bluuebunny/crossref_metadata_2025 huggingface dataset:
Distinct post-receptor alterations generate gene- and signal-selective adaptation and cross-adaptation of TLR4 and TLR2 in human leukocytes
Authors: Liwu Li  , Jacinto Randy , Yoza Barbara , McCall Charles E.
Published: June 2007

Gene- and signal-specific adaptation/tolerance of blood leukocytes to lipopolysaccharide endotoxin (LPS) occurs during human and animal septicemia. These phenotypes can be modeled in vitro. LPS-TLR4-adapted human THP-1 promonocytic cells cross-adapt to lipoteichoic acid (LTA)-TLR2-induced IL-1β/TNF-α production, suggesting disruption of a common intracellular signaling event(s). A plausible explanation for homologous adaptation of TLR4 with heterologous adaptation of TLR2 is a persistent inactivation and degradation of IRAK1 following TLR4 activation. LTA stimulation of TLR2 also produces homologous adaptation of TLR2 with inactivation of IRAK1, but there is no detectable degradation of IRAK1. Strikingly, such LTA-adapted cells still respond to LPS stimulation of TLR4 with rapid activation and degradation of IRAK1, and robust IL-1β/TNFα production. Moreover, cells adapted to either LTA- or LPS-production of IL-1β/TNF-α normally produce soluble interleukin 1 receptor antagonist (sIL-1Ra) anti-inflammatory protein when stimulated by either agonist. We conclude that: (i) disruption of a unique TLR2 signaling component upstream of IRAK1, but downstream of TLR2 sensing, induces homologous adaptation to LTA; (ii) disruption of IRAK1 may induce homologous adaptation of TLR4 to LPS and cross-adaptation of TLR2 to LTA; and (iii) TLR2/TLR4 signaling events that control sIL-1Ra translation do not adapt to LPS or LTA, indicating that TLR4 and TLR2 can still function. We present a hypothetical model of adaptation based on a signalsome, with IRAK1 evolving after IRAK4 to regulate TLR4 adaptation tightly.
Endotoxin boosts the vascular endothelial growth factor (VEGF) in rabbits
Authors: Hahn Robert G.
Published: June 2007

Vascular endothelial growth factor (VEGF) is a cytokine that greatly increases vascular permeability and thereby promotes hypovolemia. The present study examines whether the plasma VEGF concentration is increased by a bolus injection of endotoxin 20 μg/kg in 30 rabbits, and whether the response is modified by vitamin A, which doubles the endotoxin clearance. The results show that endotoxin stimulates a gradual increase in the VEGF concentration to a peak 5 h later which is approximately 100 times higher than the baseline concentration. No statistically significant difference was found between the rabbits that received no further treatment ( n = 10) and the ones that were given 1000 IE/kg of vitamin A intravenously 1 h before ( n = 10) or after ( n = 10) the endotoxin. The rise in VEGF correlated with the development of fever, and the VEGF concentrations were higher in animals with a severely affected physical status as judged by the breathing pattern and changes in posture or reactivity. In conclusion, release of VEGF is part of the cytokine response to endotoxin with a peak occurring 5 h after a bolus injection, and the rise is pronounced also in the presence of a high endotoxin clearance.
Endotoxin tolerance: is there a clinical relevance?
Authors: Cavaillon Jean-Marc , Adrie Christophe , Fitting Catherine , Adib-Conquy Minou
Published: June 2007

