bluuebunny/crossref_metadata_2025 huggingface dataset:
IκB-ζ, a new anti-inflammatory nuclear protein induced by lipopolysaccharide, is a negative regulator for nuclear factor-κB
Authors: Muta Tatsushi , Yamazaki Soh , Eto Akiko , Motoyama Masaiwa , Takeshige Koichiro
Published: June 2007

Activation of nuclear factor-κ B (NF-κ B), a prominent cellular response to bacterial endotoxin or other microbial products, must be strictly regulated because excessive activation leads to overproduction of cytotoxic cytokines that culminates in septic shock. During screening for genes up-regulated upon inflammation, we identified a new member of the IκB family proteins with the ankyrin-repeats. This protein, designated Iκ B-ζ , is hardly detectable in resting cells, but is strongly induced upon stimulation by lipopolysaccharide, which stimulates cells through the Toll-like receptor 4. Interleukin-1β stimulation also results in the strong induction of IκB-ζ, but tumor necrosis factor-α does not. In contrast to IκB-α or Iκ B-β , IκB-ζ localizes in the nucleus, where it inhibits NF-κB activity. NF-κ B activity is essential for the induction of IκB-ζ, but is not sufficient. Thus, this protein is a new anti-inflammatory protein, which is specifically induced upon inflammation to regulate NF-κB activity.
Endotoxemia and Gram-negative bacteremia as predictors of outcome in sepsis: a meta-analysis using ROC curves
Authors: Hurley James C.
Published: June 2007

The prognostic value of the detection of endotoxin in plasma of patients with suspected Gram-negative sepsis is unclear despite numerous studies. Sixteen studies have reported endotoxemia detection and blood culture results with outcome data for 1937 patients with suspected Gram-negative sepsis. Comparisons of the fatality rates for the subgroups of patients with, respectively, either endotoxemia (group 3), or Gram-negative bacteremia (group 2), or both detected (group 1), versus the fatality rate for the subgroup of patients with these factors absent (group 4), were made based on summary odds ratios (and 95% CI). Summary odds ratios for the comparison of group 2 versus group 4 and group 3 versus group 4 were non-significant (2.2; 0.9-5.8 and 2.0; 0.8-4.8, respectively). The summary odds ratio was significant for group 1 versus group 4 (3.6; 2.1-6.3), and more so among studies in which non-Enterobacteriaceae were common among the Gram-negative blood culture isolates (4.8; 2.1-10.7). Endotoxemia, like Gram-negative bacteremia, is a weak prognostic indicator. Endotoxemia appears to have most prognostic significance when it is detected in the co-presence of Gram-negative bacteremia. The differing interpretations among published studies could be attributed to the variability in types of Gram-negative bacteremia observed.
TLR signaling at the intestinal epithelial interface
Authors: Abreu Maria T. , Thomas Lisa S. , Arnold Elizabeth T. , Lukasek Katie , Michelsen Kathrin S. , Arditi Moshe
Published: June 2007

The intestinal epithelium provides a critical interface between lumenal bacteria and the mucosal immune system. Whereas normal commensal flora do not trigger acute inflammation, pathogenic bacteria trigger a potent inflammatory response. Our studies emanate from the hypothesis that the intestinal epithelium is normally hyporesponsive to commensal pathogen-associated molecular patterns (PAMPs) such as LPS. Our data demonstrate that normal human colonic epithelial cells and lamina propria cells express low levels of TLR4 and its co-receptor MD-2. This expression pattern is mirrored by intestinal epithelial cell (IEC) lines. Co-expression of TLR4 and MD-2 is necessary and sufficient for LPS responsiveness in IEC. Moreover, LPS sensing occurs along the basolateral membrane of polarized IEC in culture. Expression of MD-2 is regulated by IFN-γ. Cloning of the MD-2 promoter demonstrates that promoter activity is increased by IFN-γ and blocked by the STAT inhibitor SOCS3. We conclude from our studies that the intestinal epithelium down-regulates expression of TLR4 and MD-2 and is LPS unresponsive. The Th1 cytokine IFN-γ up-regulates expression of MD-2 in a STAT-dependent fashion. The results of our studies have important implications for understanding human inflammatory bowel diseases.
Receptor cluster formation during activation by bacterial products
Authors: Triantafilou Martha , Triantafilou Kathy
Published: June 2007

The recognition of bacterial products, such as lipopolysaccharide (LPS) by the innate immune system lead to a strong pro-inflammatory response that can eventually lead to fatal sepsis syndrome in humans. Although CD14 and TLR4 have been identified as the key molecules involved in LPSinduced signal transduction, accumulating evidence indicates that multiple receptors are also involved. Our group has recently identified a cluster of receptors, involving heat-shock proteins 70 and 90, chemokine receptor 4 as well as growth differentiation factor 5, that are formed following LPS stimulation. In addition, we present data demonstrating that these molecules associate with TLR4 and accumulate in membrane microdomains following LPS ligation. Our results suggest that the entire bacterial recognition is based around the recruitment of multiple signalling molecules, in addition to CD14 and TLRs, within the lipid rafts. We propose that different combinational associations of receptors within activation clusters determine the different responses to a variety of bacterial stimuli.
Effects of bacterial cell wall components (PAMPs) on the expression of monocyte chemoattractant protein-1 (MCP-1), macrophage inflammatory protein-1α (MIP-1α) and the chemokine receptor CCR2 by purified human blood monocytes
Authors: Møller Anne-Sophie W. , Øvstebø Reidun , Westvik Åse-Brit , Joø Gun Britt , Haug Kari-Bente F. , Kierulf Peter
Published: June 2007

