bluuebunny/crossref_metadata_2025 huggingface dataset:
Endotoxin tolerance protects against local hepatic ischemia/reperfusion injury in the rat
Authors: Domínguez Fernández Emilio , Flohé Sascha , Siemers Frank , Nau Michael , Ackermann Markus , Ruwe Markus , Schade Fritz Ulrich
Published: June 2007

Liver surgery and liver transplantation as well as circulatory shock are often associated with hepatic ischemia/reperfusion (I/R) injury. Recent evidence suggests that TNF-α plays a central role in I/R injury and, therefore, down-regulation of TNF-α seems to be a promising way to protect against the deleterious consequences of I/R. Endotoxin tolerance represents a state of unresponsiveness to endotoxin and is associated with diminished TNF-α production. Thus, the effect of endotoxin tolerance on hepatic I/R injury of the liver was investigated in a rat model. I/R injury was induced by temporary ischemia of the left lateral liver lobe for 90 min followed by a 3 h observation period of reperfusion. I/R injury resulted in functional hepatic disorder characterized by a decrease both in bile flow and bile acid concentration and 50% mortality. This was prevented by induction of endotoxin tolerance. Hepatic TNF-α mRNA expression after I/R of the liver was determined by RT-PCR. In untreated rats, TNF-α mRNA was induced in the liver 60 min after reperfusion and further increased until 3 h after reperfusion. In contrast, in endotoxin-tolerant rats, no increases in TNF-α mRNA expression were detected. This suggests that induction of endotoxin tolerance protects against hepatic I/R injury possibly via down-regulation of intra-organ TNF-α expression.
Induction of TNF-α and MnSOD by endotoxin: effect of E5531, a synthetic non-toxic lipid A analog
Authors: White Julie E. , Tsan Min-Fu
Published: June 2007

E5531, a synthetic lipid A analog, has been shown to inhibit endotoxin (lipopolysaccharide, LPS)-induced tumor necrosis factor-α (TNF-α) production by human monocytes and murine macrophages. Whether it also inhibits LPS induction of manganese superoxide dismutase (MnSOD) is not clear. In the current study, we demonstrated that E5531, while having no effect on TNF-α and MnSOD mRNAs by itself, markedly inhibited LPS- and lipid A-, but not TNF-α-, induced increases in TNF-α and MnSOD mRNAs in human monocytes. In contrast, E5531 at concentrations and conditions that markedly inhibit LPS-induced increases in TNF-α and MnSOD mRNAs, and TNFα production by human monocytes, had no effect on murine peritoneal macrophages. These results demonstrate that E5531 is a potent LPS antagonist in human monocytes. However, it does not show antagonist action against LPS in murine macrophages in the range of concentrations tested, suggesting that E5531 is a more potent antagonist in humans than in mice.
Innate recognition of lipopolysaccharide by Toll-like receptor 4/MD-2 and RP105/MD-1
Authors: Miyake Kensuke , Ogata Hirotaka , Nagai Yoshinori , Akashi Sachiko , Kimoto Masao
Published: June 2007

The Toll family of receptors has been implicated in innate recognition and subsequent activation of defense programs against pathogens such as bacteria and fungi. TLR4, for example, signals the presence of lipopolysaccharide (LPS), a membrane constituent of Gram-negative bacteria. LPS signaling via TLR4 is greatly enhanced by a molecule referred to as MD-2, which is associated with the extracellular domain of TLR4. The TLR4/MD-2 complex, therefore, recognizes LPS. RP105, another member of the Toll family, has a striking similarity to TLR4 in that it is associated with an MD-2-like molecule MD-1. B-cells lacking RP105 are severely impaired in LPS-induced proliferation and antibody production. Studies employing transfectants showed that RP105/MD-1, like MD-2, enhances the LPS signaling via TLR4. RP105/MD-1 thus constitutes an LPS-signaling complex on B-cells. These results suggest that a variety of cell surface molecules regulate LPS recognition/signaling by TLR4.
CD14: a bridge between innate immunity and adaptive IgE responses
Authors: Vercelli Donata , Baldini Mauro , Stern Debra , Lohman I. Carla , Halonen Marilyn , Martinez Fernando
Published: June 2007

Total IgE levels are known to be under genetic control. Linkage studies have indicated that one or more loci on chromosome 5q may control total IgE, as well as asthma and bronchial hyper-responsiveness to non-specific stimuli. Our group has undertaken a systematic analysis of chromosome 5q, and has recently characterized five single nucleotide polymorphisms at position —1619, —1359, —1145, —809, and —159 in the promoter of the gene encoding CD14, the myeloid pattern recognition receptor that is critical for efficient innate immune responses to lipopolysaccharide and bacterial ligands. Individuals homozygous for the three major CD14 haplotypes found in the Children Respiratory Study population ( n = 390) were analyzed for serum levels of total IgE and soluble CD14. A strong inverse correlation was found between these two parameters, i.e. carriers of the —1359T/—1145A/—159C haplotype had the highest levels of IgE, and the lowest levels of sCD14. Conversely, carriers of the —1359G/—1145G/—159T haplotype had the highest levels of sCD14 and the lowest IgE values. Our results suggest that genetic variation in CD14, a key gene of innate immunity, may modulate the effects that exposure to bacterial ligands has on the development of Th2 responses.
A monoclonal antibody with specificity for the genus Klebsiella binds to a common epitope located in the core region of Klebsiella lipopolysaccharide
Authors: Brade Lore , Podschun Rainer , Brade Helmut
Published: June 2007

