Relationship between thallium(I)-mediated plasma membrane fluidification and cell oxidants production in Jurkat T cells
Introduction
Thallium (Tl) is a heavy metal present in the earth's crust as salts and minerals, at very low concentrations. This metal is widely used in the manufacture of electronic devices, in smelting plants, in cement factories, and in certain medical procedures (ATSDR, 1999). As a consequence of these human activities, Tl can be released to the environment (Douglas et al., 1990, Repetto et al., 1998, Heim et al., 2002, Xiao et al., 2004), exposing humans to its noxious effects. For example, it has been described that Tl can affect several tissues and systems, including the epidermal, gastrointestinal, cardiovascular, reproductive and renal systems (Douglas et al., 1990, Moore et al., 1993, Galvan-Arzate and Santamaria, 1998, Heim et al., 2002). It can also cross the blood–brain barrier (Galvan-Arzate and Rios, 1994, Galván-Arzate et al., 2000), and deposit in the brain where it causes neurodegeneration, demyelination, and the accumulation of end products of lipid oxidation (Galván-Arzate et al., 2000).
Recently, we demonstrated that both Tl cationic species (Tl+ and Tl3+) can interact with membrane phospholipids causing alterations in their physical properties (Villaverde and Verstraeten, 2003). Opposed to Tl3+, Tl+ has a relatively low charge density and, therefore, it has a poor tendency to establish ionic bonding to phospholipid headgroups. However, we found that Tl+ can induce a modest increase in liposome surface potential, followed by a decrease in the fluidity of lipid domains enriched in negatively charged phospholipids (Villaverde and Verstraeten, 2003).
The aim of the present work was to investigate the hypothesis that, similarly to our previous findings in liposomes, Tl+ could either affect cells plasma membrane fluidity through direct or indirect mechanisms. Working with human leukemia T cells (Jurkat), the effects of increasing Tl+ concentrations (5–100 μM) on plasma membrane integrity and fluidity were investigated. Also, the effects of this cation on the formation of cellular oxidant levels and the release of lipid oxidation products were evaluated, as well as their possible association with plasma membrane fluidity. In the range of concentrations assessed, and after 72 h of Tl+ addition, a significant increase in plasma membrane fluidity was observed. No alterations in cell membrane permeability to bulky molecules were found. In addition, Tl+ increased the content of cytoplasmic oxidant species and oxidation products release to the media. This latter significantly correlated with the increase in plasma membrane fluidity and with the average cells size. Together, the experimental results suggest that in Jurkat T cells, Tl+ exacerbates the production of oxidant species available to oxidize membrane lipids and alter their fluidity, effect that could ultimately lead to a dysfunction of plasma membrane-associated metabolic processes observed in Tl+ toxicity.
Section snippets
Chemicals
Human leukemia T cells (Jurkat) were obtained from the American Type Culture Collection (A.T.C.C., Rockville, MD, USA). RPMI 1640 medium and fetal bovine serum (FBS) were purchased from Gibco BRL (Grand Island, NY, USA). Thallium(I) nitrate was from Fluka (Milwaukee, WI, USA). The fluorescent probes 6-(9-anthroyloxy)stearic acid (6-AS), 5(6)-carboxy-2′,7′-dichlorodihydrofluorescein diacetate (DCDCDHF), merocyanine 540 (MC), and propidium iodide (PI) were purchased from Molecular Probes Inc.
Effects of Tl+ on cell viability
The possibility that Tl+ could affect the functional integrity of Jurkat T cells plasma membrane was investigated 72 h after a single administration of Tl+ (5–100 μM). The exclusion of the dye Trypan Blue, a method commonly used for the determination of cell viability was first assessed. In the range of concentrations studied, Tl+ did not affect cell viability (Fig. 1A). Similarly, the ability of these cells to reduce the dye MTT was comparable to that found in control cells (Fig. 1A). The
Discussion
Tl is a non-essential heavy metal that contaminates soils and water due to its elimination into the environment (Heim et al., 2002, Hoffman, 2003). In fact, the high content of Tl in the soil is an environmental concern in certain countries (Heim et al., 2002, Xiao et al., 2004). It has been described that Tl+ intoxication results in the alteration of a considerable amount of biological processes. Among others, Tl+ can induce a local increase of lipid oxidation products content in five
Acknowledgements
This work was supported by grants from the University of Buenos Aires (B 072) and the Agencia Nacional de Promoción Científica y Tecnológica (PICT 12285), Argentina.
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