The Sensitivity Difference Of Avian Species To The Toxicity Of Dioxin-like Organic Pollutants

For the past half century scientists, scientific regulators, industry and the public have been concerned with the environmental pollution and adverse effects on human and animals induced by dioxins, dioxin-like compounds (DLCs) and other organic pollutants. The sensitivity to toxic effects of dioxins and DLCs varies among different avian species, even with a large difference of up to a thousand times. Predicting effectively the sensitivity difference to toxicity of dioxins and DLCs will greatly reduce the uncertainty of ecological risk assessment. It will consume large amounts of live birds or fertile eggs for obtaining toxicity data from traditional toxicology tests by feeding or egg injection. This approach not only is contrary to ethics, but also brings huge workload to scientists. And it encounters inevitably the challenges of working with rare or endangered species. New commodity chemicals have been constantly emerging, which increases scientists’burden for toxicity screening and ecological risk assessment. A rapid, highly-efficient and all avian species-applicable alternative is needed instead of animal testing.The recent studies indicate that the difference in sensitivity among avian species to dioxin-like toxicity is based upon the identities of the amino acids at two sites324and380within the ligand binding domain (LBD) of aryl hydrocarbon receptor1(AHR1). Based on these sequences, and data from in vitro and egg injection studies, birds could be grouped into three general classes of sensitivity to DLCs:type1(chicken-like; Ile324_Ser380), type2(ring-necked pheasant [Phasianus colchicus]-like; Ile324_Ala380), or type3(Japanese quail [Coturnix japonica]-like; Val324_Ala380). Accordingly, one novel avian AHRl-luciferase reporter gene (LRG) assay was developed to predict the species-specific dioxin-like toxicity of one single dioxin or DLC in birds, and the sensitivity difference to the toxicity for birds.Therefore, the present study was carried out in the following aspects:1) To test whether World Health Organization (WHO)-toxic equivalency factors (TEFs) which are commonly used in avian ecological risk assessment of dioxins and DLCs may be representative of relative dioxin-like potency of dioxins and DLCs in all avian species using a recently developed avian AHR1-LRG assay; and to determine whether the LRG assay is a useful tool for predicting the avian species-specific toxicity of DLC mixtures and relative sensitivity difference to the toxicity, in addition to individual dioxin and DLC;2) To determine the avian species-specific toxicity of18polychlorinated diphenyl sulfides (PCDPSs) and19hydroxylated/methoxylated polybrominated diphenyl ethers (HO-/MeO-PBDEs), and relative sensitivity difference of avian species to the toxicity;3) From the perspective of biophysics, some OH-/MeO-PBDEs were taken for example to explore preliminarily the reason why the avian sensitivity difference to the dioxin-like toxicity of some DLCs is dissimilar to that of typical dioxins (such as2,3,7,8-tetrachlorodibenzo-p-dioxin) in the atomic level. The main conclusions are shown as follows.1) The study of Aroclors showed that the six tested Aroclors could be divided into two groups:high potency Aroclors (1248,1242, and1254) and low potency Aroclors (1260,1016, and1221). Bioassay-derived TEQs for the LRG and Ethoxyresorufin O-deethylase (EROD) assays in the chicken were significantly correlated with TEQs for Aroclors calculated using WHO-TEFs. Differences between the calculated and chicken bioassay-derived TEQs for each Aroclor were within1order of magnitude. This is consistent with WHO-TEFs being derived primarily from chicken toxicity data. The results indicate that the LRG assay is a useful tool for evaluating the toxicity of dioxins and DLCs mixtures in avian species, in addition to individual dioxin and DLC. However, LRG-TEQs for the Japanese quail AHR1were up to2orders of magnitude higher than the calculated TEQs for certain Aroclors, suggesting that the WHO-TEFs may not be representative of relative potency of dioxins and DLCs in all avian species. These results have important implications for the general application of the WHO-TEFs in estimating the embryolethal effects of environmental DLC mixtures to wild avian species. While the WHO-TEF values can be used to estimate toxicity of DLC mixtures to the chicken and other type1species (Ile324_Ser380), they may at times over-or underestimate dioxin-like toxicity for type2(Ile324_Ala380) and type3(Val324_Ala380) species, depending on the composition of the mixture.2) The results of the study of PCDPSs and OH-/MeO-PBDEs indicate that they could induce very different dioxin-like toxicity via activating AHR1-mediated molecular toxicological pathways. And parts of them have fair or even higher dioxin-like toxicity than OctaCDD, OctaCDF and dioxin-like PCBs. The derived avian species-specific relative potency (ReP) values of PCDPSs suggested that the ReP values increased with the number of substituted Cl atoms increases for all the three broad categories of birds (type1(Ile324_Ser380), type2(Ile324_Ala380) and type3(Val324_Ala380)). Relative sensitivity (ReS) values derived indicated that the avian sensitivity order was different from that induced by TCDD for some of the PCDPSs and OH-/MeO-PBDEs. This may be because differences of amino acid sequence in the AHR1-LBD combined with differences in the structure of the ligand compounds lead to differences in Iigand-receptor conformation, thereby resulting in varied co-activator recruitment and interactions with xenobiotic response elements. RePavg values of PCDPSs and OH-/MeO-PBDEs derived from the avian AHR1s-LRG and CEH-EROD assays were much greater (even up to4orders of magnitude) than that from the H4IIE-luc (recombinant rat hepatoma cells) assays, which was consistent with the case of dioxins and DLCs. This indicated that it was inappropriate to use H4IIE-luc assays to estimate the TEQ value of the extraction of dioxins and DLC mixtures from environmental samples to conduct risk assessment for birds, which tends to at times underestimate the dioxin-like toxicity. Therefore, parts of PCDPSs and OH-/MeO-PBDEs should be ranked as potential high priority environmental pollutants and novel DLCs given their considerable persistence and environmental mobility reported previously as well as the dioxin-like toxicity demonstrated in the present study.3) The results of molecular dynamics simulation of AHR1-LBDs and OH-/MeO-PBDEs complexes indicated that amino acid at site324within the LBD of AHR1could affect the conformation change of subsequent No.4helix connected to it, thereby affecting AHR1activation. Amino acid at site324within the LBD of AHR1is valine for Japanese quail, while it is isoleucine for chicken and ring-necked pheasant instead. The results of molecular dynamics simulation indicated that the steric hindrance of valine is bigger than that of isoleucine, which results in bigger rotational energy and greater stability for the subsequent No.4helix. This may be the reason why the AHR1of Japanese quail is more likely to be activated than chicken’s and ring-necked pheasant’s when exposed to some OH-/MeO-PBDEs, which then leads to that the avian sensitivity difference to the dioxin-like toxicity of the OH-/MeO-PBDEs is dissimilar to that of TCDD. For chicken and ring-necked pheasant, the amino acid at site324within AHR1-LBD is the same, but the amino acid at site380is different (Serine in chicken; alanine in ring-necked pheasant). Because the380site amino acid residue is far from the No.4helix in spatial position, it is speculated that the difference of the380site amino acid residue leads to the variance of interaction between avian AHRls and ligand, thereby influencing the movement of No.4helix which is mainly determined by the amino acid at site324. It may be the reason why the situation of the avian sensitivity difference to dioxin-like toxicity of organic pollutants is complex and varied. But whether this rule could be used for other ligand compounds still needs further study.