Functional consequences of the Asp299Gly Toll-like receptor-4 polymorphism.

Toll-like receptor-4 (TLR4) is a pattern-recognition receptor not only for exogenous ligands such as lipopolysaccharide (LPS) of Gram-negative bacteria, but also for endogenous ligands such as fibronectin, heat shock proteins and hyaluronan oligosaccharides. The Asp299Gly allele of the TLR4 gene has been associated with increased risk for severe infections, but reduced progression of atherosclerosis. We have investigated the consequences of the presence of Asp299Gly polymorphism after stimulation of mononuclear cells with lipopolysaccharide (LPS), the non-LPS TLR4 microbial stimuli Aspergillus fumigatus and Cryptococcus neoformans, and the endogenous TLR4 ligand heat shock protein 60. No differences in either production of the proinflammatory cytokine TNF or the antiinflammatory cytokine interleukin-10 were observed between volunteers with the wild-type allele, volunteers heterozygous for the Asp299Gly allele and one volunteer homozygous for the TLR4 variant. In conclusion, the presence of the Asp299Gly TLR4 polymorphism does not result in defective pro and antiinflammatory cytokine production after stimulation with either exogenous (LPS and non-LPS) or endogenous TLR4 ligands, and alternative explanations are likely to be responsible for the epidemiological data showing associations with inflammatory conditions. In addition, this is the first study to demonstrate that even homozygosity for the Asp299Gly mutation does not confer hyporesponsiveness to stimulation with TLR4 stimuli.

Chlamydia pneumoniae stimulates IFN-gamma synthesis through MyD88-dependent, TLR2- and TLR4-independent induction of IL-18 release.

Recent studies suggest that inflammation plays a central role in the pathogenesis of atherosclerosis, and IFN-gamma is a prominent proinflammatory mediator in this context. However, it is unclear what stimuli are responsible for initial stimulation of IFN-gamma synthesis in the vessel wall. In the present study, we demonstrate that Chlamydia pneumoniae is an important stimulus for IFN-gamma synthesis, and this production depends on release of endogenous IL-18, IL-12, and IL-1, but not of TNF. The production of the proinflammatory cytokines TNF and IL-1beta from PBMC by sonicated C. pneumoniae was mediated through TLR2-dependent pathways. In contrast, C. pneumoniae stimulated the production of IL-18 through MyD88-dependent, TLR2-, TLR4-, and CD14-independent pathways, mediated by posttranscriptional mechanisms not involving de novo protein synthesis. In conclusion, C. pneumoniae is a potent stimulus of IFN-gamma production, in addition to the proinflammatory cytokines TNF and IL-1beta, which may contribute to its proatherogenic effects. Most interestingly, C. pneumoniae is also a potent inducer of IL-18 production through pathways independent of TLR2 and TLR4.

Human lipoproteins have divergent neutralizing effects on E. coli LPS, N. meningitidis LPS, and complete Gram-negative bacteria.

The use of lipoproteins has been suggested as a treatment for Gram-negative sepsis because they inhibit lipopolysaccharide (LPS)-mediated cytokine production. However, little is known about the neutralizing effects of lipoproteins on cytokine production by meningococcal LPS or whole Gram-negative bacteria. We assessed the neutralizing effect of LDLs, HDLs, and VLDLs on LPS- or whole bacteria-induced cytokines in human mononuclear cells. A strong inhibition of Escherichia coli LPS-induced interleukin-1beta (IL-1beta), tumor necrosis factor-alpha, and IL-10 by LDL and HDL was seen, whereas VLDL had a less pronounced effect. In contrast, Neisseria meningitidis LPS, in similar concentrations, was neutralized much less effectively than E. coli LPS. Effective neutralization of meningococcal LPS required a longer interaction time, a lower concentration of LPS, or higher concentrations of lipoproteins. The difference in neutralization was independent of the saccharide tail, suggesting that the lipid A moiety accounted for the difference. Minimal neutralizing effects of the lipoproteins were observed on whole E. coli or N. meningitidis bacteria under all conditions tested. These results indicate that efficient neutralization of LPS depends on the type of LPS, but a sufficiently long interaction time, a low LPS concentration, or high lipoprotein concentration also inhibited cytokines by the less efficiently neutralized N. meningitidis LPS. Irrespective of these differences, whole bacteria showed no neutralization by lipoproteins.

Parenteral administration of medium- but not long-chain lipid emulsions may increase the risk for infections by Candida albicans.

Intravenous administration to volunteers of an emulsion of medium-chain lipids, but not of an emulsion of pure long-chain lipids or a placebo, increased the growth of Candida albicans in serum and modulated Candida-induced cytokine production by mononuclear cells in a way suggesting that medium-chain, but not long-chain, triglycerides increase the risk for infections by Candida.

Native LDL potentiate TNF alpha and IL-8 production by human mononuclear cells.

Native LDL (nLDL) increases expression of adhesion molecules on endothelial cells through induction of Ca(2+) mobilization. Ca(2+) mobilization is also involved in the induction of proinflammatory cytokines, important mediators involved in atherogenesis. The aim of the study was to evaluate the capacity of nLDL to affect spontaneous and lipopolysaccharide (LPS)-stimulated cytokine production. Preincubation of human peripheral blood mononuclear cells (PBMC) with nLDL for 24 h did not influence spontaneous production of tumor necrosis factor alpha (TNF alpha) or interleukin-8 (IL-8), but significantly potentiated LPS-induced production of these cytokines. nLDL preincubation of PBMC did not increase the expression of the LPS receptors Toll-like receptor-4, CD14, or CD11c/CD18. Potentiation of cytokine production by nLDL was mediated through induction of Ca(2+) mobilization, because: a) nLDL induced a sustained pattern of repetitive Ca(2+) transients in human PBMC; b) the Ca(2+) chelator fura 2-acetoxymethyl ester, 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid, an intracellular Ca(2+) chelator, inhibited the potentiating effect of nLDL on LPS-induced cytokine synthesis; c) induction of Ca(2+) mobilization by thapsigargin potentiated LPS-induced cytokine production. nLDL are able to potentiate LPS-induced production of cytokines by human PBMC, and this effect is probably mediated through induction of Ca(2+) mobilization. This may represent an important pathogenetic mechanism in atherogenesis induced by hyperlipoproteinemia.