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In vitro immune toxicity of depleted uranium: effects on murine macrophages, CD4+ T cells, and gene expression profiles.

ABSTRACT: Depleted uranium (DU) is a by-product of the uranium enrichment process and shares chemical properties with natural and enriched uranium. To investigate the toxic effects of environmental DU exposure on the immune system, we examined the influences of DU (in the form of uranyl nitrate) on viability and immune function as well as cytokine gene expression in murine peritoneal macrophages and splenic CD4+ T cells. Macrophages and CD4+ T cells were exposed to various concentrations of DU, and cell death via apoptosis and necrosis was analyzed using annexin-V/propidium iodide assay. DU cytotoxicity in both cell types was concentration dependent, with macrophage apoptosis and necrosis occurring within 24 hr at 100 microM DU exposure, whereas CD4+ T cells underwent cell death at 500 microM DU exposure. Noncytotoxic concentrations for macrophages and CD4+ T cells were determined as 50 and 100 microM, respectively. Lymphoproliferation analysis indicated that macrophage accessory cell function was altered with 200 microM DU after exposure times as short as 2 hr. Microarray and real-time reverse-transcriptase polymerase chain reaction analyses revealed that DU alters gene expression patterns in both cell types. The most differentially expressed genes were related to signal transduction, such as c-jun, NF- kappa Bp65, neurotrophic factors (e.g., Mdk), chemokine and chemokine receptors (e.g., TECK/CCL25), and interleukins such as IL-10 and IL-5, indicating a possible involvement of DU in cancer development, autoimmune diseases, and T helper 2 polarization of T cells. The results are a first step in identifying molecular targets for the toxicity of DU and the elucidation of the molecular mechanisms for the immune modulation ability of DU.

SUBMITTER: Wan B

PROVIDER: S-EPMC1332661 | biostudies-literature | 2006 Jan

REPOSITORIES: biostudies-literature

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In vitro immune toxicity of depleted uranium: effects on murine macrophages, CD4+ T cells, and gene expression profiles.

Wan Bin B Fleming James T JT Schultz Terry W TW Sayler Gary S GS

Environmental health perspectives 20060101 1

Depleted uranium (DU) is a by-product of the uranium enrichment process and shares chemical properties with natural and enriched uranium. To investigate the toxic effects of environmental DU exposure on the immune system, we examined the influences of DU (in the form of uranyl nitrate) on viability and immune function as well as cytokine gene expression in murine peritoneal macrophages and splenic CD4+ T cells. Macrophages and CD4+ T cells were exposed to various concentrations of DU, and cell d ...[more]

PMID: 16393663

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Project description:BACKGROUND: Mercury is known to bioaccumulate and to magnify in marine mammals, which is a cause of great concern in terms of their general health. In particular, the immune system is known to be susceptible to long-term mercury exposure. The aims of the present study were (1) to determine the mercury level in the blood of free-ranging harbour seals from the North Sea and (2) to examine the link between methylmercury in vitro exposure and immune functions using seal and human mitogen-stimulated peripheral blood mononuclear cells (T-lymphocytes). METHODS: Total mercury was analysed in the blood of 22 harbour seals. Peripheral blood mononuclear cells were isolated from seals (n = 11) and from humans (n = 9). Stimulated lymphocytes of both species were exposed to functional tests (proliferation, metabolic activity, radioactive precursor incorporation) under increasing doses of methylmercury (0.1 to 10 microM). The expression of cytokines (IL-2, IL-4 and TGF-beta) was investigated in seal lymphocytes by RT-PCR and by real time quantitative PCR (n = 5) at methylmercury concentrations of 0.2 and 1 microM. Finally, proteomics analysis was attempted on human lymphocytes (cytoplasmic fraction) in order to identify biochemical pathways of toxicity at concentration of 1 microM (n = 3). RESULTS: The results showed that the number of seal lymphocytes, viability, metabolic activity, DNA and RNA synthesis were reduced in vitro, suggesting deleterious effects of methylmercury concentrations naturally encountered in free-ranging seals. Similar results were found for human lymphocytes. Functional tests showed that a 1 microM concentration was the critical concentration above which lymphocyte activity, proliferation and survival were compromised. The expression of IL-2 and TGF-beta mRNA was weaker in exposed seal lymphocytes compared to control cells (0.2 and 1 microM). Proteomics showed some variation in the protein expression profile (e.g. vimentin). CONCLUSION: Our results suggest that seal and human PBMCs react in a comparable way to MeHg in vitro exposure with, however, larger inter-individual variations. MeHg could be an additional cofactor in the immunosuppressive pollutant cocktail generally described in the blood of seals and this therefore raises the possibility of additional additive effects in the marine mammal immune system.

Project description:Toxoplasmosis, which is one of the most widespread zoonoses worldwide, has a high incidence and infection can result in severe disease in humans and livestock. Citrate synthase (CS) is a component of nearly all living cells that plays a vital role in the citric acid cycle, which is the central metabolic pathway of aerobic organisms. In the present study, the citrate synthase I gene of Toxoplasma gondii (T. gondii) (TgCSI) was cloned and characterized. The TgCSI gene had an open reading frame of 1665 bp nucleotides encoding a 555 amino acid protein with a molecular weight of 60 kDa. Using western blotting assay, the recombinant protein was successfully recognized by the sera of rats experimentally infected with T. gondii, while the native protein in the T. gondii tachyzoites was detected in sera from rats immunized with the recombinant protein of TgCSI. Binding of the protein to murine macrophages was confirmed by immuno fluorescence assay. Following incubation of macrophages with rTgCSI, the rTgCSI protein was found to have a dual function, with low concentrations (5-10 μg/mL) enhancing phagocytosis and high levels (80 μg/mL) inhibiting phagocytosis. Investigation of murine macrophage apoptosis illustrated that 5 μg/mL rTgCSI protein can significantly induce early apoptosis and late stage apoptosis (*p < 0.05), while 10 μg/mL rTgCSI protein significantly induced early apoptosis, but had no effect on late stage of apoptosis (**p < 0.01), and 80 μg/mL rTgCSI protein inhibited late stage apoptosis of macrophages (*p < 0.05). Cytokine detection revealed that the secretion of interleukin-10, interleukin-1β, transforming growth factor-β1 and tumor necrosis factor-α of macrophages increased after the cells were incubated with all concentration of rTgCSI, with the exception that 5 μg/mL rTgCSI had no effect on the secretion of interleukin-10 and interleukin-1β. However, secretion of NO and cell proliferation of the macrophages were substantially reduced. Taken together, these results suggested that TgCSI can affect the immune functions of murine macrophages by binding to the cells in vitro.

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In vitro immune toxicity of depleted uranium: effects on murine macrophages, CD4+ T cells, and gene expression profiles.

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In vitro immune toxicity of depleted uranium: effects on murine macrophages, CD4+ T cells, and gene expression profiles.

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