Regulation of chemokine production and the expression of chemokine receptors play an important role during inflammation and infectious diseases. The present study was designed to study the effects of five different bacterial cell wall components (PAMPs) on the production of MCP-1 and MIP-1α and the expression of CCR2 by highly purified human blood monocytes. All five PAMPs induced high expression of mRNA and protein synthesis of both chemokines. Generally, MCP-1 mRNA and protein levels were higher than MIP-1α levels. Expression of MCP-1 and MIP-1α differed both at the mRNA and at the protein levels, MIP-1α always showing a more rapid initial increase, attaining lower protein levels than MCP-1. Antibodies against CD14 significantly inhibited the inducing effects of all the PAMPs used. Antibody against TLR2 inhibited the chemokine production induced by LTA and AraLAM by more than 36% ( P < 0.05) while chemokine production induced by Escherichia coli-LPS, purified E. coli-LPS and Neisseria meningitidis-LPS was inhibited by more than 60% by antibody against TLR4 ( P < 0.05). The inducing effects of all five PAMPs could be inhibited by rIL-4, rIL-10 and rIL-13. rIL-4 was the most effective. Generally, IC50 of these anti-inflammatory cytokines were lower for the MIP-1α than for the MCP-1 production. The cell surface expression of CCR2 was significantly down-regulated by all five PAMPs in addition to a decrease in cytosolic free calcium and binding of rMCP-1. We conclude that MCP-1 and MIP-1α as well as the MCP-1 receptor CCR2 will be substantially regulated upon monocyte contact with various cell wall components (PAMPs) from Gram-negative and Gram-positive bacteria as well as from mycobacteria.
The dual role of LBP and CD14 in response to Gram-negative bacteria or Gram-negative compounds
Authors: Heumann Didier , Lauener Roger , Ryffel Bernard
Published: June 2007

Innate immunity initiates protection of the host organism against invasion of micro-organisms by specific recognition mechanisms. This article reviews the dual role of LBP/CD14 in innate immunity, focusing mostly on experiments performed in mice by the authors. LPS induces uncontrolled pro-inflammatory response that kills the host and is LBP- and CD14-dependent, as neutralization of LBP and CD14 prevents lethal shock. However, surprisingly, the synthetic Pam3CysSerLys4 bacterial lipoprotein from Escherichia coli (BLP), which is well tolerated in mice, kills the mice upon LBP or CD14 blockade. Furthermore, after blockade of LBP and CD14, the mice succumb to a challenge with virulent Klebsiella pneumoniae or Salmonella typhimurium. Therefore, host responses to Gram-negative bacteria are not identical to that of LPS or BLP. When the host is in the presence of virulent Gram-negative bacteria, the invading pathogens must be held in check by the innate immune system until a specific immune response is mounted. Under these conditions, LBP, CD14, and likely Toll-like receptors (TLRs) are a prerequisite to trigger a pro-inflammatory response of macrophages, which is crucial for keeping an infection under control. These studies indicate that we are very far from understanding how the innate system works and more work needs to be done concerning LBP, CD14 or TLRs. Therefore, caution should be the rule about the use of therapeutic approaches to block the pro-inflammatory response in Gram-negative infections.
An overview of clinical studies in healthy subjects and patients with severe sepsis with IC14, a CD14-specific chimeric monoclonal antibody
Authors: Axtelle Tim , Pribble John
Published: June 2007

An overview and discussion of clinical studies with IC14, a chimeric monoclonal antibody directed against human CD14 is presented. These studies include phase 1 trials in: (i) healthy subjects; (ii) healthy subjects challenged with lipopolysaccharide (LPS); and (iii) patients with severe sepsis. The results from clinical studies of IC14 demonstrate this chimeric monoclonal antibody diminishes the systemic inflammatory response directed against LPS and may have potential as a treatment to prevent organ dysfunction in patients with severe sepsis. Furthermore, IC14 treatment was generally well-tolerated in both healthy subjects and severe sepsis patients and did not increase the incidence of secondary infections. Analysis of pharmacology data indicate that higher doses of IC14 are required to saturate membrane-bound CD14 receptors effectively in patients with severe sepsis than in healthy subjects. Finally, the results of non-clinical studies indicate that IC14 should only be administered with concurrent antimicrobial therapy in patients with infection.
Review: The plasma kallikrein/kinin and renin angiotensin systems in blood pressure regulation in sepsis
Authors: Shariat-Madar Zia , Schmaier Alvin H.
Published: June 2007

The hemodynamics of septic shock after endotoxinemiai s influenced by the plasma kallikrein/kinin and the renin angiotensin systems. In recent years, new information has improved understanding of the protein/biologically active peptide interactions between these two systems. The plasma kallikrein/kinin system, more commonly known as the contact system, has undergone a re-evaluation as to how it assembles on cell membranes for physiological and pathophysiological activation and as to its role in Gram-negative sepsis. It has been proposed that it counterbalances the plasma renin angiotensin system. Furthermore, more knowledge about the renin angiotensin system has become available on how it either opposes the actions of the kallikrein/kinin system or, in some cases, summates with it. Understanding the interactions between these two systems may lead to development of better pharmacological treatments for endotoxin-induced shock.
Molecular cloning and characterization of mouse LITAF cDNA: role in the regulation of tumor necrosis factor-α (TNF-α) gene expression
Authors: Bolcato-Bellemin Anne-Laure , Mattei Marie-Genevieve , Fenton Matthew , Amar Salomon
Published: June 2007