A mouse monoclonal antibody (mAb) which has been obtained after immunization of mice with heat-killed Klebsiella pneumoniae strain R20/O1— followed by standard plasmacytoma cell fusion protocols was investigated for its ability to identify various species of the genus Klebsiella. Based on the published observation that the antibody binds to an epitope located in the core region of lipopolysaccharide (LPS) of strain R20/O1—, we tested whether this epitope is shared and exposed by other species of the genus Klebsiella. The antibody was able to bind to LPS of clinical isolates of K. pneumoniae ( n = 77), K. oxytoca ( n = 50), K. terrigena ( n = 49) and K. planticola ( n = 50) in 93%, 98%, 96% and 100%, respectively, but did not bind to LPS of other Gram-negative genera ( n = 159) as tested by Western blots and dot blots using proteinase K-digested whole cell lysates as antigens. Western blot analyses indicated that the antibody bound only to those LPS molecules which did not carry an O-antigen and that the antibody is thus different from those already published.
Influence of acyl chain fluidity on the lipopolysaccharide-induced activation of complement
Authors: Wiese Andre , Grünewald Petra , Schaper Klaus-Jürgen , Seydel Ulrich
Published: June 2007

Lipopolysaccharides (LPSs, endotoxins) are the major amphiphilic constituents of the outer leaflet of the outer membrane of Gram-negative bacteria. They are known to activate the complement cascade to form lytic membrane pores. Here, we study the influence of the fluidity of the acyl chains of LPSs and lipid As on the formation of lytic pores. To this end, we have performed electrical measurements on asymmetric planar endotoxin/phospholipid bilayers as a reconstitution model of the outer membrane using two deep rough mutant LPSs (from Escherichia coli strains WBB01 and WBB25) and two lipid As (from E. coli WBB25 and Rhodobacter sphaeroides). The two LPSs and the two lipid As each differ in their acylation pattern which is correlated with the fluidity. The addition of human serum to the endotoxin side of the bilayers led to the formation of membrane pores, and pore formation correlated in each case with acyl chain fluidity, i.e. time required for the first lytic pore to be formed was shorter for the more fluid endotoxin. Furthermore, in the case of LPSs, the activation rate was higher for the more fluid membrane and the respective bacteria had a higher susceptibility to the growth inhibitory action of serum.
Pan-caspase inhibitor zVAD enhances cell death in RAW246.7 macrophages
Authors: Sung Ouk Kim  , Jiahuai Han
Published: June 2007

Even though the pan-caspase inhibitor zVAD has been widely used as an anti-apoptotic agent, inefficient prevention or even enhancement of cell death has been reported in certain cells. To further investigate its effects on cell death, three different cell types were exposed to various apoptotic stimuli in the presence or absence of zVAD. In Jurkat cells, zVAD protected against cell death induced by tumor necrosis factor (TNF), sodium nitroprusside (SNP) and etoposide, whereas in L929 cells cell death was increased. In RAW246.7 macrophages, zVAD showed similar effects as in L929 cells. However, unlike L929 cells, in which the cell death by TNF is known to be necrosis, RAW246.7 cells manifested features of apoptosis such as chromatin condensation and nuclear fragmentation. Induction of cell death by zVAD in lipopolysaccharide (LPS)-activated RAW246.7 cells also showed the same features as those observed in SNP- and etoposide-treated cells. Initiation of an apoptotic process by zVAD not only disputes the sole role of caspases in apoptosis but also suggests an anti-apoptotic function of certain caspase(s). Death of LPS-activated macrophages may be controlled by an anti-apoptotic caspase.
Toll-like receptors (TLRs) signalling and expression pattern
Authors: Muzio Marta , Mantovani Alberto
Published: June 2007

Toll is a Drosophila gene essential for ontogenesis and antimicrobial resistance. Several homologues of Toll have been identified and cloned in vertebrates, namely Toll-like receptors (TLRs). TLRs are structurally characterized by a cytoplasmic Toll/interleukin-1R (TIR) domain and by extracellular leucine rich repeats. TLRs characterized so far activate the MyD88/IRAK signalling cascade which activates NF-κB transcription factor. Genetic, gene transfer, and dominant negative approaches have involved TLR family members (TLR2 and TLR4) in distinct bacterial components for recognition and signalling. However, very little information is available regarding other TLRs. Here we propose to classify TLRs based on their expression pattern, in ubiquitous (TLR1), restricted (TLR2, 4, 5) and specific (TLR3) TLRs. Differential expression and regulation as well as distinct, though overlapping, ligand recognition patterns may underlie the existence of a such a large, seemingly redundant, TLR family.
CD14 and LBP in endotoxemia and infections caused by Gram-negative bacteria
Authors: Heumann Didier
Published: June 2007

It is now well recognized that plasma LPS-binding protein (LBP) and membrane CD14 present at the surface of cells of the myelo/monocytic lineage are central molecules of the innate immune system, in response to LPS or to bacterial products. This paper reviews the role of LBP and CD14 in models of endotoxemia and infection.
Phase variable changes in genes lgtA and lgtC within the lgtABCDE operon of Neisseria gonorrhoeae can modulate gonococcal susceptibility to normal human serum
Authors: Shafer William M. , Datta Anup , Kumar Kolli V.S. , Mahbubur Rahman M. , Balthazar Jacqueline T. , Martin Larry E. , Veal Wendy L. , Stephens David S. , Carlson Russell
Published: June 2007

