Robert Barouki

Developmental origins of non-communicable disease: Implications for research and public health.

This White Paper highlights the developmental period as a plastic phase, which allows the organism to adapt to changes in the environment to maintain or improve reproductive capability in part through sustained health. Plasticity is more prominent prenatally and during early postnatal life, i.e., during the time of cell differentiation and specific tissue formation. These developmental periods are highly sensitive to environmental factors, such as nutrients, environmental chemicals, drugs, infections and other stressors. Nutrient and toxicant effects share many of the same characteristics and reflect two sides of the same coin. In both cases, alterations in physiological functions can be induced and may lead to the development of non-communicable conditions. Many of the major diseases – and dysfunctions – that have increased substantially in prevalence over the last 40 years seem to be related in part to developmental factors associated with either nutritional imbalance or exposures to environmental chemicals. The Developmental Origins of Health and Disease (DOHaD) concept provides significant insight into new strategies for research and disease prevention and is sufficiently robust and repeatable across species, including humans, to require a policy and public health response. This White Paper therefore concludes that, as early development (in utero and during the first years of postnatal life) is particularly sensitive to developmental disruption by nutritional factors or environmental chemical exposures, with potentially adverse consequences for health later in life, both research and disease prevention strategies should focus more on these vulnerable life stages.

The aryl hydrocarbon receptor, more than a xenobiotic-interacting protein

The aryl hydrocarbon (dioxin) receptor (AhR) has been studied for several decades largely because of its critical role in xenobiotic‐induced toxicity and carcinogenesis. Albeit this is a major issue in basic and clinical research, an increasing number of investigators are turning their efforts to try to understand the physiology of the AhR under normal cellular conditions. This is an exciting area that covers cell proliferation and differentiation, endogenous mechanisms of activation, gene regulation, tumor development and cell motility and migration, among others. In this review, we will attempt to summarize the studies supporting the implication of the AhR in those endogenous cellular processes.

Dietary Polyphenols Increase Paraoxonase 1 Gene Expression by an Aryl Hydrocarbon Receptor-Dependent Mechanism

ABSTRACT Human paraoxonase 1 (PON-1) is a serum high-density lipoprotein-associated enzyme mainly secreted by the liver. It has endogenous and exogenous substrates and displays protective properties with respect to cardiovascular disease and organophosphate intoxication. In the HuH7 human hepatoma cell line, PON-1 activity and mRNA levels were increased by dietary polyphenolic compounds such as quercetin but also by toxic ligands of the aryl hydrocarbon receptor (AhR) such as 3-methylcholanthrene (3-MC). However, the 2,3,7,8-tetrachlorobenzo(p)dioxin (TCDD) was a poor inducer. Transient and stable transfection assays indicated that these compounds increased the PON-1 gene promoter activity in an AhR-dependent manner, since their effect was inhibited by 7-keto-cholesterol and AhR-directed short interfering RNA. Deletions and mutations studies showed that a xenobiotic responsive element (XRE)-like sequence within the PON-1 promoter mediated the effect of 3-MC and quercetin. In contrast with consensus XREs from the cytochrome P450 1A1 gene, the PON-1 XRE-like element mediated preferentially the effect of quercetin compared to the results seen with TCDD. Furthermore, AhR binding to this element was preferentially activated by quercetin. These observations provide a molecular mechanism for the regulation of the cardioprotective enzyme PON-1 by polyphenols. They suggest also that AhR ligands may differentially regulate gene expression depending on the DNA target sequence.

Down-regulation of Cytochrome P450 1A1 Gene Promoter by Oxidative Stress CRITICAL CONTRIBUTION OF NUCLEAR FACTOR 1

Oxidative stress interferes with several cellular functions, in particular transcriptional regulation. We show here that the human cytochrome P450 1A1 (CYP1A1) is down-regulated at the transcriptional level by oxidative stress. Basal as well as 2,3,7,8-tetrachloro-p-dioxin-induced promoter activities are strongly impaired by H2O2 treatment or glutathione depletion with l-buthionine-(S,R)-sulfoximine. Tumor necrosis factor α inhibits CYP1A1 expression, and this inhibition is prevented by the antioxidant pyrrolidine dithiocarbamate. We show that these regulations depend on the integrity of the nuclear factor 1 (NFI) site located in the proximal promoter. We therefore examined the redox regulation of this transcription factor. Treatment of human HepG2 or rat H4 hepatoma cells with H2O2 orl-buthionine-(S,R)-sulfoximine inactivates the binding of the NFI transcription factor to its DNA consensus sequence. Furthermore, H2O2 treatment leads to a dose-dependent decrease of reporter gene expressions driven by promoters containing NFI binding sites. Glutathione depletion and catalase inhibition also repress a NFI-driven promoter. Under the same conditions, the CP-1 transcription factor activity is not affected by oxidative stress. Thus, NFI seems particularly sensitive to oxidative stress. This accounts, at least partially, for the regulation ofcyp1A1 gene expression.