The inflammatory response to bacteria and bacterial products, such as lipopolysaccharides (LPSs), is mediated by a variety of secreted factors, but cytotoxic effects of LPS have been ascribed to the tumor necrosis factor alpha (TNF-α) activity. TNF-α is probably the most pleiotropic cytokine and, given the deleterious effects to the host of this factor, it has been postulated that its expression must be tightly regulated. Our laboratory has recently isolated, cloned and characterized a novel human transcription factor named LITAF or LPS-induced TNF-alpha factor. The present study reports the isolation, cloning and characterization of the mouse LITAF cDNA. Chromosomal localization revealed that mouse LITAF mapped to mouse chromosome 16, in a region highly homologous with the area on which human LITAF was previously located. Northern blot analysis shows that mouse LITAF is already expressed at embryonic day 7 of development, and is highly expressed in adult liver, heart and kidney. Moreover, upon LPS stimulation, we show that: (i) LITAF expression is increased in a mouse monocyte/macrophage cell line; and (ii) TNF-α expression is reduced in ES cell-derived macrophages lacking one copy of LITAF gene. Taken together, these results highlight the important role of LITAF in the regulation of TNF-α gene expression and suggest a potential role of LITAF in mouse organogenesis.
Endotoxin induces leukocyte transmigration and changes in permeability of the airway epithelium via protein-kinase C and extracellular regulated kinase activation
Authors: Serikov Vladimir B. , Choi Hyon , Schmiel Kenneth , Skaggs Cody , Fleming N.W. , Wu Reen , Widdicombe Jonathan H.
Published: June 2007

Lipopolysaccharide (LPS) endotoxin of Gram-negative bacteria compromises the integrity of the airway epithelial barrier and initiates migration of leukocytes across the epithelium. The goal of the present study was to identify the role of extracellular regulated kinase (ERK1/2) transduction pathways in these processes. The first aim was to determine whether LPS induces ERK1/2 activation and changes in epithelial permeabilityin epithelial cells alone or only in the presence of immune cells. The second aim was to determine whether the changes in the epithelial permeability were diminished by ERK1/2 blockade. The third aim was to investigatethe role of protein kinase C (PKC) activation as an upstream event in activation of ERK1/2. In vitro 20 μg/ml LPS challenge reduced epithelial barrier function, and induced ERK1/2 phosphorylation in primary cultures of bovine tracheal epithelium and in the transformed human airway epithelial cell line, Calu-3. LPS initiated migration of neutrophil-like and monocyte-like transformed HL-60 cell across sheets of Calu-3 cells. The migration rate and the associated changes in the electrical resistance, permeability to albumin, and ERK1/2 phosphorylation were all blocked by calphostin C, the specific blocker of PKC and by PD98059 (2'-amino-3'methoxyflavone), a selective cell-permeable inhibitor of MAP kinase kinase. In rats, in vivo perfusion of the lumen of an isolated segment of trachea with LPS (0.1 mg/ml) initiated migration of neutrophils and increased the permeability to albumin. Again, these effects were markedly inhibited by PD98059 and calphostin C (by > 50%). We conclude that epithelial ERK1/2 is activated by endotoxin via PKC and is an important pathway in regulation of epithelial permeability.
A novel assay to detect nucleotide receptor P2X7 genetic polymorphisms influencing numerous innate immune functions
Authors: Denlinger Loren C. , Schell Kathleen , Angelini Giuditta , Green Dawn , Guadarrama Arturo , Prabhu Usha , Coursin Douglas B. , Hogan Kirk , Bertics Paul J.
Published: June 2007

The importance of accessory signaling pathways amplifying endotoxin responses has recently been highlighted by genetic studies describing LPS-hyporesponsive individuals despite carrying the common allele for TLR4. The nucleotide receptor P2X7 modulates the production of numerous LPS-stimulated inflammatory mediators. We have recently described the largest phenotypic screen known for genetic polymorphisms associated with the nucleotide receptor P2X7, a global regulator of leukocyte function. This required the development of a novel monocyte pore assay with numerous advantages over previous methods and with the potential to facilitate rapid (< 3 h), multiplex analysis of clinical samples. This paper addresses aspects pertinent to the development of the monocyte pore assay, briefly summarizes our results suggestingthat P2X7 alleles modulate LPSstimulated cytokine production, and discusses a model wherein P2X7 may serve as an amplification loop of innate immunity.
rBPI(10—193) is secreted by CHO cells and retains the activity of rBPI21
Authors: Horwitz Arnold H. , Ammons W. Steve , Bauer Robert J. , Dedrick Russell , Nadell Rossana , Williams Robert E. , Liu Pei-Syan
Published: June 2007

rBPI23, a recombinant N-terminal fragment of human bactericidal/permeability-increasing protein (BPI), kills Gram-negative bacteria and neutralizes endotoxin. rBPI 21, a variant in which cysteine 132 is changed to alanine, retains the activities of rBPI23. Analysis of certain purified rBPI 21 preparations revealed that some of the molecules had lost nine amino acids from the amino terminus. To explore the effect of this modification on structure and activity, we cloned and expressed a variant of rBPI 21, designated rBPI(10-193), which lacks the first nine amino acids. A monoclonal antibody believed to recognize the amino terminus of rBPI 21 cross-reacted with rBPI21, but not with rBPI(10-193) or full length recombinant BPI (rBPI). These results demonstrated that the antibody recognizes the first nine amino acids of rBPI21 and that this region of the holoprotein (rBPI) is inaccessible to the antibody (as suggested by the known 3-D structure). Purified rBPI(10193) and rBPI 21 were similarly potent in in vitro assays measuring bactericidal, endotoxin binding and neutralization activities. In a mouse model of lethal bacteremia, rBPI(10-193) and rBPI21 were similarly potent whereas in a mouse endotoxin challenge model, rBPI(10-193) appeared to be at least 2-fold more potent than rBPI21. In conscious rats, a rapid bolus dose of 40 mg/kg of rBPI21 caused a significant transient decrease in blood pressure while the same dose of rBPI(10-193) caused no blood pressure decrease. We conclude from these studies that the first nine amino acids of rBPI21 are not essential for the anti-infective and endotoxin-neutralizing activities of BPI.
Enzymology of lipid A palmitoylation in bacterial outer membranes
Authors: Bishop Russell E. , Lo Eileen I. , Adil Khan M. , El Zoeiby Ahmed , Jia Wenyi
Published: June 2007