The α-chain of the core oligosaccharide of the lipo-oligosaccharide (LOS) produced by Neisseria gonorrhoeae can undergo reversible and rapid changes in structure due to phase-variable production of certain enzymes employed in the biosynthesis of the lacto- N-neotetraose structure. Five of these enzymes are encoded by the lgtABCDE operon, and polynucleotide tracts within three of these genes ( lgtA, lgtC and lgtD) can be substrates for slipped-strand mispairing events that lead to nucleotide insertions or deletion events which result in variable production of their respective gene products. We now report that phase-variable synthesis of the lgtA and lgtC gene products in strain FA19 results in the production of elongated LOS α-chains and that the presence of these LOS species can result in gonococci being sensitive to the bacteriolytic action of serum-antibody and complement. Hence, phase variation within the lgtABCDE operon can significantly impact the ability of gonococci to subvert this important host defense system.
Inhibition of histone deacetylation enhances endotoxin-stimulated transcription but does not reverse endotoxin tolerance
Authors: Yoza Barbara K. , Hu Jean Y.-Q. , McCall Charles E.
Published: June 2007

Covalent modification of histones and the subsequent remodeling of chromatin have emerged as important mechanisms in regulating gene expression. In particular, recent identification of the enzyme families responsible for the steady-state balance of histone acetylation has served to redefine our understanding of these modifications as fundamental biochemical processes regulating transcription. Current evidence suggests that histone acetylation correlates positively with gene activation, while histone deacetylation acts to repress transcription. In this study, we examined the role of histone modification in the inflammatory response to endotoxin. We focused on the endotoxin-stimulated expression of the interleukin-1β promoter and tested the hypotheses that persistent histone deacetylation was responsible for the decreased expression of this promoter observed after prolonged exposure to endotoxin, a manifestation of a phenomenon known as endotoxin tolerance. We found that histone deacetylase inhibitors enhanced endotoxin-stimulated transcription; however, deacetylation inhibitors could neither block the development of tolerance nor restore endotoxin sensitivity in a tolerant cell. Deacetylase inhibitors could not restore LPS-mediated transcription in tolerant cells. These results show that histone acetylation/deacetylation regulates, at least in part, the endotoxin-induced expression of inflammatory genes and that repressed transcription observed in endotoxin tolerance is not caused by enhanced activity of histone deacetylases.
Cellular mechanism underlying LPS-induced inhibition of in vitro L-leucine transport across rabbit jejunum
Authors: Abad B. , Mesonero J.E. , Salvador M.T. , Garcia-Herrera J. , Rodriguez-Yoldi M.J.
Published: June 2007

Lipopolysaccharide(LPS) is a known causative agent of sepsis. In previous studies, we have shown that it reduces L-leucine mediated transport across the rabbit jejunum by about 30%. In this study, the mechanism(s) of LPS inhibition on amino acid transport were analysed in detail. LPS did not inhibit L-leucine transport across brush border membrane vesicles, suggesting the need for an intracellular step. The inhibitory effect of LPS was not altered by the addition of protein kinase A (PKA) inhibitor (IP20, 10—7 M) or an analog of cAMP (DB-cAMP, 3 × 10—4 M), indicating that the PKA signal transduction pathway was not involved in the LPS effect. However, the inhibitory effect of LPS was suppressed by trifluoroperazine (10—7 M), a Ca2+/calmodulin inhibitor and staurosporine (10—7 M), an protein kinase C (PKC) inhibitor. Likewise, LPS inhibition disappeared in media without calcium. These results suggest that LPS could inhibit the intestinal uptake of L-leucine across the small intestine in vitro by intracellular processes related to calcium, involving PKC and calmodulin protein.
Alterations in membrane-associated CD14 expression and the simultaneous liberation of soluble CD14 fragment in adherent macrophages mediated by a leukocyte carboxyl/aspartate protease
Authors: Coyne C.P. , Howell Trey , Smodlaka Hrvoje , Willetto Carla , Fenwick Brad W. , Chenney Erle
Published: June 2007

Investigations sought to discover the biochemical mechanisms in macrophages that mediate the `shedding' of soluble CD14 fragment. Stimulated macrophages display both increased liberation of soluble CD14 fragment and decreases in residual membrane-associated CD14 complexes following exposure to activating agents ( fMLP/A23187). Application of `class-specific' protease inhibitors revealed that a thiol/cysteine was involved in the biochemical production of soluble CD14 fractions and that a metalloprotease enzymatically degraded soluble CD14 fragment. Exposure of macrophages to individual proteases revealed that both cathepsin-D and elastase promoted variable depletion of membrane-associated CD14 complexes. Additionally, cathepsin-D, and to a lesser extent elastase, generated soluble CD14 fragment. Related studies isolated a carboxyl/aspartate protease from activated macrophages using pepstatin-A affinity chromatography. The physical and functional properties of macrophage pepstatin-A binding protein fractions closely corresponded with the known characteristics of cathepsin-D with respect to: (i) cellular origin; (ii) binding-avidity of carboxyl/aspartate proteases for pepstatin-A; (iii) non-specific proteolysis of haemoglobin detected by Hb-PAGE zymography; and (iv) hydrolysis of a synthetic cathepsin-D-specific peptide substrate. Interpretation of these findings collectively implies that activated leukocytes can biochemically alter membrane-associated CD14 complex expression and promote the liberation of soluble CD14 fragment in both activated and non-activated cell populations.
Requirement of TNF and TNF receptor type 2 for LPS-induced protection from lethal septic peritonitis
Authors: Echtenacher Bernd , Männel Daniela N.
Published: June 2007

Pretreatment of mice with low quantities of LPS induces endotoxin t olerance characterized by enhanced resistance to lethal doses of LPS and to a number of infectious challenges. M ice subjected to cecal ligation and puncture (CLP) survived the ensuing septic peritonitis significantly better when they had been pretreated with LPS. This LPS-induced protection was dependent on endogenous TNF production capacity since LPS pretreatment did not protect TNF-deficient mice from death after CLP. W hile mice deficient in the TNF receptor type 2 (p75TNFR) were as sensitive to CLP-induced mortality as control mice, LPS pretreatment could not reduce mortality in p75TNFR-deficient mice after CLP. Therefore, activation of the TNF receptor type 2 by endogenous TNF constitutes an important interaction for the development of LPS-induced resistance to bacterial infection.
Modulating macrophage function with IgG immune complexes
Authors: Anderson Charles F. , Gerber Jeffrey S. , Mosser David M.
Published: June 2007