Beeson (1946) first defined endotoxin tolerance as a reduced endotoxin-induced fever following repeated injections of typhoid vaccine. Freudenberg and Galanos (1988) demonstrated that endotoxin tolerance that can protect against a lethal challenge of lipopolysaccharide (LPS) involves the participation of macrophages. Evans and Zuckerman (1991) reported a role for glucocorticoids in endotoxin tolerance. Prostaglandins, interleukin-(IL-)10, and transforming growth factor-β are other players of in vivo endotoxin tolerance. Dramatic reduction of plasma tumor necrosis factor (TNF) (Mathison et al. 1990) and other cytokines in response to LPS parallels endotoxin tolerance. The reduced capacity to produce TNF and other cytokines can be mimicked in vitro by pretreatment of monocytes or macrophages with LPS. It is not a specific phenomenon and can be induced by other agents or events. Cross-tolerance between LPS, TLR2 specific ligands, IL-1 and TNF has been regularly reported. A similar loss of LPS-reactivity has been repeatedly reported in leukocytes of septic patients and in patients with non-infectious systemic inflammation response syndrome (SIRS; e.g . surgery, trauma, cardiac arrest and resuscitation, etc.). Studies on cellular signaling within leukocytes from septic and SIRS patients reveal numerous alterations of the activation pathways reminiscent of those observed in endotoxin-tolerant cells. While endotoxin tolerance prevents severity of infections and ischemia-reperfusion damage, it has been suggested that the immune dysregulation observed in SIRS patients was associated with an enhanced sensitivity to nosocomial infections. In conclusion, in vitro and in vivo endotoxin tolerance, either experimental or due to clinical status, are similar but not identical.
Synergistic effects of lipopolysaccharide and interferon-γ in inducing interleukin-8 production in human monocytic THP-1 cells is accompanied by up-regulation of CD14, Toll-like receptor 4, MD-2 and MyD88 expression
Authors: Tamai Riyoko , Sugawara Shunji , Takeuchi Osamu , Akira Shizuo , Takada Haruhiko
Published: June 2007

Lipopolysaccharide (LPS) and interferon (IFN)-γ synergistically induced interleukin-8 (IL-8) production in human monocytic THP-1 cells. IFN-γ-primed THP-1 cells produced higher levels of IL-8 on stimulation with LPS than non-primed cells and the level correlated with duration of priming up to 24 h, although the level of IL-8 induced was most comparable to that induced by co-stimulation with LPS and IFN-γ . Unstimulated THP-1 cells were shown by flow cytometry to be practically devoid of membrane CD14 (mCD14). LPS and IFN-γ enhanced mCD14 and Toll-like receptor (TLR) 4 expression in THP-1 cells, respectively, and co-stimulation with LPS and IFN-γ induced higher levels of mCD14 and TLR4 expression than stimulation with either agent alone. LPS and IFN-γ alone each augmented MD-2 and MyD88 mRNA expression in THP-1 cells, and co-stimulation with LPS and IFN-γ markedly enhanced MD-2 and MyD88 mRNA expression in the cells compared to those with either LPS or IFN-γ alone. Anti-CD 14 and anti-TLR4 monoclonal antibodies almost completely inhibited IL-8 production induced by LPS plus IFN-γ in THP-1 cells. These findings suggest that combined stimulation of THP-1 cells with LPS and IFN-γ up-regulate mCD14, TLR4, MD-2 and MyD88 expression by these cells, which might be involved in synergistic IL-8 production by the cells.
Macrophage recognition of zymosan particles
Authors: Underhill David M.
Published: June 2007

Zymosan particles have served as a model for recognition of microbes by the innate immune system for over 50 years. Zymosan induces inflammatory signals in macrophages through Toll-like receptors TLR2 and TLR6. In addition, phagocytic receptors on macrophages bind zymosan and stimulate particle engulfment. We have further examined the requirements for induction of inflammatory responses such as TNF-α production and NF-κ B activation by zymosan in mouse macrophages. We have observed that direct particle contact is required (excluding a role for soluble components of zymosan preparations) and that contact with a single particle is sufficient to trigger cytokine production. Further, ablation of the Toll-like receptor-stimulating activity of zymosan does not affect the ability of phagocytic receptors to internalise the particle.
Review: Lipopolysaccharide of Burkholderia cepacia complex
Authors: Vinion-Dubiel Arlene D. , Goldberg Joanna B.
Published: June 2007