The enzymology of palmitate addition to lipid A can be traced to the early discovery of monosaccharide lipid A precursors, but the functional importance of lipid A palmitoylation in bacterial resistance to the host immune response has emerged only recently. Lipid A palmitoylationin enterobacteriais determined by a PhoP/PhoQ-activated gene pagP, which encodes an unusual outer membrane enzyme of lipid A biosynthesis. PagP structure and dynamics have now been elucidated by both NMR spectroscopy and X-ray crystallography. PagP is an 8-stranded antiparallel β-barrel preceded by an N-terminal amphipathic α-helix. The PagP barrel axis is uniquely tilted by 30° with respect to the membrane normal. An interior hydrophobic pocket in the upper half of the molecule functions as a hydrocarbon ruler, which allows the enzyme to distinguish palmitate from other acyl chains found in phospholipids. Internalization of a phospholipid palmitoyl group within the barrel appears to occur by lateral diffusion from the outer leaflet through non-hydrogen bonded regions between β-strands. The MsbA-dependent trafficking of lipids from the inner membrane to the outer membrane outer leaflet is necessary for lipid A palmitoylation in vivo. Efforts to determine the PagP catalytic mechanism may lead to the development of inhibitors for the treatment of infections.
The nexus between systemic inflammation and disordered coagulation in sepsis
Authors: Opal Steven M.
Published: June 2007

The innate immune system is designed as an early defense system that recognizes and clears potential microbial invaders. The coagulation system co-evolved with the innate immune system from a common ancestral cellular origin, and these two systems continue to exhibit a remarkable degree of integration in their signaling pathways and regulatory circuits following tissue injury and microbial invasion. Inflammatory mediators generate procoagulant signals and intravascular thrombosis activates multiple components of the innate immune system. The success of recombinant human activated protein C for the treatment of human septic shock is likely attributable to its combined effects as an endogenous anticoagulant and an anti-inflammatory molecule. Further advances in the management of severe infections may accrue with a better understanding of the integral link between coagulation and innate immunity.
C-reactive protein: a predominant LPS-binding acute phase protein responsive to Pseudomonas infection
Authors: Ng Patricia M.L. , Zhenxiao Jin  , Tan Sandra S.H. , Ho Bow , Ding Jeak L.
Published: June 2007

As a structural component of the outer membrane of Gram-negative bacteria, endotoxin, also known as lipopolysaccharide (LPS) exhibits strong immunostimulatory properties, rendering it a pivotal role in the pathogenesis of Gram-negative septicaemia. Our attempt to identify LPS-binding proteins from the hemolymph of the horseshoe crab led to the isolation and identification of C-reactive protein (CRP) as the predominant LPS-recognition protein during Pseudomonas infection. CRP is an evolutionarily ancient member of a superfamily of `pentraxins'. It is a major protein in acute phase of infection in humans. Our investigation of CRP response to Pseudomonas aeruginosa unveiled a robust innate immune system in the horseshoe crab, which displays rapid suppression of a dosage of 106 CFU of bacteria in the first hour of infection and effected complete clearance of the pathogen by 3 days. Such a high dose would have been lethal to mice. Full-length CRP cDNA was cloned. Analysis of the untranslated regions suggests their crucial role in post-transcriptional regulation of CRP transcript levels. Northern blot analysis demonstrated an acute up-regulation of CRP by about 60-fold in 6—48 h of Pseudomonas infection. Taken together, our results provide new insights into the importance of CRP as a conserved molecule for pathogen recognition.
A descriptive study of the variation in baseline levels of anti-endotoxin core antibodies between US and UK populations
Authors: Down J.F. , Barclay G.R. , Bennett-Guerrero E. , Hamilton-Davies C. , Stephens R. , Grocott M.P.W. , Mythen M.G.
Published: June 2007

Low levels of naturally occurring antibodies to the core section of endotoxin (EndoCAb) have been shown to be predictors of poor outcome following major surgery. We performed a retrospective study comparing pre-operative levels in US surgical patients, UK surgical patients and healthy volunteers. Both IgM and IgG EndoCAb levels were higher in the US surgical patients when compared with the other groups (approximately twice as high in the case of IgG EndoCAb). This may reflect genetic or environmental variability between the patient groups, differences in the disease processes, the disparity in the delivery of health care between the two countries or degradation of the samples in transfer.
Porphyromonas gingivalis fimbriae are pro-inflammatory but do not play a prominent role in the innate immune response to P. gingivalis
Authors: Graves Dana T. , Naguib Ghada , Huafei Lu  , Desta Tesfahun , Amar Salomon
Published: June 2007

The fimA gene encodes the major fimbrial protein of Porphyromonas gingivalis. It has been shown to stimulate adhesion to salivary proteins and other bacteria. It is also thought to play a major role in invading and stimulating host cells. To determine whether the fimA gene represents one of the principal molecules of P. gingivalis that induces inflammation, we tested purified FimA protein and a mutant P. gingivalis (DPG3) that lacks the fimA gene versus wild-type (WT) P. gingivalis. When injected into connective tissue of the scalp, purified FimA protein induced TNF-α and MIP-2 expression confirming that it is pro-inflammatory. WT P. gingivalis induced TNF-α expression and recruitment of PMNs in the same model. However, DPG3 P. gingivalis stimulated TNF expression and PMN recruitment to the same extent. The latter was consistent with similar induction of the chemokine MIP-2. Similar results were obtained with diabetic mice that have a more prolonged inflammatory response to bacterial stimulation. These results indicate that FimA is a potent inducer of inflammatory cytokine expression but, in the context of P. gingivalis infection, it is not a principal stimulator of the innate host response.
Invited review: Detoxifying endotoxin: time, place and person
Authors: Munford Robert S.
Published: June 2007

