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International audience; Although soil characteristics modulate metal mobility and bioavailability to organisms, they are often ignored in the risk assessment of metal transfer. This paper aims to determine the ability of chemical methods to assess and predict cadmium (Cd), lead (Pb) and zinc (Zn) environmental bioavailability to the land snail Cantareus aspersus. Snails were exposed in the laboratory for 28 days to 17 soils from around a former smelter. The soils were selected for their range of pH, organic matter, clay content, and Cd, Pb and Zn concentrations. The influence of soil properties on environmental availability (estimated using HF-HClO(4), EDTA, CaCl(2), NH(4)NO(3), NaNO(3), free ion activity and total dissolved metal concentration in soil solution) and on environmental bioavailability (modelled using accumulation kinetics) was identified. Among the seven chemical methods, only the EDTA and the total soil concentration can be used to assess Cd and Pb environmental bioavailability to snails (r²(adj)=0.67 and 0.77, respectively). For Zn, none of the chemical methods were suitable. Taking into account the influence of the soil characteristics (pH and CEC) allows a better prediction of Cd and Pb environmental bioavailability (r²(adj)=0.82 and 0.83, respectively). Even though alone none of the chemical methods tested could assess Zn environmental bioavailability to snails, the addition of pH, iron and aluminium oxides allowed the variation of assimilation fluxes to be predicted. A conceptual and practical method to use soil characteristics for risk assessment is proposed based on these results. We conclude that as yet there is no universal chemical method to predict metal environmental bioavailability to snails, and that the soil factors having the greatest impact depend on the metal considered.

Transfer and toxic effects of two cadmium (Cd) forms, inorganic (CdCl2 dosed rat food) or organic (contaminated snail-based rat food) were studied in Wistar rat. Cd concentrations in rat food were 0 and 2.5 μg Cd g−1 for both inorganic and organic forms and a high concentration of 100 μg Cd g−1 was also tested for the inorganic form. Rats were exposed for four weeks to contaminated food. Both forms of Cd were bioavailable to rats, with a percentage of transfer from food to rats of around 1% for all contaminated groups. Cd concentrations in rat tissues increased with increasing Cd concentrations in the food. Rats fed with organic form of Cd accumulated significantly more Cd in the main organ for Cd toxicity, the kidney, than those eating the inorganic form. Survival was not affected for any rat group but a decrease in growth and food consumption was observed for the inorganic form. As a defence system against Cd toxicity, rats increased their metallothionein (MT) synthesis at the highest Cd concentration in the target organs (kidney, liver and small intestine) and even did the same at low Cd concentrations (2.5 μg Cd g−1) in the kidney. At this low Cd concentration, MT induction was lower in the small intestine of rats ingesting organic Cd than those ingesting inorganic Cd. Bioavailability of organic and inorganic forms of Cd was similar, but subsequent Cd distribution within organs was different. This quantification of the trophic transfer of both inorganic and organic forms of a toxicant is a basis for a better assessment of the fate and effects of chemicals in food webs. Keywords: Bioavailability, Ecotoxicology, Metallothioneins

International audience; The proper use of bioaccumulation in the assessment of environmental quality involves accounting for chemical fluxes in organisms. Cadmium (Cd) accumulation kinetics in a soil-plant-snail food chain were therefore investigated in the field under different soil contamination (from 0 to 40 mg kg(-1)), soil pH (6 and 7) and season. Allowing for an accurate and sensitive assessment of Cd transfer to snails, toxicokinetics appears an interesting tool in the improvement of risk assessment procedures and a way to quantify metal bioavailability for a defined target. On the basis of uptake fluxes, snails proved to be sensitive enough to distinguish moderate soil contaminations. The soil pH did not appear, in the range studied, as a modulating parameter of the Cd transfer from soil to snail whereas the season, by influencing the snail mass, may modify the internal concentrations. The present data specifying a time integrated assessment of environmental factors on metal bioavailability and transfer to terrestrial snails should ensure their rational use in environmental biomonitoring.

Angiopoietin-like 4 (ANGPTL4) is a secreted protein that belongs to the angiopoietin family and is involved in angiogenesis and metabolism regulation. We previously reported the induction of angptl4 by hypoxia in endothelial cells and in human ischemic tissues from peripheral artery disease. We here observed in a mouse model of hindlimb ischemia that the mRNA upregulation in the vessels correlates with the accumulation of the full-length protein in ischemic tissues. We then investigated its functions in endothelial cells. In response to hypoxia, endogenous ANGPTL4 accumulates in the subendothelial extracellular matrix (ECM). Although the secreted protein undergoes proteolysis leading to truncated fragments present in the medium, only full-length ANGPTL4 interacts with the ECM. Competition and direct binding assays indicate that the strong interaction of ANGPTL4 with the ECM is heparin/heparan sulfate proteoglycan dependent. The balance between matrix-associated and soluble forms of ANGPTL4 points to the role of the ECM in the regulation of its bioavailability. The angiogenic function of the ECM-bound full-length protein was investigated using either the form associated with the conditioned ECM from ANGPTL4-transfected HEK293 cells or the purified immobilized protein. We show that matrix-associated and immobilized ANGPTL4 limit the formation of actin stress fibers and focal contacts in the adhering endothelial cells and inhibit their adhesion. Immobilized ANGPTL4 also decreases motility of endothelial cells and inhibits the sprouting and tube formation. Altogether, these findings show that hypoxic endothelial cells accumulate ANGPTL4 in the ECM, which in turn negatively regulates their angiogenic capacities through an autocrine pathway.