BurKholderia cepacia complex (Bcc) is a group of phenotypically similar, genetically distinct bacteria that are beneficial to the environment but can also cause severe human infections. Bcc are being exploited for use as bioremediation agents and as a way to combat agricultural plant diseases. However, Bcc can cause lung infections in patients with chronic granulomatous disease or cystic fibrosis often resulting in mortality of these patients. Since it is unclear what bacterial components are necessary for causing human infections, studies of Bcc have focused on identifying putative virulence factors. As in other Gram-negative bacteria, the lipopolysaccharide (LPS) of Bcc induces a strong immune response that can contribute to host cell damage. The unusual structure of Bcc LPS lowers the anionic charge of the Bcc cell surface, which inhibits the binding and subsequent effects of cationic antibiotics. These distinguishing features include the substitution of a Ko for a Kdo residue in the inner core oligosaccharide and Ara4N residues bound to phosphates of the lipid A backbone. The structures of O antigen subunits and the consequent serotypes will also be discussed, with particular reference to the O antigen biosynthetic loci of two Bcc strains.
β-Glucosidase enzymatic activity of crystal polypeptide of the Bacillus thuringiensis strain 1.1
Authors: Papalazaridou A. , Charitidou L. , Sivropoulou A.
Published: June 2007

The crystals of Bacillus thuringiensis strain 1.1 consist of the 140 kDa δ-endotoxin, which exhibits β-glucosidase enzymatic activity, based on the following data. (i) Purified crystals exhibit β-glucosidase enzymatic activity. When the crystals are reacted with specific antibodies directed either against the commercial (almond purified) β-glucosidase or against the 140 kDa polypeptide, then considerable reduction of enzymatic activity is observed almost at the same level with both antibodies. (ii) Commercial β-glucosidase and the 140 kDa crystal polypeptide share antigenic similarities; in Western immunoblots, the 140 kDa crystal polypeptide is recognized by anti-β -glucosidase antibodies, and commercial β-glucosidase is recognized by anti-140-kDa antibodies. (iii) The enzymatic properties of commercial β-glucosidase and that resident in the crystals of B. thuringiensis strain 1.1 are very similar. Thus, both enzymes hydrolyze a wide range of substrates (aryl-β -glucosides, disaccharides with α- or β-linkage polysaccharides) and have an optimum activity at 40°C and pH 5. Both enzymes are relatively thermostable and are resistant to end-product inhibition by glucose. Additionally, they show the same pattern of inhibition or activation by several chemical compounds. (iv) The crystals and commercial β-glucosidase show almost equivalent levels of insecticidal activity against Drosophila melanogaster larvae and, furthermore, cause reduction in adult flies that emerge from larvae surviving treatment.
Pseudomonas aeruginosa lipid A diversity and its recognition by Toll-like receptor 4
Authors: Ernst Robert K. , Hajjar Adeline M. , Tsai Jeff H. , Moskowitz Samuel M. , Wilson Christopher B. , Miller Samuel I.
Published: June 2007

Lipid A is the pro-inflammatory component of bacterial lipopolysaccharide, the major surface component of Gram-negative bacteria. Gram-negative bacteria alter the structure of lipid A in response to specific environmental conditions including those found upon colonization of a host. The opportunistic pathogen Pseudomonas aeruginosa synthesizes a unique hexa-acylated lipid A containing palmitate and aminoarabinose during adaptation to the cystic fibrosis airway. Different lipid A species are observed in P. aeruginosa isolated from non-cystic fibrosis associated infections. Here we report that P. aeruginosa isolates from the airway of a cystic fibrosis patient with severe pulmonary disease synthesized a novel hepta-acylated lipid A. Cystic fibrosis-specific P. aeruginosa lipid A modifications result in resistance to host antimicrobial peptides and increased recognition by human Toll-like receptor 4 (TLR4). Using P. aeruginosa lipid A with different levels of acylation, we identified a 222 amino acid region in the extracellular portion of human TLR4 that is required for the differential recognition of cystic fibrosis-specific lipid A. P. aeruginosa adaptation to the human airway may, therefore, play a fundamental role in the progressive lung damage associated with cystic fibrosis.
Review: Molecular mechanisms of endotoxin tolerance
Authors: Hongkuan Fan  , Cook James A.
Published: June 2007