Animals that cannot sense endotoxin may die if they are infected by Gram-negative bacteria. Animals that sense endotoxin and respond too vigorously may also die, victims of their own inflammatory reactions. The outcome of Gram-negative bacterial infection is thus determined not only by an individual's ability to sense endotoxin and respond to its presence, but also by numerous phenomena that inactivate endotoxin and/or prevent harmful reactions to it. Endotoxin sensing requires the MD-2/TLR4 recognition complex and occurs principally in local tissues and the liver. This review highlights the known detoxification mechanisms, which include: (i) proteins that facilitate LPS sequestration by plasma lipoproteins, prevent interactions between the bioactive lipid A moiety and MD-2/TLR4, or promote cellular uptake via non-signaling pathway(s); (ii) enzymes that deacylate or dephosphorylate lipid A; (iii) mechanisms that remove LPS and Gram-negative bacteria from the bloodstream; and (iv) neuroendocrine adaptations that modulate LPS-induced mediator production or neutralize pro-inflammatory molecules in the circulation. In general, the mechanisms for sensing and detoxifying endotoxin seem to be compartmentalized (local versus systemic), dynamic, and variable between individuals. They may have evolved to confine infection and inflammation to extravascular sites of infection while preventing harmful systemic reactions. Integration of endotoxin sensing and detoxification is essential for successful host defense.
The antimicrobial properties of C-reactive protein (CRP)
Authors: Tan Sandra S.H. , Ng Patricia M.L. , Ho Bow , Jeak Ling Ding
Published: June 2007

C-reactive protein, CRP, is a predominant pattern-recognition receptor (PRR) in the plasma of the horseshoe crab, which recognizes lipopolysaccharide (LPS). Native CRP2 has previously been shown to exhibit agglutination activity against the polysialic capsule of Escherichia coli K1 but its role in bacterial clearance is not well characterized. In this work, the antimicrobial activity of a recombinant CRP2 isoform (rCRP2) was tested against E. coli, Pseudomonas aeruginosa and Staphylococcus aureus. rCRP2 agglutinates bacteria and exhibits bactericidal activity against Gram-negative bacteria. In addition, the antimicrobial activity of rCRP2 is calcium-independent. GST pulldown experiments suggest that in the naïve physiological state, CRP2 interacts with hemocyanin, native CRPs, a 35-kDa plasma lectin and an as yet unidentified 40-kDa protein. This interaction was enhanced upon Pseudomonas infection. We propose that rCRP2 is a PRR with potent antimicrobial activity and its interacting partners contribute to effective bacterial clearance.
Augmentation of lipopolysaccharide-induced nitric oxide production by α-galactosylceramide in mouse peritoneal cells
Authors: Ito Hiroyasu , Koide Naoki , Morikawa Akiko , Hassan Ferdaus , Islam Shamima , Tumurkhuu Gantsetseg , Mori Isamu , Yoshida Tomoaki , Kakumu Shinichi , Moriwaki Hisataka , Yokochi Takashi
Published: June 2007

The effect of α-galactosylceramide (α-GalCer) on lipopolysaccharide (LPS)-induced nitric oxide (NO) production in mouse peritoneal cells was studied. α-GalCer augmented LPS-induced NO production in mouse peritoneal cells, but not in RAW 264.7 macrophage cells. α-GalCer augmented NO production, but not tumor necrosis factor (TNF)-α production in LPS-stimulated peritoneal cells. Peritoneal cells produced a significant level of interferon (IFN)-γ in response to α-GalCer and anti-IFN-γ antibody abolished the augmentation of LPS-induced NO production by α-GalCer. Moreover, anti-IFN-γ antibody prevented the enhanced expression of an inducible type of NO synthase mRNA by α-GalCer. α-GalCer did not augment LPS-induced NO production in peritoneal cells from natural killer T (NKT)-deficient mice. Therefore, it was suggested that α-GalCer might augment LPS-induced NO production in peritoneal cells through release of IFN-γ from NKT cells.
Reprogramming of circulatory cells in sepsis and SIRS
Authors: Cavaillon J.-M. , Adrie C. , Fitting C. , Adib-Conquy M.
Published: June 2007

Immune status is altered in patients with sepsis or non-infectious systemic inflammatory response syndrome (SIRS). Reduced ex-vivo TNF production by endotoxin-activated monocytes has been regularly reported. This observation is reminiscent of the phenomenon of endotoxin tolerance, and the term `leukocyte reprogramming' well defines this phenomenon. This review will outline that the hyporesponsiveness of circulating leukocytes is not a generalized phenomenon in sepsis and SIRS. Indeed, the nature of the insult ( i.e. infectious versus non-infectious SIRS; under anesthesia [surgery] or not [trauma, burn]), the nature of the activator used to trigger leukocytes ( i.e. different Toll-like receptor ligands or whole bacteria), the nature of the cell culture ( i.e. isolated monocytes versus peripheral blood mononuclear cells versus whole blood assays), and the nature of the analyzed cytokines ( e.g. IL-1β versus IL-1ra; TNF versus IL-10) have a profound influence on the outcome of the response.
The role and regulation of apoptosis in sepsis
Authors: Wesche-Soldato Doreen E. , Lomas-Neira Joanne L. , Perl Mario , Jones Leslie , Chung Chun-Shiang , Ayala Alfred
Published: June 2007