Macrophages respond to bacterial products by releasing a large array of inflammatory mediators. We demonstrate that, in the presence of IgG immune complexes, macrophages produce high levels of IL-10 and virtually no IL-12, when they are exposed to bacterial products. The production of IL-10 by these cells can dampen innate inflammatory responses to microbial products, such as LPS. This alteration in macrophage cytokine production can also influence an adaptive immune response, preferentially inducing Th2-type immunity. Thus, immune complexes change the physiology of activated macrophages, converting them to anti-inflammatory cells that induce Th2-like immune responses. We have termed these cells type II activated macrophages.
The role of endotoxin and the innate immune response in the pathophysiology of acute graft versus host disease
Authors: Cooke Kenneth R. , Olkiewicz Krystyna , Erickson Nicole , Ferrara James L.M.
Published: June 2007

Allogeneic stem cell transplantation (SCT) is an important therapy for a number of malignant diseases, and acute graft versus host disease (GVHD) and leukemic relapse remain the two major obstacles to successful outcomes of this treatment strategy. The therapeutic potential of allogeneic SCT relies on the graft versus leukemia (GVL) effect, during which donor T lymphocytes eradicate residual malignant cells via immunological mechanisms. Unfortunately, beneficial GVL effects are closely associated with the toxicity of GVHD. The pathophysiology of GVHD is complex and fundamentally depends upon aspects of adaptive immunity and interactions between donor T cells and foreign host tissue antigens. Recent work has revealed that components of the innate immune response and the secretion of inflammatory cytokine effectors are also important. In this context, experimental studies have demonstrated that loss of gastrointestinal (GI) tract integrity plays a major role in the amplification of systemic GVHD. Specifically, translocation of endotoxin across a damaged GI tract and into the circulation promotes local and systemic cytokine release. This effect perpetuates further gut mucosal injury and endotoxin leak, thus establishing a positive feedback loop for progressive target organ injury and systemic inflammation. Data obtained using murine SCT models have shown that disruption of the cellular activating effects of lipopolysaccharide (LPS) significantly reduces cytokine secretion and GVHD severity without altering T-cell responses to host antigens. These findings support a critical role for LPS in the early inflammatory events responsible for GVHD and suggest that strategies which target the innate immune response and LPS receptor-ligand interactions may help prevent GVHD while preserving donor T-cell responses and GVL activity.
Association of LPS chemotype of Mannheimia (Pasteurella) haemolytica A1 with disease virulence in a model of ovine pneumonic pasteurellosis
Authors: Hodgson J. Christopher , Moon Gordon M. , Quirie Malcolm , Donachie William
Published: June 2007

Host responses during pneumonic pasteurellosis were compared in sheep infected with strains of Mannheimia (Pasteurella) haemolytica A1 differing in their O-antigen type. Nine-week-old, specific pathogen-free lambs were infected intratracheally with parainfluenza type 3 virus (108 TCID50) followed 7 days later by 5—6 . 107 CFU of M. haemolytica A1 possessing rough (group R, 6 lambs) or smooth (group S, 6 lambs) lipopolysaccharide, or saline (group C, 4 lambs). Group C lambs remained afebrile with no evidence of endotoxaemia or bacteraemia and biochemical parameters were normal. Group R and group S lambs became febrile within 2—3 h postinfection and the response was higher and more prolonged in group R lambs. Four group R and 2 group S lambs developed clinical pneumonic pasteurellosis within 24—48 h and the severity of disease correlated with episodes of endotoxaemia, bacteraemia and elevated eicosanoid concentrations. At postmortem, M. haemolytica (107—109 CFU/g) was isolated from the lungs of all 6 group R lambs but from only 1 group S lamb. The results indicate an association between the incidence and severity of ovine pneumonic pasteurellosis and LPS chemotype and suggest an important role for LPS chemotype in determining host-species susceptibility to lung infection.
Overexpression of CD14, TLR4, and MD-2 in HEK 293T cells does not prevent induction of in vitro endotoxin tolerance
Authors: Medvedev Andrei E. , Vogel Stefanie N.
Published: June 2007

TLR4 and MD-2 are necessary for conferring cellular responsiveness to LPS. Prior exposure to LPS induces a transient state of cell refractoriness to subsequent LPS re-stimulation, known as `endotoxin tolerance'. While induction of LPS tolerance has been reported to correlate with down-regulation of cell surface expression of TLR4/MD-2, other mechanisms of LPS tolerance have been revealed that target intracellular intermediates downstream of the TLR4/MD-2 complex. In this study, we sought to examine whether endotoxin tolerance could be induced under conditions where expression of TLR4 and MD-2 proteins is not affected by LPS. Human HEK 293T cells are completely unresponsive to LPS, but acquire high LPS sensitivity following transient transfection with CD14, TLR4, and MD-2 (293T/CD14/TLR4/MD-2 cells), as judged by NF-κB activation, ERK 1/2 phosphorylation, and TNFα gene expression. Prior exposure of 293T/CD14/TLR4/MD-2 cells to LPS resulted in a significant decrease of LPS-mediated responses, yet failed to affect expression levels of TLR4 and MD-2. Thus, altered expression and/or function of intracellular mediators downstream of the TLR4/MD-2 complex play an important role in mediating LPS tolerance.
Impact of sepsis-induced changes in plasma on LPS interactions with monocytes and plasma lipoproteins: roles of soluble CD14, LBP, and acute phase lipoproteins
Authors: Kitchens Richard L. , Thompson Patricia A.
Published: June 2007

