Cytokine Modulation in Experimental Endotoxemia: Characterization of an ex vivo Whole Blood Model

Abstract

A new model was developed to study cytokine regulation and modulation in whole blood ex vivo. The model is characterized by stable leukocyte counts and high leukocyte viability throughout the experimental period. Oxygen consumption per time decreased slowly, whereas carbon dioxide partial pressure increased accordingly throughout the experiment. In this model, the anti-inflammatory effects of recombinant human (rh) interleukin (IL)-4, rhIL-10 and rhIL-13 on lipopolysaccharide (LPS) stimulated (10 ng/ml) leukocytes were examined and compared by measuring their ability to inhibit the release and mRNA levels of tumor necrosis factor (TNF)α, IL-6 and IL-1β. rhIL-10 potently inhibited the release of TNF-α, IL-6 and IL-1β in a potent and dose-dependent manner, but did not influence the mRNA levels of these cytokines in CD14-positive cells. Also, rhIL-4 and rhIL-13 inhibited the release of IL-6 and IL-1β in a potent and dose-dependent manner, however, stronger maximal inhibition of IL-1β (85%) than of IL-6 (60%) was obtained. In contrast, rhIL-4 and rhIL-13 seemed to have both stimulatory and inhibitory effects on plasma values of TNF-α. The effects of 10 ng/ml LPS showed to be signalling through the CD14 receptor, since blood treated with a monoclonal anti-CD14 antibody did not produce any TNF-α. The whole blood model described in this study is in our opinion a useful tool for investigating immunomodulating effects on a mixed white blood cell population.

Peptidoglycan and Lipoteichoic Acid fromStaphylococcus aureusInduce Tumor Necrosis Factor Alpha, Interleukin 6 (IL-6), and IL-10 Production in Both T Cells and Monocytes in a Human Whole Blood Model

“…This is in contrast to previous observations that CD14 is involved in PepG signaling (12,32). We recently demonstrated that MAb 18D11 abolished the ability of LPS to induce cytokine production in whole blood (31). However, it is not known whether LPS and PepG share binding sites on CD14 and 18D11 may block binding of LPS, but not of PepG, to the CD14 molecule.…”

“…We have recently developed and characterized the whole-blood model used in this study (31). Briefly, venous blood from healthy volunteers was anticoagulated with Na-citrate.…”

“…We have recently developed a whole-blood model to study the cytokine network under both physiological and pathophys-iological conditions (31). In the present study, we have examined the ability of PepG and LTA isolated from S. aureus to induce release of TNF-␣, IL-6, and IL-10 in whole human blood and identified the cellular origins of these cytokines.…”

Organ Injury and Cytokine Release Caused by Peptidoglycan Are Dependent on the Structural Integrity of the Glycan Chain

“…The human whole-blood model was used as previously described (47), with some modifications. In brief, venous blood from healthy volunteers was anticoagulated with heparine (30 IU/ml of blood; Leo, Ballerup, Denmark) and incubated in 0.5-ml thin-wall tubes (ABgene House, Epsom, United Kingdom) at 37°C with slow rotation in the absence or presence of 1 g of PepG/ml or 1 g of hydrolyzed PepG material (at a hydrolase concentration of 15 ng/ml)/ml.…”

Involvement of CD14 and Toll-Like Receptors in Activation of Human Monocytes byAspergillus fumigatusHyphae

“…The whole-blood model used in this study has recently been established and characterized (38). Venous blood from healthy volunteers was anticoagulated with heparin (30 IU per ml of blood).…”