Repression of cytochrome P450 1A1 gene expression by oxidative stress: mechanisms and biological implications

Cytochrome P450 1A1 (CYP1A1) is a member of a multigenic family of xenobiotic-metabolizing enzymes. Beyond its usual role in the detoxification of polycyclic aromatic compounds, the activity of this enzyme can be deleterious since it can generate mutagenic metabolites and oxidative stress. The CYP1A1 gene is highly inducible by the environmental contaminants dioxin and benzo[a]pyrene. We discuss here the regulatory mechanisms that limit this induction. Several feedback loops control the activation of this gene and the subsequent potential toxicity. The oxidative repression of the CYP1A1 gene seems to play a central role in these regulations. The transcription factor Nuclear Factor I/CCAAT Transcription Factor (NFI/CTF), which is important for the transactivation of the CYP1A1 gene promoter, is particularly sensitive to oxidative stress. A critical cysteine within the transactivating domain of NFI/CTF appears to be the target of H(2)O(2). The DNA-binding domains of several transcription factors have been described as targets of oxidative stress. However, recent studies described here suggest that more attention should be given to transactivating domains that may represent biologically relevant redox targets of cellular signaling.

Fate and Complex Pathogenic Effects of Dioxins and Polychlorinated Biphenyls in Obese Subjects before and after Drastic Weight Loss

Background In humans, persistent organic pollutants (POPs) are stored primarily in adipose tissue. Their total body burden and their contribution to obesity-associated diseases remain unclear. Objectives We characterized POP total body burden and their redistribution in obese individuals before and after drastic weight loss and compared these values with a variety of molecular, biological, and clinical parameters. Methods Seventy-one obese subjects were enrolled and underwent bariatric surgery. Blood and adipose tissue samples were obtained at different times from these individuals as well as from 18 lean women. Results POP content (17 dioxins/furans and 18 polychlorinated biphenyl congeners) in different adipose tissue territories was similar, allowing us to assess total POP body burden from a single biopsy. Total POP body burden was 2 to 3 times higher in obese than in lean individuals. We also found increased expression of some POP target genes in obese adipose tissue. Drastic weight loss led to increased serum POPs and, within 6–12 months, to a significant 15% decrease in total polychlorinated biphenyl body burden. Importantly, serum POP levels were positively correlated with liver toxicity markers and lipid parameters, independently of age and body mass index. Conclusions POP content in adipose tissue and serum correlate with biological markers of obesity-related dysfunctions. Drastic weight loss leads to a redistribution of POPs and to a moderate decrease of their total body burden.

Toxicological Function of Adipose Tissue: Focus on Persistent Organic Pollutants

Background: Adipose tissue (AT) is involved in several physiological functions, including metabolic regulation, energy storage, and endocrine functions. Objectives: In this review we examined the evidence that an additional function of AT is to modulate persistent organic pollutant (POP) toxicity through several mechanisms. Methods: We reviewed the literature on the interaction of AT with POPs to provide a comprehensive model for this additional function of AT. Discussion: As a storage compartment for lipophilic POPs, AT plays a critical role in the toxicokinetics of a variety of drugs and pollutants, in particular, POPs. By sequestering POPs, AT can protect other organs and tissues from POPs overload. However, this protective function could prove to be a threat in the long run. The accumulation of lipophilic POPs will increase total body burden. These accumulated POPs are slowly released into the bloodstream, and more so during weight loss. Thus, AT constitutes a continual source of internal exposure to POPs. In addition to its buffering function, AT is also a target of POPs and may mediate part of their metabolic effects. This is particularly relevant because many POPs induce obesogenic effects that may lead to quantitative and qualitative alterations of AT. Some POPs also induce a proinflammatory state in AT, which may lead to detrimental metabolic effects. Conclusion: AT appears to play diverse functions both as a modulator and as a target of POPs toxicity.