To enlarge the possibilities of using organisms of the soil fauna to assess the bioaccumulative potential of chemicals, the kinetic of soil cadmium (Cd) transfer to the terrestrial gastropod Helix aspersa was investigated under laboratory conditions during a long-term experiment (6 months). During the exposure phase (3 months), juvenile snails were subjected to three different concentrations of Cd spiked in artificial ISO soil (ISO 0, 20 and 100 mg Cd kg−1) and to a field soil (ME4) industrially contaminated by 20 mg Cd kg −1. For both soils, internal steady-state Cd concentrations were reached in the viscera of the snails, the main storage organ for Cd, after 2 weeks of exposure whatever the Cd concentration in soil. The equilibrium concentrations in the viscera were 0.7 (±0.1), 11.3 (±2.4), 73.3 (±4.8) and 6.3 (±1.3) mg Cd kg−1 dry mass for ISO 0, ISO 20, ISO 100 and ME4, respectively. During the depuration phase (3 months), from 0 to 52% of the accumulated Cd in the viscera were removed by excretion or relocation in the foot. However, the snails were not able to depurate down to initial concentrations. Data were modelled by integrating a specific growth rate constant into one-compartment toxicokinetic models. This allowed the calculation of Cd uptake rates that can be used as indicators of metal bioavailability. Since this parameter was found to be lower for snails exposed to the field soil ME4, we concluded that lower Cd bioavailability in this field soil was responsible of the lower transfer to the snails compared to the ISO 20 soil, even though they were polluted to similar extents. Internal validation showed that the toxicokinetic models could be applied for predictive purposes, promising for the development of a bioaccumulation directive for terrestrial environment. Keywords: Accumulation, Depuration, Bioavailability, Modelling, Long-term exposure

International audience; Among soil invertebrates, terrestrial snails are herbivorous and detritivorous organisms exposed to polluted soils by both digestive and cutaneous routes. Using laboratory-reared snails (Helix aspersa aspersa), we describe how the effects of contaminants on survival and growth of snails can be evaluated in laboratory bioassays. A national ring test was performed to assess the effect of Cd added to the soil or to the food. The ecotoxicity of sewage sludge also was evaluated. The present results demonstrate that toxicity depends on both the pollutants and the exposure route. Cadmium was sixfold more toxic for snails exposed via food contamination (median effective concentration [EC50], 68-139 microg/g) than via soil contamination (EC50, 534-877 microg/g), whereas the opposite occurred with the sewage sludge (EC50, 55% of sludge in the food and 10% of waste in the soil). A logistic relationship linked growth inhibition and internal Cd concentrations, which can reach 2,000 microg/g in the viscera of snails exposed to 626 microg/g in the food. No clear trend was found between Cu, Zn, Pb, Cr, and Ni concentrations in the sludge and in snail tissues. These data enabled the development of an international standard, which should enhance the use of terrestrial gastropods for both fundamental research and routine risk assessment in the terrestrial environment.

International audience; Isotopic dilution techniques were initially used to evaluate the bioavailability of trace metals contained in soils to plants, i.e., the phytoavailability. Here, we use for the first time an isotopic technique to evaluate the zooavailability of cadmium (Cd), i.e., the bioavailability to an animal organism. A terrestrial invertebrate, the snail Helix aspersa, was exposed for 14 days to a polluted soil that was spiked with 109Cd. Isotopic composition of snail tissues was then determined, allowing the computing of the L value, which can be considered as a biological estimate of the bioavailable Cd pool in the soil. It showed that the bioavailable pool of Cd to H. aspersa represented 58% of the total soil Cd. The Evalue, a chemical estimate of the soil Cd labile pool, was calculated with short-term isotopic exchange kinetics experiments and a time-dependent model describing the isotopic exchange over time. The E(14 days) value indicated that the labile soil Cd pool represented 49.6% of the total soil Cd. The L value was significantly higher than the E value, showing that snails accessed the nonlabile soil Cd pool, generally considered as nonbioavailable. The nonlabile pool contributed for 16% to the total Cd accumulated by snails. These results showed that the uptake of Cd in the nonlabile pool by soil organisms could be important and should be considered in risk assessment procedures for metal polluted soils.

International audience; Juvenile Helix aspersa snails exposed in field microcosms were used to assess the transfer of Cd, Cu, Ni, Pb and Zn from forest soils amended with liquid and composted sewage sludge. Zn concentrations and contents were significantly higher in snails exposed to liquid and composted sludge after 5 and 7 weeks of exposure, when compared with control. Trends were less clear for the other metals. Present results show that Zn, among the cocktail of metallic trace elements (MTE) coming from sewage sludge disposal, represents the principal concern for food chain transfer and secondary poisoning risks. The microcosm design used in this experiment was well suited for relatively long-term (about 2 months) active biomonitoring with H. aspersa snails. The snails quickly indicated the variations of MTE concentrations in their immediate environment. Therefore, the present study provides a simple but efficient field tool to evaluate MTE bioavailability and transfer.

International audience; Juvenile Helix aspersa snails were exposed for 4 weeks to fresh rape leaves (Brassica napus), contaminated by simulating superficial deposits of increasing concentrations of cadmium (Cd). The Cd concentration in leaves was 0.1 in control and 38.4, 93.1, and 177.2 microg x g(-1) (dry mass) in Cd-treated food. The concentration in snail tissues increased with increasing Cd concentrations in the food. The bioaccumulation factors ranged from 4.8 (control) to 2.4 (highest exposed group), indicating a biomagnification of Cd in this food chain. The growth of the snails treated with Cd was reduced by 17, 24, and 43% respectively, compared to the control group. Comparison of these results with those obtained with snails exposed to similar Cd concentrations in a vegetable flour revealed that accumulation and effects were relatively consistent, demonstrating a comparable bioavailability of Cd in the two diets. Tests using growing H. aspersa snails exposed to metals in flour or fresh leaves can be useful for risk assessment purposes.