Today, sepsis continues to be a growing problem in the critically ill patient population. A number of laboratories have been interested in understanding how changes in immune cell apoptosis during sepsis appear to contribute to septic morbidity. Consistently, it has been found that immune cell apoptosis is altered in a variety of tissue sites and cell populations both in experimental animals and humans. While divergent mediators, such as steroids and TNF, contribute to some of these apoptotic changes, their effects are tissue and cell population selective. Inhibition of FasL—Fas signaling (by either FasL gene deficiency, in vivo gene silencing [siRNA] or with FasL binding protein) protects septic mice from the onset of marked apoptosis and the morbidity/mortality seen in sepsis. Further, this extrinsic apoptosis response appears to utilize aspects of the Bid-induced mitochondrial pathway. This is in keeping with the findings that pan-specific caspase inhibition or the overexpression of Bcl-2 also protect these animals from the sequellae of sepsis.
Evidence for the requirement for CD40—CD154 interactions in resistance to infections with attenuated Salmonella
Authors: Fernandez-Cabezudo Maria J. , Ullah Azim , Flavell Richard A. , al-Ramadi Basel K.
Published: June 2007

Salmonella species include facultative intracellular pathogens which reside preferentially within cells of the host's reticulo-endothelial system. Resistance to Salmonella involves a collaboration between cells of the innate and adaptive immune systems, and protective immunity requires cell-mediated and humoral-immune responses. CD40—CD154 interactions are of central importance in the induction of cellular immune responses. In the present study, CD154-deficient (CD154—/—) mice were used to assess the role of CD40—CD154 interactions in immunity to Salmonella infection. Compared to C57BL/6 (CD154+/+) controls, CD154 —/—mice were hypersusceptible to infection by an attenuated strain of Salmonella enterica serovar Typhimurium ( S. typhimurium), as evidenced by a significantly decreased survival rate. CD154—/— mice exhibited a defect in the production of IFN-γ and NO in the acute phase of the disease, which resulted in a failure to control bacterial replication. We conclude that intercellular communications via the CD40—CD154 pathway play a critical role in the induction type-1 cytokine responses and protection against primary infections with attenuated Salmonella.
Synthesis of lipid A and its analogues for investigation of the structural basis for their bioactivity
Authors: Fujimoto Yukari , Adachi Yo , Akamatsu Masao , Fukase Yoshiyuki , Kataoka Mikayo , Suda Yasuo , Fukase Koichi , Kusumoto Shoichi
Published: June 2007

As a step to elucidate the structural requirements for the endotoxic and antagonistic activity of lipid A derivatives, we have focused, in the present study, on the effects of the acyl moieties and acidic groups at the 1- and 4′- positions. We synthesized a new analogue corresponding to Rubrivivax gelatinosus lipid A, which has a characteristic symmetrical distribution of acyl groups on the two glucosamine residues with shorter acyl groups (decanoyl groups [C10] and lauryl groups [C12]) than Escherichia coli lipid A. Carboxymethyl analogues in which one of the phosphates was replaced with a carboxymethyl group were also synthesized with different distribution of acyl groups. Biological tests revealed that the distribution of acyl groups strongly affected the bioactivity. The synthetic Ru. gelatinosus type lipid A showed potent antagonistic activity against LPS, whereas its 1- O-carboxymethyl analogue showed weak endotoxic activity. These results demonstrated that when the lipid A has shorter (C10, C 12) hexa-acyl groups, the bioactivity of lipid A is easily affected with small structural difference, such as the difference of acidic group or the distribution of acyl groups, and the bioactivity changes from endotoxic to agonistic or vice versa at this structural boundary for the bioactivity. We also designed, based on molecular mechanics calculations, and synthesized lipid A analogues possessing acidic amino acid residues in place of the non-reducing end phosphorylated glucosamine. Definite switching of the endotoxic or antagonistic activity was also observed depending on the difference of the acidic groups (phosphoric acid or carboxylic acid) in the lipid A analogues.
Reduced toxicity of lipo-oligosaccharide from a phoP mutant of Neisseria meningitidis: an in vitro demonstration
Authors: Rustam Tarick , McClean Stephen , Newcombe Jane , McFadden Johnjoe , Eales-Reynolds Lesley-Jane
Published: June 2007

PhoP is part of a two-component regulatory system, which we have previously demonstrated in Neisseria meningitidis and shown to be an important regulator of virulence in an in vivo model. The phoP mutant clearly induced cross-species reactive antibodies and lacks the obvious toxic effects of the wild-type strain. In the current study, we demonstrate distinct differences between the wild-type and mutant strains in an in vitro model of toxicity. At concentrations likely to be present early in an infection, the mutant was more efficient at stimulating an inflammatory response than the wild-type. However, at the concentrations likely to be found at the site of a fulminant infection, the mutant showed significantly weaker ability to stimulate the release of pro-inflammatory cytokines and the production of reactive oxygen and nitrogen intermediates. SDS-PAGE analysis of the isolated LOS from the wild-type and mutant showed a difference in the level of expression of two major species of LOS, a finding which was supported by preliminary MALDI-TOF analysis. These results suggest that the altered toxicity of the mutant may be due to the increased expression of a conformationally altered LOS species, which shows less affinity and avidity for the cellular receptors responsible for the inflammatory response to endotoxin.
TLR4 agonists as immunomodulatory agents
Authors: Alderson Mark R. , McGowan Patrick , Baldridge Jory R. , Probst Peter
Published: June 2007