Sepsis-induced changes in human plasma decrease LPS association with monocytes by regulating dynamic interactions among LPS, monocytes, and plasma lipoproteins. In the physiological environment of undiluted human serum, we have found that: (i) LPS binds transiently to monocytes and is released into plasma lipoproteins; (ii) the release of LPS from monocytes is dependent upon lipoprotein acceptors and is enhanced by soluble CD14 (sCD14); and (iii) both lipoproteins and sCD14 can attenuate cytokine responses in monocytes that have already bound LPS. Whereas LPS binding protein (LBP) also inhibited LPS responses after LPS had bound to monocytes, this did not require extensive release of cell-bound LPS as was observed with sCD14. In the serum of septic patients, both free LPS and monocyte-bound LPS were usually transferred to lipoproteins at an accelerated rate. In spite of a sharp decline in HDL levels, HDL remained the dominant LPS acceptor in many severely septic patients, whereas in some cases LPS binding shifted largely to a non-HDL lipoprotein fraction that co-eluted according to size with very low-density lipoprotein (VLDL). Preliminary data suggest that these lipoproteins have a very low density, and they contain apolipoprotein E and higher than normal proportions of the total lipoprotein cholesterol, phospholipid, apolipoprotein B, and serum amyloid A. The data suggest that the VLDL fraction contains acute phase lipoproteins of significantly altered composition that can replace HDL as the dominant LPS acceptor during sepsis when HDL levels are low.
Lessons from an LPS-deficient Neisseria meningitidis mutant
Authors: van der Ley Peter , Steeghs Liana
Published: June 2007

In the pathogen Neisseria meningitidis, a completely LPS-deficient but viable mutant can be obtained by insertional inactivation of the lpxA gene, encoding UDP-GlcNAc acyltransferase required for the first step of lipid A biosynthesis. The expression and assembly of integral outer membrane proteins in the absence of LPS is largely unaffected. However, the expression of iron limitation-inducible, cell surface-exposed lipoproteins is greatly reduced. Major changes were seen in the phospholipid composition, with a shift towards PE and PG species containing mostly shorter chain, saturated fatty acids. The presence of the capsular polysaccharide turned out to be essential for viability without LPS. The immunogenicity of outer membrane proteins in mice was greatly reduced for the LPS-deficient mutant, showing the importance of LPS as an internal adjuvant in such vaccines. Stimulation of MM6 cells and peripheral blood mononuclear cells showed that induction of TNF-α by whole meningococci was greatly reduced for the LPS-deficient mutant. However, even without LPS the mutant strain could still induce a significant inflammatory response.
Differential modulation of the induction of inflammatory mediators by antibiotics in mouse macrophages in response to viable Gram-positive and Gram-negative bacteria
Authors: Wei Cui  , Lei Mei-Guey , Silverstein Richard , Morrison David C.
Published: June 2007

We have investigated effects of β-lactam antibiotics on TNF-α, and iNOS production from mouse peritoneal macrophages following co-culture with Escherichia coli or Staphylococcus aureus bacteria. Ceftazidime and aztreonam enhanced TNF-α secretion from macrophages stimulated with E. coli; however, imipenem does not alter either the kinetics or magnitude of TNF-α in E. coli -treated macrophages. Similar treatments with S. aureus co-cultured with macrophages markedly altered profiles of TNF-α response characterized by apparent early TNF-α peak relative to untreated S. aureus. All antibiotics increased E. coli-induced iNOS expression as assessed by both mRNA and protein. These same antibiotics significantly reduced S. aureus-induced iNOS levels of RNA. Both ceftazidime and aztreonam enhanced LPS release from E. coli in comparison to low-level LPS release from imipenem-treated bacteria, consistent with observed differences in TNF-α release. Incubation of all three antibiotics with S. aureus similarly increased levels of the cell wall constituent protein A detected in supernatants at early time points indicating microbial lysis. In parallel, S. aureus culture supernatants from 2-h incubation with antibiotics enhanced TNF-α release. These results indicate that different cellular mechanisms contribute to antibiotic-mediated regulation of TNF-α and iNOS secretion in mouse macrophages in response to E. coli versus S. aureus.
Role of lipopolysaccharide (LPS) in asthma and other pulmonary conditions
Authors: Michel Olivier
Published: June 2007

Endotoxin significantly contaminates house dust and is an enhancing factor for asthma severity. Natural exposure to endotoxin in early life could influence immune development and protect from the risk of developing atopy. This article will focus on published data showing that home environmental contamination by endotoxin can participate in chronic airways diseases, in particular asthma.
Molecular basis for lipopolysaccharide mimetic action of Taxol™ and flavolipin
Authors: Kawasaki Kiyoshi , Gomi Kazunori , Kawai Yohko , Shiozaki Masao , Nishijima Masahiro
Published: June 2007

We previously reported that Taxol™, which mimics the action of LPS on murine macrophages, induces signals via mouse TLR4/MD-2, but not via human TLR4/MD-2. Here we investigated the molecular basis for this species-specific action of Taxol™. Expression of mouse MD-2 conferred both LPS and Taxol™ responsiveness on HEK293 cells expressing mouse TLR4, whereas expression of human MD-2 conferred LPS responsiveness alone, suggesting that MD-2 is responsible for the species-specificity of Taxol™ responsiveness. Furthermore, mouse MD-2 mutants, in which Gln-22 was changed to other amino acids, showed dramatically reduced ability to confer Taxol™ responsiveness, although their ability to confer LPS responsiveness was not affected. These results indicated that Gln-22 of mouse MD-2 is essential for Taxol™ signaling, but not for LPS signaling. In this study, we also found that the TLR4/MD-2 complex, together with CD14, mediated signal transduction induced by flavolipin, an amino acid-containing lipid unique to Flavobacterium meningosepticum.
Role of transforming growth factor-β1 (TGF-β1) in endotoxin-induced hepatic failure after extensive hepatectomy in rats
Authors: Yoshimoto Noboru , Togo Shinji , Kubota Toru , Kamimukai Nobuyuki , Saito Shuji , Nagano Yasuhiko , Endo Itaru , Sekido Hitoshi , Nagashima Yoji , Shimada Hiroshi
Published: June 2007

