Rhodobacter capsulatus PG Lipopolysaccharide Blocks the Effects of a Lipoteichoic Acid, a Toll-Like Receptor 2 Agonist.

Lipopolysaccharides (LPS) and lipoteichoic acids (LTA) are the major inducers of the inflammatory response of blood cells caused by Gram-negative and some Gram-positive bacteria. CD14 is a common receptor for LPS and LTA that transfers the ligands to TLR4 and TLR2, respectively. In this work, we have demonstrated that the non-toxic LPS from Rhodobacter capsulatus PG blocks the synthesis of pro-inflammatory cytokines during the activation of blood cells by Streptococcus pyogenes LTA through binding to the CD14 receptor, resulting in the signal transduction to TLR2/TLR6 being blocked. The LPS from Rhodobacter capsulatus PG can be considered a prototype for developing preparations to protect blood cells against the LTA of gram-positive bacteria.

The Effect of a Lipopolysaccharide from Rhodobacter capsulatus PG on Inflammation Caused by Various Influenza Strains.

The development of a specific inflammation in mice that had been infected by two influenza virus strains, A/chicken/Kurgan/5/2005 (H5N1) and A/Hamburg/2009 MA (H1N1), was studied. We investigated the effect of a non-toxic lipopolysaccharide from Rhodobacter capsulatus PG on the survival and body weight of the mice, production of IgG antibodies, and the induction of pro- and anti-inflammatory cytokines in blood serum. The administration of the R. capsulatus PG lipopolysaccharide was shown to induce interferon-ß synthesis, both in healthy and influenza A virus-infected mice, and to promote production of antiviral antibodies in the blood of the influenza-infected animals.

In vivo Proinflammatory Cytokine Production by CD-1 Mice in Response to Equipotential Doses of Rhodobacter capsulatus PG and Salmonella enterica Lipopolysaccharides.

The capacities of relatively nontoxic lipopolysaccharide (LPS) from Rhodobacter capsulatus PG and highly potent LPS from Salmonella enterica serovar Typhimurium to evoke proinflammatory cytokine production have been compared in vivo. Intravenous administration of S. enterica LPS at a relatively low dose (1 mg/kg body weight) led to upregulation of TNF-α, IL-6, and IFN-γ production by non-sensitized CD-1 mice. LPS from R. capsulatus PG used at a four-times higher dose than that from S. enterica elicited production of almost the same amount of systemic TNF-α; therefore, the doses of 4 mg/kg LPS from R. capsulatus PG and 1 mg/kg LPS from S. enterica were considered to be approximately equipotential doses with respect to the LPS-dependent TNF-α production by CD-1 mice. Rhodobacter capsulatus PG LPS was a weaker inducer of the production of TNF-α, IL-6, and IFN-γ, as compared to the equipotential dose of S. enterica LPS. Administration of R. capsulatus PG LPS before S. enterica LPS decreased production of IFN-γ, but not of TNF-α and IL-6, induced by S. enterica LPS. Rhodobacter capsulatus PG LPS also suppressed IFN-γ production induced by S. enterica LPS when R. capsulatus PG LPS had been injected as little as 10 min after S. enterica LPS, but to a much lesser extent. Rhodobacter capsulatus PG LPS did not affect TNF-α and IL-6 production induced by the equipotential dose of S. enterica LPS. In order to draw conclusion on the endotoxic activity of particular LPSs, species-specific structure or arrangement of the animal or human immune systems should be considered.

Artifacts Arising from Using Leukocytic Fc Receptor Blocking Buffer.

We studied the effects of Human TruStain FcX buffer (Fcγ receptor blocking solution) in experiments on evaluation of TLR4 level with labeled monoclonal antibodies, intracellular immunofluorescent staining of NF-κB p50, and TNF-α synthesis on human isolated monocytes and whole blood cells. The influence of the blocking buffer on the measured parameters should be taken into account and appropriateness of its use in experiments on isolated cells and whole blood should be considered.

Dynamics of Antagonistic Potency of Rhodobacter capsulatus PG Lipopolysaccharide against Endotoxin-Induced Effects.

The dynamics of antagonistic potency of lipopolysaccharide (LPS) isolated from Rhodobacter capsulatus PG on the synthesis of proinflammatory (TNF-α, IL-1ß, IL-8, IL-6, IFN-γ) and antiinflammatory (IL-10, IL-1Ra) cytokines induced by highly stimulatory endotoxins from Escherichia coli or Salmonella enterica have been studied. Using human whole blood, we have shown that R. capsulatus PG LPS inhibited most pronouncedly the endotoxin-induced synthesis of TNF-α, IL-1ß, IL-8, and IL-6 during the first 6 h after endotoxin challenge. Similarly, the endotoxin-induced release of IFN-γ was abolished by R. capsulatus PG LPS as well (24 h). In contrast to the above-mentioned cytokines, the relatively weak antagonistic activity of R. capsulatus PG LPS against endotoxin-triggered production of IL-6 and IL-8 was revealed. Since R. capsulatus PG LPS displays more potent antagonistic activity against deleterious effects of S. enterica LPS than those of E. coli LPS in the cases of such cytokines as IL-1ß (6 and 24 h), IL-6 and IL-8 (4 h), we conclude that the effectiveness of protective action of antagonist is mostly determined by the primary lipid A structure of the employed agonist.

