Global gene expression profiling in human lung cells exposed to cobalt.

International audienceBACKGROUND: It has been estimated that more than 1 million workers in the United States are exposed to cobalt. Occupational exposure to 59 Co occurs mainly via inhalation and leads to various lung diseases. Cobalt is classified by the IARC as a possible human carcinogen (group 2B). Although there is evidence for in vivo and in vitro toxicity, the mechanisms of cobalt-induced lung toxicity are not fully known. The purpose of this work was to identify potential signatures of acute cobalt exposure using a toxicogenomic approach. Data analysis focused on some cellular processes and protein targets that are thought to be relevant for carcinogenesis, transport and biomarker research. RESULTS: A time course transcriptome analysis was performed on A549 human pulmonary cells, leading to the identification of 85 genes which are repressed or induced in response to soluble 59 Co. A group of 29 of these genes, representing the main biological functions, was assessed by quantitative RT-PCR. The expression profiles of six of them were then tested by quantitative RT-PCR in a time-dependent manner and three modulations were confirmed by Western blotting. The 85 modulated genes include potential cobalt carriers (FBXL2, ZNT1, SLC12A5), tumor suppressors or transcription factors (MAZ, DLG1, MYC, AXL) and genes linked to the stress response (UBC, HSPCB, BNIP3L). We also identified nine genes coding for secreted proteins as candidates for biomarker research. Of those, TIMP2 was found to be down-regulated and this modulation was confirmed, in a dose-dependent manner, at protein level in the supernatant of exposed cells. CONCLUSION: Most of these genes have never been described as related to cobalt stress and provide original hypotheses for further study of the effects of this metal ion on human lung epithelial cells. A putative biomarker of cobalt toxicity was identified

Caracterisation et purification a l'aide d'un anticorps monoclonal, d'un nouveau marqueur d'activation des lymphocytes T et B murins

SIGLEINIST T 73224 / INIST-CNRS - Institut de l'Information Scientifique et TechniqueFRFranc

Overview of the TRANSAT (TRANSversal Actions for Tritium) project

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Poorly soluble cobalt oxide particles trigger genotoxicity via multiple pathways

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Bioaccumulation and biological effects of hydrogenated cement particles in the marine bivalve, Mytilus galloprovincialis

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Low Doses of Gamma-Irradiation Induce an Early Bystander Effect in Zebrafish Cells Which Is Sufficient to Radioprotect Cells

International audienceThe term ''bystander effect'' is used to describe an effect in which cells that have not been exposed to radiation are affected by irradiated cells though various intracellular signaling mechanisms. In this study we analyzed the kinetics and mechanisms of bystander effect and radioadaptation in embryonic zebrafish cells (ZF4) exposed to chronic low dose of gamma rays. ZF4 cells were irradiated for 4 hours with total doses of gamma irradiation ranging from 0.01-0.1 Gy. In two experimental conditions, the transfer of irradiated cells or culture medium from irradiated cells results in the occurrence of DNA double strand breaks in non-irradiated cells (assessed by the number of c-H2AX foci) that are repaired at 24 hours post-irradiation whatever the dose. At low total irradiation doses the bystander effect observed does not affect DNA repair mechanisms in targeted and bystander cells. An increase in global methylation of ZF4 cells was observed in irradiated cells and bystander cells compared to control cells. We observed that pre-irradiated cells which are then irradiated for a second time with the same doses contained significantly less c-H2AX foci than in 24 h gamma-irradiated control cells. We also showed that bystander cells that have been in contact with the pre-irradiated cells and then irradiated alone present less c-H2AX foci compared to the control cells. This radioadaptation effect is significantly more pronounced at the highest doses. To determine the factors involved in the early events of the bystander effect, we performed an extensive comparative proteomic study of the ZF4 secretomes upon irradiation. In the experimental conditions assayed here, we showed that the early events of bystander effect are probably not due to the secretion of specific proteins neither the oxidation of these secreted proteins. These results suggest that early bystander effect may be due probably to a combination of multiple factors

A new procedure for high precision isotope ratio determinations of U, Cu and Zn at nanogram levels in cultured human cells: What are the limiting factors?

The monitoring of isotopic fractionations in in vitro cultured human cell samples is a very promising and under-exploited tool to help identify the metabolic processes leading to disease-induced isotopic fractionations or decipher metabolic pathways of toxic metals in these samples. One of the limitations is that the analytes are often present at small amounts, ranging from tens to hundreds of ng, thus making challenging low-uncertainty isotope ratio determinations. Here we present a new procedure for U, Cu and Zn purification and isotope ratio determinations in cultured human neuron-like cells exposed to natural U. A thorough study of the influence of the limiting factors impacting the uncertainty of δ238U, δ66Zn and δ65Cu is also carried out. These factors include the signal intensity, which determines the within-day measurement reproducibility, the procedural blank correction and the matrix effects, which determine the accuracy of the mass bias correction models. Given the small Cu and U amounts in the cell samples, 15–30 and 20 ng respectively, a highly efficient sample introduction system was employed in order to improve the analyte transport to the plasma and, hence, the signal intensity. With this device, the procedural blanks became the main uncertainty source of δ238U and δ65Cu values, accounting over 65% of the overall uncertainty. The matrix effects gave rise to inaccuracies in the mass bias correction models for samples finally dissolved in the minimal volumes required for the analysis, 100–150 µL, leading to biases for U and Cu. We will show how these biases can be cancelled out by dissolving the samples in volumes of at least 300 µL for Cu and 450 µL for U. Using our procedure, expanded uncertainties (k = 2) of around 0.35‰ for δ238U and 0.15‰ for δ66Zn and δ65Cu could be obtained. The analytical approach presented in this work is also applicable to other biological microsamples and can be extended to other elements and applications

Cobalt chloride speciation, mechanisms of cytotoxicity on human pulmonary cells, and synergistic toxicity with zinc.

Cobalt is used in numerous industrial sectors, leading to occupational diseases, particularly by inhalation. Cobalt-associated mechanisms of toxicity are far from being understood and information that could improve knowledge in this area is required. We investigated the impact of a soluble cobalt compound, CoCl(2)*6H(2)O, on the BEAS-2B lung epithelial cell line, as well as its impact on metal homeostasis. Cobalt speciation in different culture media, in particular soluble and precipitated cobalt species, was investigated via theoretical and analytical approaches. The cytotoxic effects of cobalt on the cells were assessed. Upon exposure of BEAS-2B cells to cobalt, intracellular accumulation of cobalt and zinc was demonstrated using direct in situ microchemical analysis based on ion micro-beam techniques and analysis after cell lysis by inductively coupled plasma mass spectrometry (ICP-MS). Microchemical imaging revealed that cobalt was rather homogeneously distributed in the nucleus and in the cytoplasm whereas zinc was more abundant in the nucleus. The modulation of zinc homeostasis led to the evaluation of the effect of combined cobalt and zinc exposure. In this case, a clear synergistic increase in toxicity was observed as well as a substantial increase in zinc content within cells. Western blots performed under the same coexposure conditions revealed a decrease in ZnT1 expression, suggesting that cobalt could inhibit zinc release through the modulation of ZnT1. Overall, this study highlights the potential hazard to lung function, of combined exposure to cobalt and zinc