Monophosphoryl lipid A (MPL®) is a potent vaccine adjuvant derived from Salmonella minnesota that was recently licensed in Europe as a component of an improved vaccine for hepatitis B (Fendrix®). MPL, like lipopolysaccharide from which it is derived, signals via the TLR4/MD-2 complex. We have produced a series of synthetic Toll-like receptor 4 (TLR4) agonists that are based upon the structure of the major hexa-acylated congener contained within MPL. These TLR4 agonists, termed the aminoalkyl glucosaminide phosphates (AGPs), stimulate the production of various cytokines by human peripheral blood mononuclear cells in vitro and up-regulate cell surface markers on monocytes, NK cells and B cells. In addition, AGPs provide non-specific resistance to challenge with viral and bacterial pathogens when administered to the upper airways of mice. Structure—activity relationship studies have shown that the activation of innate immune effectors by AGPs depends primarily on the length of the secondary acyl chains and the nature of the functional group attached to the aglycon component. Moreover, AGPs can act as potent adjuvants for mucosal administration of vaccine antigens, enhancing both antigen-specific antibody and cell-mediated immune responses. Thus, by combining the adjuvant and non-specific resistance induction properties of AGPs it may be possible to generate mucosal vaccines that provide innate protection immediately following administration together with long-term acquired immunity.
Pretreatment of curcumin attenuates coagulopathy and renal injury in LPS-induced endotoxemia
Authors: Chen Hsiang-Wen , Kuo Hung-Tien , Chai Chee-Yin , Ou Jian-Liang , Yang Rei-Cheng
Published: June 2007

Disseminated intravascular coagulation (DIC) is a lethal situation in severe infections, characterized by the systemic formation of microthrombi complicated with bleeding tendency and organ dysfunction. Current clinical trials are not promising. In this study, we investigated the protective effect of curcumin in a lipopolysaccharide (LPS)-induced DIC model in rats. Experimental DIC was induced by sustained infusion of LPS (10 mg/kg body weight) for 4 h through the tail vein. Curcumin (60 mg/kg body weight) was given intraperitoneally 3 h before LPS infusion. Results showed that, in vivo, curcumin reduced the mortality rate of LPS-infused rats by decreasing the circulating TNF-α levels and the consumption of peripheral platelets and plasma fibrinogen. Furthermore, in vivo curcumin also has the effect of preventing the formation of fibrin deposition in the glomeruli of kidney. These results reveal the therapeutic potential of curcumin in infection-related coagulopathy of DIC.
Investigation of the chemical structure and biological activity of oligosaccharides isolated from rough-type Xanthomonas campestris pv. campestris B100 lipopolysaccharide
Authors: Kaczynski Zbigniew , Braun Sebastian , Lindner Buko , Niehaus Karsten , Holst Otto
Published: June 2007

The rough-type lipopolysaccharide (LPS) of the phytopathogenic bacterium Xanthomonas campestris pv. campestris B 100 was isolated utilizing the hot phenol-water method and successively de-acylated by treatment with hydrazine and hot potassium hydroxide. Four compounds were separated by preparative high-performance anion-exchange chromatography and studied by sugar analysis and by 1D and 2D homonuclear and heteronuclear 1H-, 13C- and 31P-NMR spectroscopy as well as ESI FT-MS. The two main products were a heptasaccharide and a pentasaccharide of the structures α-D-Man p-(1→3)-α-D-Man p-(1→4)-β-D-Glc p-(1→4)-α-D-Man p-3 P -(1→5)-α-Kdo-(2→6)-β-D-Glc pN-4 P-(1→6)-α-D-Glc pN-1 P (1) and β-D-Glc p-(1→4)-α-D-Man p-3 P-(1→5)-α-Kdo-(2→6)-β-D-Glc pN-4 P-(1→6)-α-D-Glc pN-1 P (2), respectively. The products in smaller amounts were a heptasaccharide and pentasaccharide possessing the above structures plus a phosphate group at C-4 of the Kdo residue (compounds 3 and 4). Both, heptasaccharide 1 and pentasaccharide 2 were able to induce an oxidative burst in cell cultures of the non-host plant tobacco.
LPS-induced mucin expression in human sinus mucosa can be attenuated by hCLCA inhibitors
Authors: Hauber Hans-Peter , Goldmann Torsten , Vollmer Ekkehard , Wollenberg Barbara , Hung Hsiao-Ling , Levitt Roy C. , Zabel Peter
Published: June 2007

Background: hCLCA1 is a member of the calcium-activated chloride channel family and is associated with disease-inducible mucus expression. Niflumic acid (NFA) and a closely related chemical structure are reported inhibitors of calcium-activated chloride channels and endotoxin-inducible mucus expression in the mouse. Therefore, we tested the hypothesis that hCLCA1 may be involved in lipopolysaccharide (LPS) induced mucin up-regulation in human airways. We also investigated the effect of NFA and MSI-2216 on LPS-induced mucin up-regulation.

Materials and Methods : Explanted human airways and the muco-epidermoid cell line Calu-3 were stimulated with LPS. Different concentrations of NFA or MSI-2216 were added to LPS-stimulated airway mucosa and Calu-3 cells. Expression of hCLCA1 and MUC5AC mRNA and protein was quantified in human airways using real-time PCR and PAS staining. In addition, immunohistochemistry was performed for quantification of inflammatory cells (lymphocytes, monocytes, eosinophils, and neutrophils) in the submucosa of the airways. Expression of hCLCA1 protein in Calu-3 cells was analysed by FACS.

Results: LPS significantly induced hCLCA1 and MUC5AC mRNA and protein expression in human airway mucosa ( P < 0.05). NFA and MSI-2216 significantly decreased LPS-induced mucus expression in explanted airway mucosa in a dose-dependent manner ( P < 0.05). In Calu-3 cells, LPS significantly increased hCLCA1 surface expression whereas intracellular expression was significantly decreased ( P < 0.05). In Calu-3 cells, NFA and MSI-2216 also significantly reduced MUC5AC mRNA expression ( P < 0.05).

