attack of the immune system. In addition, the AhR was suggested to be part of a defense system towards exogenous factors e.g. in the skin. Like-
Abstracts / Toxicology Letters 258S (2016) S39–S53
Workshop W09: Deciphering the role of the aryl hydrocarbon receptor in toxicity and its emerging functions in physiology
W09-2 The AhR: A regulator of liver fibrosis?
W09-1 Insights into novel functions of the dioxin receptor in cell differentiation and pluripotency
Xavier Coumoul 1,∗ , Stéphane Pierre 2 , Aline Chevallier 2 , Fatima Teixeira Clerc 3 , Ariane Ambolet Camoit 2 , Linh Chi Bui 2 , Anne Sophie Bats 4 , Jean Christophe Fournet 5 , Pedro Fernandez Salguero 6 , Robert Barouki 7 , Sophie Lotersztajan 3 , Martine Aggerbeck 1
Pedro M. Fernández Salguero 1,∗ , Nuria Moreno Marin 1 , Antonio Morales Hernandez 1 , Ana Nacarino Palma 1 , Beroe Paniagua 1 , Ascensión Infante Campos 2 , Aurea Gomez Duran 2 , Inmaculada Catalina Fernández 2 , Jaime M. Merino 1 1
Department of Biochemistry and Molecular Biology, Faculty of Sciences, University of Extremadura, Badajoz, Spain 2 Department of Pathology, Infanta Cristina University Medical Center, Badajoz, Spain The aryl hydrocarbon/dioxin receptor (AhR) is known for its critical role in the toxic and carcinogenic response to dioxin and, more recently, for its contribution to the homeostasis of the hepatic, vascular, reproductive and immune systems. AhR is required to maintain proper numbers of undifferentiated cancer stem-like cells in melanoma and human embryonic carcinoma. Here, we have used wild type and AhR-null mice to investigate how AhR affects the regenerative potential of the liver and lungs following acute exposure to carbon tetrachloride (Cl4C) or naphthalene (NPH), respectively. We also analyzed the long-term response of the liver to the carcinogen dimethylnitrosamine (DEN). In the lungs, NPH exhausted the bronchoalveolar epithelium in both genotypes. AhRnull mice restored basal conditions earlier than wild type mice and such process was associated with increased cell proliferation, higher numbers of CK14+ and Clara cells and enhanced expression of OCT4. In the liver, Cl4C damaged the parenchyma and induced necrosis in both AhR genotypes although lack of AhR accelerated regeneration by increasing proliferation and the number of liver stem-like Oval cells. These results suggest that, upon exposure to a toxic insult, AhR deficiency upregulates undifferentiated/stemlike cells in the liver and lung, likely accelerating regeneration. The fact that AhR-null mice treated with DEN developed massive liver tumors with respect to wild type mice, further supports that AhR deficiency promotes an undifferentiated status prone to tumor progression. We propose that AhR negatively regulates the generation and/or maintenance of undifferentiated cells presumed critical for tissue regeneration after toxin-induced damage. http://dx.doi.org/10.1016/j.toxlet.2016.06.1279
1
INSERM UMR-S 1124, Toxicologie Pharmacologie et Signalisation Cellulaire, Paris, France 2 Université Paris Descartes, Sorbonne Paris Cité, Paris, France 3 IRMB, INSERM U955, Hopital Henri Mondor, 94010 Creteil, France 4 AP-HP, Hôpital Européen Georges Pompidou, Service de Chirurgie Gynécologique Cancérologique, Paris, France 5 AP-HP, Hôpital Necker-Enfants Malades, Service d’Anatomo-Pathologie, Paris, France 6 Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias, Universidad de Extremadura, Badajoz, Spain 7 AP-HP, Hôpital Necker-Enfants Malades, Service de Biochimie Métabolique, Paris, France The contribution of pollutants to liver fibrosis is an important and poorly explored issue. In vitro studies suggest that 2,3,7,8tetrachlorodibenzo-p-dioxin (TCDD) and other aryl hydrocarbon receptor (AhR) ligands induce genes that are known to be upregulated during liver fibrosis. Our aim was to determine whether exposure to such pollutants can lead to liver fibrosis and to characterize the mechanisms of action. Mice were treated once a week with 25 g/kg of TCDD. Gene and protein expression as well as liver histology were investigated for each treatment. Treatment of mice with TCDD for 2 weeks modified the hepatic expression of markers of fibrosis (collagen-1A1 and ␣-smooth muscle actin). This is not observed in AhR knockout mice. Following 6 weeks of treatment, histological features of murine hepatic fibrosis became apparent. In parallel, the levels of inflammatory cytokines and markers of activated fibroblasts were found to be upregulated. Interestingly, we also found increased expression of genes of the TGF- pathway and a concomitant decrease of miR-200a levels. Because the transcription factors of the Snail family were shown to be involved in liver fibrosis, we studied their regulation by TCDD. Two members of the Snail family were increased, whereas their negative targets, the epithelial marker E-cadherin and Claudin 1, were decreased. Further, the expression of mesenchymal markers was increased. Finally, we confirmed that Snai2 is a direct transcriptional target of TCDD in the human hepatocarcinoma cell line, HepG2. http://dx.doi.org/10.1016/j.toxlet.2016.06.1280 W09-3 Small immune-modulating molecules interacting with the AhR system Dieter Schrenk Food Chemistry and Toxicology, University of Kaiserslautern, Germany The role of the arylhydrocarbon-receptor (AhR) as a modulator of immune and defense functions is an emerging issue. Kynurenine was identified as the first endogenous prototype modulator which can suppress certain immune functions via the AhR. The secretion of kynurenine by tumor cells was suggested to play a role in the resistance of certain tumors towards the attack of the immune system. In addition, the AhR was suggested to be part of a defense system towards exogenous factors e.g. in the skin. Like-