Activation of the dioxin/aryl hydrocarbon receptor (AhR) modulates cell plasticity through a JNK-dependent mechanism

Environmental chemicals such as dioxin adversely affect immune, neurological and reproductive functions and have been implicated in cancer development. However, the mechanisms responsible for dioxin toxicity are still poorly understood. Here, we show that dioxin and related pollutants trigger a marked morphological change in epithelial cells that remodel their cytoskeleton to increase interaction with extra cellular matrix while loosening cell–cell contacts. Furthermore, dioxin-treated cells show increased motility. These dioxin-mediated effects are mimicked by constitutive expression and activation of the intracellular dioxin receptor (aryl hydrocarbon receptor (AhR)). They correlate with activation of the Jun NH2-terminal kinase (JNK) and are reverted by treatment with a JNK inhibitor. Dioxin-induced effects occur 48 h post-treatment initiation, a time scale, which argues for a genomic effect of the AhR, linked to induction of target genes. This novel Ahr action on cell plasticity points to a role in cancer progression.

Inflammatory pathway genes belong to major targets of persistent organic pollutants in adipose cells.

Background: Epidemiological studies emphasize the possible role of persistent organic pollutants (POPs) in obesity and the metabolic syndrome. These pollutants are stored in adipose tissue (AT). Objectives: Our aim was to study the effects of POPs on human adipose cells and rodent AT. Methods: Using human multipotent adipose-derived stem cells, we carried out large-scale gene expression analysis to identify the major pathways modified by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), polychlorinated biphenyl (PCB) congener 126 (PCB-126), and PCB-153 and to evaluate their toxic effects. The effects of TCDD on gene expression and AT histology were also assessed in mice. Results: The most significantly regulated genes in both precursor cells and adipocytes were those involved in the inflammatory/immune response, cancer, and metabolism pathways. Interestingly, the fold induction and the number of modulated genes were higher in precursors than in adipocytes, suggesting that the former could be more sensitive to the effect of pollutants. When cells were treated with combinations of pollutants, the effects of the AhR ligands TCDD and PCB-126 were dominant compared with those of the non-dioxin-like PCB-153. The effects of AhR ligands were reduced by the AhR antagonist α-naphthoflavone. The regulation of inflammatory pathway was observed in wild-type AT but not in AhR-knockout mice. Conclusions: Both in vitro and in vivo studies showed that adipose cells were targets of AhR ligands and suggest that inflammation is one of the main regulated pathways. These observations suggest a possible contribution of pollutants to low-grade AT inflammation that accompanies the pathogenesis of metabolic diseases.

An Autoregulatory Loop Controlling CYP1A1 Gene Expression: Role of H 2 O 2 and NFI

ABSTRACT Cytochrome P450 1A1 (CYP1A1), like many monooxygenases, can produce reactive oxygen species during its catalytic cycle. Apart from the well-characterized xenobiotic-elicited induction, the regulatory mechanisms involved in the control of the steady-state activity of CYP1A1 have not been elucidated. We show here that reactive oxygen species generated from the activity of CYP1A1 limit the levels of induced CYP1A1 mRNAs. The mechanism involves the repression of theCYP1A1 gene promoter activity in a negative-feedback autoregulatory loop. Indeed, increasing the CYP1A1 activity by transfecting CYP1A1 expression vectors into hepatoma cells elicited an oxidative stress and led to the repression of a reporter gene driven by the CYP1A1 gene promoter. This negative autoregulation is abolished by ellipticine (an inhibitor of CYP1A1) and by catalase (which catalyzes H2O2 catabolism), thus implying that H2O2 is an intermediate. Down-regulation is also abolished by the mutation of the proximal nuclear factor I (NFI) site in the promoter. The transactivating domain of NFI/CTF was found to act in synergy with the arylhydrocarbon receptor pathway during the induction of CYP1A1 by 2,3,7,8-tetrachloro-p-dibenzodioxin. Using an NFI/CTF-Gal4 fusion, we show that NFI/CTF transactivating function is decreased by a high activity of CYP1A1. This regulation is also abolished by catalase or ellipticine. Consistently, the transactivating function of NFI/CTF is repressed in cells treated with H2O2, a novel finding indicating that the transactivating domain of a transcription factor can be targeted by oxidative stress. In conclusion, an autoregulatory loop leads to the fine tuning of theCYP1A1 gene expression through the down-regulation of NFI activity by CYP1A1-based H2O2 production. This mechanism allows a limitation of the potentially toxic CYP1A1 activity within the cell.