Postoperative infections after hepatectomy sometimes lead to fatal hepatic failure, but the mechanism of the hepatic failure is unclear. Wistar rats underwent 90% hepatectomy, and were then divided into three groups: (i) the SAL group, injected with normal saline; (ii) the LPS group, injected with lipopolysaccharide (LPS) every day for 1 week; and (iii) the LPS plus TGF-Ab (LPS+TGF-Ab) group, injected with LPS with anti-transforming growth factor-β1 (TGF-β1) antibody. We investigated survival rates, TGF-β1 expression in the liver, liver regeneration by proliferating cell nuclear antigen labeling index, hepatocyte apoptosis by single stranded DNA labeling index, and perisinusoidal fibrosis using Masson's trichrome staining. The LPS group (30.4%) had a significantly lower survival rate than the SAL group (84%) and tended to be lower than the LPS+TGF-Ab group (49.4%). Liver regeneration in the LPS group was significantly lower than in the other groups. In the LPS group, hepatocyte apoptosis and perisinusoidal fibrosis was significantly more remarkable, and TGF-β1 expression was significantly higher than in the SAL group. TGF-β1 enhanced by LPS plays an important role in the mechanism of hepatic failure by infections after hepatectomy, especially in inhibition of liver regeneration, and induction of hepatocyte apoptosis and perisinusoidal fibrosis.
Legionella pneumophila mediated activation of dendritic cells involves CD14 and TLR2
Authors: Braedel-Ruoff Sibylla , Faigle Marion , Hilf Norbert , Neumeister Birgid , Schild Hansjörg
Published: June 2007

In this study, we analyzed the activation of bone-marrow derived dendritic cells (BMDCs) from mice lacking the cd14-gene with purified Legionella pneumophila lipopolysaccharide and with viable or formalin-killed L. pneumophila .We found that low concentrations of LPS and doses of L. pneumophila that are relevant to infection are dependent on CD14 to activate BMDCs. Higher concentrations of LPS are able to overcome the lack of CD14 indicating that other receptors are involved. We, therefore, included studies using BMDCs from mice lacking functional TLR2 and/or TLR4 molecules. We found that purified L. pneumophila LPS as well as L. pneumophila either viable or formalin-killed are able to activate BMDCs from TLR4-deficient C3H/HeJ mice but fail to activate BMDCs from TLR2-knockout mice. Our data show that not only purified LPS from L. pneumophila but also the microorganism itself stimulate BMDCs via TLR2 and that this stimulation is dependent on CD14 in this mouse model.
Iso-osmolar prehydration shifts the cytokine response towards a more anti-inflammatory balance in human endotoxemia
Authors: Dorresteijn Mirrin J. , van Eijk Lucas T. , Netea Mihai G. , Smits Paul , van der Hoeven Johannes G. , Pickkers Peter
Published: June 2007

Clinical experience suggests that the administration of fluids in human endotoxemia reduces symptoms. In the present study, the effects of a standardised fluid protocol on symptoms, inflammatory and hemodynamic parameters in human endotoxemia are determined. With approval of the local ethics committee, 16 healthy volunteers received 2 ng/kg of Escherichia coli endotoxin (O:113). After an overnight fast, nine subjects received 1.5 l of 2.5% glucose/0.45% NaCl the hour prior to the endotoxin administration and 150 ml/h during the course of the experiment (`prehydrated group'). Seven subjects only received a continuous infusion of 75 ml/h during the experiment (`non-prehydrated group'). The course of inflammatory parameters and symptoms were determined and mean arterial pressure, heart rate and forearm blood flow were measured. In the prehydrated group, TNF-α increased to 522 ± 63 pg/ml (mean ± SEM) while the maximum in the non-prehydrated group was 927 ± 187 pg/ml ( P < 0.04). IL-10 increased similarly in both groups (non-prehydrated 117 ± 18 pg/ml and prehydrated 99 ± 18 pg/ml; P = NS). The prehydrated group had a significantly lower ( P < 0.004) symptom score and recovered sooner ( P = 0.004). Endotoxin-induced changes in hemodynamics revealed no significant differences between groups. We demonstrate that prehydration in experimental human endotoxemia significantly shifts the cytokine balance towards a more anti-inflammatory pattern. This effect is associated with a reduction in symptoms, whereas the changes in hemodynamic parameters are not influenced by prehydration.
Endotoxin: physical requirements for cell activation
Authors: Mueller M. , Lindner B. , Dedrick R. , Schromm A.B. , Seydel U.
Published: June 2007

