Influence of EDTA and heparin on lipopolysaccharide binding and cell activation, evaluated at single‐cell level in whole blood

Brunialti, Milena Karina Coló; Kallás, Esper G.; Freudenberg, Marina A.; Galanos, Chris; Salomão, Reinaldo

doi:10.1002/cyto.10049.abs

Cited by 11 publications

(10 citation statements)

References 4 publications

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“…The test was performed as described above. Previous studies in our lab showed that anti-CD14 monoclonal antibody (clone MP9) binding to CD14 molecules on the monocyte surface was not influenced by the presence of LPS, even at concentrations of as high as 50,000 ng/ml (8).…”

Section: Methodsmentioning

confidence: 73%

Peripheral Blood Mononuclear Cell Activation Induced byLeptospira interrogansGlycolipoprotein

et al. 2002

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Section: Methodsmentioning

confidence: 73%

Peripheral Blood Mononuclear Cell Activation Induced byLeptospira interrogansGlycolipoprotein

et al. 2002

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“…Blood samples (10 ml) were taken by venous puncture with heparin as the anticoagulant. Heparin was chosen over EDTA since EDTA can interfere with cell activation (Brunialti et al ., ).…”

Section: Methodsmentioning

confidence: 94%

In vitro evaluation of the effects of perfluorooctanesulfonic acid (PFOS) and perfluorooctanoic acid (PFOA) on IL‐2 production in human T‐cells

Midgett

Peden‐Adams

Gilkeson

et al. 2014

J of Applied Toxicology

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Perfluorinated compounds, such as perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA), have been shown to alter various immune functions suggesting they are immunotoxic. This study assessed the effects of PFOS and PFOA on interleukin (IL)-2 production in the human Jurkat T-cell line and PFOS in healthy human primary T cells. Jurkat cells were stimulated with phytohemagglutinin (PHA)/phorbol myristate acetate (PMA), anti CD-3/anti CD-28, or anti CD-3, and dosed with 0, 0.05, 0.1, 0.5, 1, 5, 10, 50, 75, or 100 μg ml−1 PFOS or 0, 0.005, 0.01, 0.05, 0.1, 0.5, 1, 5, or 10 μg ml−1 PFOA. Jurkat cells stimulated with PHA/PMA or anti CD-3 exhibited decreased IL-2 production beginning at 50 μg PFOS ml−1 and 5 μg PFOS ml−1 respectively, but stimulation with anti-CD3/anti-CD28 resulted in no changes compared with the control. Addition of the PPAR-alpha antagonist GW6471 to PFOS-dosed cells stimulated with PHA/PMA resulted in decreases in IL-2 production starting at 50 μg PFOS ml−1, which suggests PFOS affected T-cell IL-2 production via PPAR-alpha-independent mechanisms. Exposure to PFOA, PFOA + GW6471, or PFOS + PFOA in Jurkat cells resulted in no significant differences in IL-2 production. In vitro dosing studies using healthy primary human CD4+ T cells were consistent with the Jurkat results. These data demonstrated that PFOA did not impact IL-2 production, but PFOS suppressed IL-2 production in both a human cell line and human primary cells at dose levels within the high end of the human exposure range. A decrease in IL-2 production is characteristic of autoimmune diseases such as systemic lupus erythematosus and should be further investigated.

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“…For ex vivo TLR stimulation experiments, all blood samples were collected in Vacutainers containing the anticoagulant EDTA, as previous studies have demonstrated that the presence of heparin contributes to augmented monocyte responses to LPS via the activity of heparin-binding protein (10,19). For phagocytosis and oxidative-burst assays, blood samples were collected in Vacutainers containing the anticoagulant heparin to prevent the inhibitory effects of EDTA on these functions (4).…”

Section: Blood Collectionmentioning

confidence: 99%

Simian Immunodeficiency Virus-Induced Alterations in Monocyte Production of Tumor Necrosis Factor Alpha Contribute to Reduced Immune Activation in Sooty Mangabeys

Mir

Bosinger

Gasper

et al. 2012

J Virol

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g Human immunodeficiency virus type 1 (HIV-1) infection is characterized by persistent viral replication in the context of CD4؉ T cell depletion and elevated immune activation associated with disease progression. In contrast, simian immunodeficiency virus (SIV) infection of African-origin sooty mangabeys (SM) generally does not result in simian AIDS despite high viral loads and therefore affords a unique model in which to study the immunologic contributions to a nonpathogenic lentiviral disease outcome. A key feature of these natural SIV infections is the maintenance of low levels of immune activation during chronic infection. Our goal was to delineate the contribution of monocytes to maintaining low levels of immune activation in SIV-infected SM. Utilizing an ex vivo whole-blood assay, proinflammatory cytokine production was quantified in monocytes in response to multiple Toll-like receptor (TLR) ligands and a specific, significant reduction in the tumor necrosis factor alpha (TNF-␣) response to lipopolysaccharide (LPS) was observed in SIV-infected SM. In contrast, monocytes from hosts of pathogenic infections (HIV-infected humans and SIV-infected Asian macaques) maintained a robust TNF-␣ response. In SIV-infected SM, monocyte TNF-␣ responses to low levels of LPS could be augmented by the presence of plasma from uninfected control animals. The impact of LPS-induced TNF-␣ production on immune activation was demonstrated in vitro, as TNF-␣ blocking antibodies inhibited downstream CD8؉ T cell activation in a dose-dependent manner. These data demonstrate an association between nonpathogenic SIV infection of SM and a reduced monocyte TNF-␣ response to LPS, and they identify a role for monocytes in contributing to the suppressed chronic immune activation observed in these natural hosts.

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Influence of EDTA and heparin on lipopolysaccharide binding and cell activation, evaluated at single‐cell level in whole blood

Cited by 11 publications

References 4 publications

Peripheral Blood Mononuclear Cell Activation Induced byLeptospira interrogansGlycolipoprotein

Peripheral Blood Mononuclear Cell Activation Induced byLeptospira interrogansGlycolipoprotein

In vitro evaluation of the effects of perfluorooctanesulfonic acid (PFOS) and perfluorooctanoic acid (PFOA) on IL‐2 production in human T‐cells

Simian Immunodeficiency Virus-Induced Alterations in Monocyte Production of Tumor Necrosis Factor Alpha Contribute to Reduced Immune Activation in Sooty Mangabeys

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