Role of CD11b/CD18 in priming of human leukocytes by endotoxin glycoforms from Escherichia coli.

The primary objective of this study was to determine the role of ß2 integrin α-subunit (CD11b) in the mechanism of human polymorphonuclear leukocyte (PML) priming by S or Re endotoxin glycoforms from Escherichia coli for fMLP-induced respiratory burst. Similar priming activity of S and Re endotoxin glycoforms for fMLP-induced reactive oxygen species (ROS) generation from primed PML was found. Anti-CD11b antibodies (clone ICRF 44) as well as isotype-matched immunoglobulin G1 (clone MOPC-21) do not influence the fMLP-induced ROS generation from unprimed PML. Antibodies against CD11b do not change fMLP-induced ROS generation from endotoxin-primed PML as well. The involvement of different isoforms of Fcγ receptors in fMLP-induced ROS generation from activated PML is proposed.

Involvement of toll-like receptor 4 and Fc receptors gamma in human neutrophil priming by endotoxins from escherichia coli.

By using the fMLP-induced respiratory burst approach, the involvement of Toll-like receptor 4 (TLR4) in human neutrophil priming by S- or Re-glycoforms of endotoxin from Escherichia coli has been elucidated. The priming effect of Re-glycoform is more pronounced than that of the S-glycoform. Unexpectedly, fMLP-triggered generation of reactive oxygen species (ROS) by endotoxin primed neutrophils was amplified by preincubation of the cells with anti-TLR4 (HTA125) antibodies or with isotype-matched immunoglobulin IgG2a. The most significant finding of our study is that neutrophils exposed to anti-TLR4 antibodies retain their ability to distinguish between S- or Re-glycoforms being primed, respectively. Moreover, differentiated effect of HTA125 antibodies on functional responses of neutrophils during their priming and fMLP stimulation was revealed. Taking these results into consideration, it is reasonable to assume that there is a contribution of Fcγ receptors to fMLP-induced ROS generation by neutrophils preincubated with HTA125 or IgG2a and primed by endotoxins.

Structural analysis of lipopolysaccharides from Gram-negative bacteria.

This review covers data on composition and structure of lipid A, core, and O-polysaccharide of the known lipopolysaccharides from Gram-negative bacteria. The relationship between the structure and biological activity of lipid A is discussed. The data on roles of core and O-polysaccharide in biological activities of lipopolysaccharides are presented. The structural homology of some oligosaccharide sequences of lipopolysaccharides to gangliosides of human cell membranes is considered.

[The effect of composition of the core region of Escherichia coli K-12 lipopolysaccharides on the surface properties of cells].

The pH dependences of electrokinetic potentials (EKP) of the cells of two Escherichia coli K-12 strains (D21 and JM 103) with known lipopolysaccharide (LPS) core composition have been determined by the method of microelectrophoresis. At pH 4.6-5.2, the negative surface charge of the cells with Re core LPS was reliably higher. It was shown that the interaction of bacteria with lysozyme results in a decrease of optical density of suspensions due to higher sensitivity of the cells with complete LPS core to hypotonic shock. LPS release from bacterial cell wall depended also on bacterial LPS core composition and increased with LPS core extension. Electrokinetic measurements and the study of the interaction of cells with lysozyme suggest that higher negative surface charge of E. coli JM 103 cells (Re type LPS) is associated with higher quantity and density of LPS packing in the cell wall as compared with the cells of E. coli D21 (Ra type LPS).

[Relations between the chemotype and the cell electrophoretic properties in Rhodobacter capsulatus strains].

The cells of two Rhodobacter capsulatus strains, B10 and PG, and the LPS of their cell walls were studied by electrophysical and biochemical methods. Strain B10 was found to belong to the R chemotype, and strain PG, to the RS chemotype. A relation was revealed between the chemotype of the photosynthesizing bacteria Rhodobacter capsulatus and the electrophoretic properties of their cells.

Effect of chemotype on Escherichia coli interactions with bactericidal proteins.

We studied the effects of bactericidal proteins (lysozyme and lactoferrin) on endotoxin release from cell wall and inhibition of the growth of Escherichia coli colonies of different chemotypes. The structure of LPS core was found to be essential for the mechanisms of the interactions of the studied proteins with the cell wall. Cell viability after contact with cationic proteins is determined not by the amount of released LPS, but by the mechanism of damage to the cell wall.

Use of colorimetric method for evaluation of LPS of different structure.

We studied the possibility of using the colorimetric method with carbocyanin dye for quantitative measurements of LPS of different structure in aqueous solutions. The absorption spectra (intensity and position of absorption peaks) of LPS-carbocyanin complexes depend not only on the presence of polysaccharide fragment in LPS structure, but also on the core structure.

Effect of various E. coli LPS chemotypes on apoptosis and activation of human neutrophils.

We studied the effects of various Escherichia coli LPS chemotypes on the production of reactive oxygen species and regulation of apoptosis in human neutrophils. A correlation was found between the increase in chemiluminescence (Re-LPS