Lipopolysaccharide (LPS) is the eminent lipid component of the outer leaflet of the outer membrane of Gram-negative bacteria and the major initiator of innate immune response to bacterial infection. Below the critical micellar concentration (CMC), LPS is exclusively present as a monomer. Above this concentration, aggregates are formed. Increasing the concentration beyond the CMC leads to an increase in aggregate concentration, whereas the concentration of monomers remains constant or even decreases. The question how LPS activates immune cells and whether the aggregate or the monomer is the biologically active unit has been and still is controversial. To prepare clearly defined monomeric solutions, we utilized a dialysis set-up consisting of a donor and an acceptor chamber, separated by a dialysis diaphragm with a cut-off of 5 kDa, thus allowing only monomers to pass. Human mononuclear cells (MNCs) were then stimulated with equal concentrations of aggregates and monomers, respectively, of deep rough mutant LPS from Escherichia coli strain F515 (Re LPS) and TNF-α release was determined. In contrast to earlier and very recent work of others, we started with a preparation of aggregate-suspensions and pure monomer-solutions and show that monomers are significantly less active than aggregates in the absence and presence of serum proteins at identical concentrations. In our model, we propose that LPS aggregates are detected by membrane-associated LBP and intercalated into the cell membrane to bring LPS into close proximity to signaling proteins in the membrane, thus finally leading to cell activation. To support this model, we present data showing that LBP is indeed present in or at the cell membrane of human macrophages.
Peptidoglycan recognition in innate immunity
Authors: Dziarski Roman , Gupta Dipika
Published: June 2007

The innate immune system recognizes micro-organisms through a series of pattern recognition receptors that are highly conserved in evolution. Peptidoglycan (PGN) is a unique and essential component of the cell wall of virtually all bacteria, is not present in eukaryotes, and is an excellent target for the innate immune system. Indeed, higher eukaryotes, including mammals, have several PGN recognition molecules, including CD14, Toll-like receptor 2 (TLR2), nucleotide oligomerization domain (Nod)-containing proteins, a family of peptidoglycan recognition proteins (PGRPs), and PGN-lytic enzymes (lysozyme and amidase). These molecules induce host responses to micro-organisms, degrade PGN, or have direct antimicrobial effects.
Peptidoglycan recognition by the Drosophila Imd pathway
Authors: Kaneko Takashi , Golenbock Douglas , Silverman Neal
Published: June 2007

The structural requirements for recognition of peptidoglycan (PGN) by PGRP-LC and activation of the Drosophila IMD pathway are not yet clear. In order to examine this question more carefully, the activity of peptidoglycan from different types of bacteria was compared in cell-based and whole animal assays. Drosophila S2* cells, but not adult flies, responded to Lys-type Micrococcus luteus PGN, but with significantly less potency compared to Dap-type Escherichia coli PGN, while intact Lys-type PGN from Staphylococcus aureus was inactive. After treatment with lysostaphin, which digests the cross-bridging peptides, S. aureus PGN weakly stimulated the IMD pathway, similar to M. luteus PGN. Further digestion with mutanolysin, which creates monomeric PGN fragments, abolished the activity of S. aureus PGN. On the other hand, monomeric E. coli PGN, generated by mutanolysin digestion, was still active but required different isoforms of PGRP-LC for recognition. Polymeric PGN required only PGRP-LCx, while monomeric E. coli PGN required both the PGRP-LCa and PGRP-LCx isoforms. These results suggest that the recognition by PGRP-LCx alone requires polymeric PGN, and that polymeric Dap-type PGN is a more potent PGRP-LCx agonist, compared to Lys-type PGN. These results also suggest that the heteromeric PGRP-LCa/LCx receptor complex recognizes monomeric Dap-type, but not Lys-type, PGN.
Mutations in TLR4 signaling that lead to increased susceptibility to infection in humans: an overview
Authors: Vogel Stefanie N. , Awomoyi Agnes A. , Rallabhandi Prasad , Medvedev Andrei E.
Published: June 2007

In this overview, we will present current information on known mutations in the TLR4 signaling pathway that have been associated with increased susceptibility to disease. To date, mutations in the extracellular domain of TLR4 itself, IRAK-4, NEMO (IKKγ), and IκBα have been identified and profoundly affect the host response to infection.
Structure/function relationships of lipoteichoic acids
Authors: Morath Siegfried , von Aulock Sonja , Hartung Thomas
Published: June 2007

The role of lipoteichoic acids (LTAs) from Gram-positive bacteria as immunostimulatory molecules was controversial for many years, as inadequate preparation methods as well as heterogeneous and endotoxin-contaminated commercial preparations led to conflicting results. An improved purification methodology for LTA now yields potent bioactive and chemically defined material, which is currently being characterized in various models. A synthetic analogue of Staphylococcus aureus LTA has proven the structure/function relationship. The key role of D-alanine esters for the immune response of LTA was confirmed by synthetic derivatives. The glycolipid anchor of LTA plays a central role analogous to the lipid A of LPS. Methodological aspects and criteria for quality assessment of LTA preparations are discussed.
Pro-atherogenic properties of lipopolysaccharide from the periodontal pathogen Actinobacillus actinomycetemcomitans
Authors: Lakio Laura , Lehto Markku , Tuomainen Anita M. , Jauhiainen Matti , Malle Ernst , Asikainen Sirkka , Pussinen Pirkko J.
Published: June 2007

An association between cardiovascular and periodontal disease may be due to lipopolysaccharide (LPS)-promoted release of inflammatory mediators, adverse alterations of the lipoprotein profile, and an imbalance in cholesterol homeostasis. Since periodontopathogenic potential differs between serotypes of a major periodontal pathogen, Actinobacillus actinomycetemcomitans, we studied the pro-atherogenic properties of LPS preparations from serotypes b and d strains on macrophages (RAW 264.7). A. actinomycetemcomitans LPS preparations induced a time-dependent release of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β). LPS induced foam cell formation and cholesteryl ester accumulation from native low density lipoprotein in the following order: A. actinomycetemcomitans strains JP2 (serotype b) > Y4 (serotype b) > IDH781 (serotype d). mRNA expression levels of scavenger receptor class B, type-I, and ATP-binding cassette transporter-1, receptors mediating cholesterol efflux from macrophages, were decreased by LPS preparations. The results suggest that the pro-atherogenic potential of A. actinomycetemcomitans LPS may depend on the infecting strain and correlate with the periodontopathogenic potential of the pathogen.
Nitric oxide induces SOCS-1 expression in human monocytes in a TNF-α-dependent manner
Authors: González-León M. Carmen , Soares-Schanoski Alessandra , del Fresno Carlos , Cimadevila Agata , Goméz-Piña Vanesa , Mendoza-Barberá Elena , García Felipe , Marín Elvira , Arnalich Francisco , Fuentes-Prior Pablo , López-Collazo Eduardo
Published: June 2007