Conclusions: These data suggest that hCLCA1 may play a role in LPS-induced mucin expression in human airway mucosa. Calcium-activated chloride channel inhibitors significantly decreased LPS-induced mucus expression both ex vivo and in vitro . Therefore, blocking of hCLCA1 may offer a therapeutic approach to reduce bacterial-induced mucus hypersecretion.
Invited review: Priming, induction and modulation of plant defence responses by bacterial lipopolysaccharides
Authors: Newman Mari-Anne , Dow J. Maxwell , Molinaro Antonio , Parrilli Michelangelo
Published: June 2007

Bacterial lipopolysaccharides (LPSs) have multiple roles in plant—microbe interactions. LPS contributes to the low permeability of the outer membrane, which acts as a barrier to protect bacteria from plant-derived antimicrobial substances. Conversely, perception of LPS by plant cells can lead to the triggering of defence responses or to the priming of the plant to respond more rapidly and/or to a greater degree to subsequent pathogen challenge. LPS from symbiotic bacteria can have quite different effects on plants to those of pathogens. Some details are emerging of the structures within LPS that are responsible for induction of these different plant responses. The lipid A moiety is not solely responsible for all of the effects of LPS in plants; core oligosaccharide and O-antigen components can elicit specific responses. Here, we review the effects of LPS in induction of defence-related responses in plants, the structures within LPS responsible for eliciting these effects and discuss the possible nature of the (as yet unidentified) LPS receptors in plants.
Function of CD14 as a peptidoglycan receptor: differences and similarities with LPS
Authors: Dziarski Roman , Gupta Dipika
Published: June 2007

Peptidoglycan (PGN) is a macrophage activator from Gram-positive bacteria. PGN activates cells of hemopoietic origin through CD14 since: (i) PGN-unresponsive CD14-negative cells become PGNresponsive after transfection with CD14 and expression of membrane CD14; (ii) PGN binds to CD14 with high affinity; and (iii) anti-CD14 mAbs inhibit both binding of PGN to CD14 and activation of CD14-positive cells by PGN. However, there are several differences in the function of CD14 as PGN and LPS receptor: (i) the kinetics of binding are different; (ii) the affinity of binding in the absence of LPS-binding protein (LBP) is higher for PGN than LPS; (iii) LBP does not increase the affinity of binding of PGN to CD14 and does not enhance cell activation by PGN (in contrast to LPS); (iv) the regions of CD14 needed for binding and activation are partially similar and partially different for PGN and LPS; (v) sCD14:PGN complexes, in contrast to sCD14:LPS complexes, do not activate CD14-negative cells; (vi) PGN, in contrast to LPS, does not activate CHO cells expressing mCD14; and (vii) PGN and LPS induce differential activation of MAP kinases, but activate similar transcription factors (NF-κB, ATF1/CREB, and AP-1).
Contribution of 252Cf-plasma desorption mass spectrometry to structural analysis of lipids A: examples of non-conservatism in lipid A structure
Authors: Caroff M. , Aussel L. , Zarrouk H. , Perry M.B. , Karibian D.
Published: June 2007

The great majority of lipids A studied so far have the enterobacterial lipid A-type skeleton: a bisphosphorylated glucosamine disaccharide, with fatty acids amidating the 2 amino groups and esterifying the C3 and C3′ positions. Differences between these lipids A occur in the nature and localization of the fatty acids. Such differences have been put forward as a possible taxonomic tool. It is now relatively easy to determine these differences using plasma desorption mass spectrometry (PDMS) if one knows the overall composition of the pure, native lipids A. We have used this technique to compare the lipids A of 2 or more species of several bacterial genera and found considerable conservatism within genera and sometimes between closely related genera ( Salmonella and Escherichia ). An exception was Bordetella of which different species varied in the nature and/or the localization of their fatty acids. B. parapertussis , like B. pertussis, had a single C10OH but at a different location, and quite unusually had a non-hydroxylated fatty acid (C16 ) directly esterifying glucosamine I. On the other hand, 2 strains of B. bronchiseptica had a C12OH in place of the C 10OH of B. pertussis, whereas a third strain replaced the C 12OH by a C12, again a primary non-hydroxylated fatty acid. PDMS has allowed us to conclude that the combined pattern of fatty acids in lipids A is not a reliable taxonomic tool.
LPS induced translocation of NF-κB occurs only in a subpopulation of CD14-positive mononuclear cells
Authors: ten Hove Tessa , Vervoordeldonk Margriet J.B.M. , Dekkers Pascale E.P. , Reitsma Pieter H. , van Deventer Sander J.H.
Published: June 2007

CD14-receptor occupancy is a potent activator of the ubiquitous transcription factor NF-κB. In resting cells, NF-κB exists as a heterodimer in the cytoplasm bound to its inhibitor IκB. Upon stimulation with lipopolysaccharide (LPS), NF-κB translocates to the nucleus. By means of immunohistochemistry, nuclear translocation of the p65 subunit was detected in LPS-stimulated mononuclear cells at the single-cell level. Double-staining experiments showed nuclear translocation of p65 in only a fraction (< 50%) of the LPS-stimulated CD14-positive cells. In vivo, similar results were obtained. In blood from healthy volunteers who were infused with LPS, only a fraction of the CD14-positive cells showed translocation of the p65 subunit. In conclusion, we identified the translocation of NF-κB after LPS stimulation at the single cell level in vitro and in vivo. Surprisingly, only a subpopulation of CD14-positive cells showed translocation. These data indicate that the mere expression of CD14 alone is insufficient for LPS-responsiveness.