In contrast to the thoroughly characterized mechanisms of positive regulation within cytokine signaling pathways, our knowledge of negative feedback loops is comparatively sparse. We and others have previously reported that IRAK-M down-regulates inflammatory responses to multiple stimuli. In particular, we could show that the nitric oxide (NO) donor, GSNO, induces IRAK-M overexpression in human monocytes. Here we study the expression of another important negative regulator of cytokine signaling, SOCS-1, in human monocytes exposed to GSNO. The NO donor induced significant levels of SOCS-1 mRNA and protein, 6 h and 16 h after stimulation, respectively. Monocytes stimulated with GSNO for longer periods (24 h and 48 h) failed to express IL-6 and IP-10 upon LPS challenge. In addition, and in line with previous reports of NO-mediated induction of TNF-α, we have found that exposure to this cytokine induces SOCS-1 mRNA in human monocytes. A blocking antibody against TNF-α impaired SOCS-1 expression upon GSNO treatment and re-instated IL-6 and IP-10 mRNA levels after LPS challenge in cultures pretreated with the NO donor. We conclude that NO stimulates SOCS-1 overexpression in a pathway at least partially regulated by TNF-α.
Anti-endotoxin properties of a cinnamon bark-derived compound and its effect on the endotoxin shock model
Authors: Azumi Satoko , Tanamoto Ken-ichi
Published: June 2007

An endotoxin inhibitor derived from cinnamon bark was characterized chemically and tested for anti-LPS properties. Chemical analysis suggested that the active center of the inhibitor was in the lipid portion. Upon incubation with LPS molecule, the inhibitor reduced the ability of LPS to induce TNFα and generate nitric oxide from various cells in vitro, and Limulus gelation activity. The lethal toxicity of LPS in galactosamine-sensitized mice and pyrogenicity of LPS in a rabbit model were reduced 1000- and 100-fold by pre-incubation with the inhibitor, respectively. Simultaneous but separate injection of the inhibitor with a lethal dose of LPS also protected the majority of mice. Protection against LPS was seen when the inhibitor was given to mice 1 h before the LPS challenge. Furthermore, the inhibitor significantly suppressed the induction of fever by simultaneous administration with LPS without prior mixing. These results suggest that the inhibitor may be a useful potent blocker of bacterial endotoxin.
Endotoxin-induced cross-tolerance to Gram-positive sepsis
Authors: Guyton Kelly , Bond Robert , Romeo Cristina , Southern Rodney , Cochran Joel , Teti Giuseppe , Cook James A.
Published: June 2007

The manifestations of Gram-positive sepsis and Gram-negative sepsis share some common clinical features suggesting common pathways of activation. The goal of this study was to assess whether lipopolysaccharide (LPS) can produce cross-tolerance to Gram-positive sepsis induced by group B streptococcus (GBS). Thromboxane (TxB2), tumor necrosis factor (TNFα), and nitric oxide (NO) production by in vitro LPS- and heat killed GBS-stimulated rat peritoneal macrophages were measured. Since our previous studies have demonstrated altered macrophage activation of extracellular signal-regulated kinases 1 and 2 (ERK 1/2) in tolerance, we also examined the effect of LPS and killed GBS on ERK 1/2 activation in normal and LPS tolerant macrophages. Tolerance was induced in rats by intraperitoneal injection of Salmonella enteritidis LPS or vehicle for two consecutive days at doses of 0.1 and 0.5 mg/kg body weight. Three days after the second LPS dose, rats were injected intravenously with viable GBS (5 x l09 cfu/kg) and D-galactosamine (1 g/kg). LPS tolerance significantly prolonged ( P <0.05) mean survival time to severe GBS sepsis in D-galactosamine sensitized rats from 12.9 ± 1.7 h in control rats to 44.0 ± 8.9 h in tolerant rats. Peritoneal macrophages from LPS tolerant rats exhibited suppressed LPS induced in vitro TxB2 and TNFα production ( P <0.05). Tolerance also decreased in vitro heat killed GBS-induced TNFα production, but did not significantly affect TxB2 production. NO production stimulated by LPS (10 µg/ml was not impaired in LPS tolerance; however at lower doses (0.02—1.25 µg/ml), NO production was significantly decreased ( P <0.05). NO production was augmented ( P <0.05) in response to stimulation with GBS (10 µg/ml) and unaltered at lower doses (0.02—1.25 µg/ml) in tolerant cells. LPS activated ERK 1/2 in control macrophages, but activation of ERK 1/2 was suppressed in LPS tolerance. GBS did not significantly affect ERK 1/2 activity in control or tolerant macrophages. Nevertheless, the selective mitogen-activated kinase (MAPK)/ERK kinase (MEK) inhibitor, PD 98059 blocked ( P <0.05) both GBS- and LPS-induced TNFα and TxB2 production, but not NO production. Thus, some level of ERK 1/2 activity appears essential for GBS- and LPS-induced macrophage activation. In conclusion, LPS tolerance induces partial cross-tolerance to Gram-positive sepsis induced lethality, and alters LPS- and GBS-induced in vitro peritoneal macrophage mediator production. This suggests common pathways of cellular activation for GBS and LPS that are altered by LPS tolerance.