The phenomenon of endotoxin tolerance has been widely investigated, but to date, the molecular mechanisms of endotoxin tolerance remain to be resolved clearly. The discovery of the Toll-like receptor (TLR) family as the major receptors for lipopolysaccharide (LPS) and other bacterial products has prompted a resurgence of interest in endotoxin tolerance mechanisms. Changes of cell surface molecules, signaling proteins, pro-inflammatory and anti -inflammatory cytokines and other mediators have been examined. During tolerance expression of LPS-binding protein (LBP), CD14, myeloid differentiation protein-2 (MD-2) and TLR2 are unchanged or up-regulated, whereas TLR4 is transiently suppressed or unchanged. Proximal post-receptor signaling proteins that are altered in tolerance include augmented degradation of interleukin-1 receptor-associated kinase (IRAK), and decreased TLR4-myeloid differentiation factor 88 (MyD88) and IRAK-MyD88 association. Tolerance has also been shown to be associated with decreased Gi protein content and activity, decreased protein kinase C (PKC) activity, reduction in mitogen-activated protein kinase (MAP kinase) activity, and reduced activator protein-1 (AP-1) and nuclear factor kappa B (NF-κB) induced gene transactivation. However, not all signaling proteins and pathways are suppressed in tolerance and induction of specific anti-inflammatory proteins and signaling pathways may serve important counter inflammatory functions. The latter include induction of IRAK-M and suppressor of cytokine-signaling-1 (SOCS-1), phosphoinositide-3-kinase (PI3K) signaling, and increased or maintained expression of inhibitor-κB (IκB) isoforms. Also at the nuclear level, increase in the NFκB subunit p50 homodimer expression and increased activation of peroxisome-proliferatoractivated receptors-γ (PPARγ) have been linked to tolerance phenotype. Although there are species and cellular variations in manifestation of the LPS tolerant phenotype, it is clear that the tolerance phenomena have evolved as a complex orchestrated counter regulatory response to inflammation.
Teasing apart structural determinants of `toxicity' and `adjuvanticity': implications for meningococcal vaccine development
Authors: Steeghs Liana , Tommassen Jan , Leusen Jeanette H.W. , van de Winkel Jan G.J. , van der Ley Peter
Published: June 2007

The use of lipopolysaccharide (LPS) as an adjuvant is limited by its high endotoxic activity. In particular, the fatty-acyl pattern of the lipid A part of LPS has been demonstrated to determine its biological activity. By genetic modification of the lipid A biosynthesis pathway in Neisseria meningitidis, a panel of recombinant strains with specific alterations in the lipid A acylation pattern, as well as a strain completely lacking LPS were isolated. Whereas all variations in the fatty-acyl pattern resulted in reduced endotoxic activity, as measured by TNF-α induction in the human macrophage cell line MM6, the adjuvant activity of the modified LPS was, in most cases, barely affected. The in vivo adjuvant properties of N. meningitidis wild-type and mutant LPS was found to correlate with induction of co-stimulatory molecules, in particular CD80 and CD40, and with IL-12 production by LPS-stimulated bone marrow-derived BALB/c dendritic cells in vitro. Our results suggest that the ability of LPS to stimulate pro-inflammatory cytokine induction is not necessarily linked to its adjuvant activity. The availability of this novel set of lipid A variants with improved pharmacological properties will be of great importance for the improvement of future outer membrane vesicle vaccines against N. meningitidis.
LPS, dsRNA and the interferon bridge to adaptive immune responses: Trif, Tram, and other TIR adaptor proteins
Authors: Hoebe Kasper , Beutler Bruce
Published: June 2007

Toll-like receptors (TLRs) expressed on antigen-presenting cells (APCs), form a critical link between innate and the adaptive immune responses. Activation of TLRs by LPS and dsRNA results in up-regulation of co-stimulatory molecules (UCM) essential for the generation of robust T-cell responses. It is now evident that type I interferons (IFNs) play an important role in UCM and in the subsequent maturation of APCs. The recently identified adaptor molecules Trif and Tram, unlike their counterparts MyD88 and MAL/Tirap, induce type I IFN via the TLR4 signaling pathway, whereas Trif appears to be the sole adaptor molecule involved in TLR3 signaling, resulting in subsequent production of type I IFN. Here, we discuss how Trif and type I IFN are involved in the optimization of APC-T cell interaction in response not only to viral but also bacterial stimuli.
Bacterial endotoxin modifies heat shock factor-1 activity in RAW 264.7 cells: implications for TNF-α regulation during exposure to febrile range temperatures
Authors: Singh Ishwar S. , He Ju-Ren , Hester Lisa , Fenton Matthew J. , Hasday Jeffrey D.
Published: June 2007

Recent studies have identified heat shock factor (HSF)-1, the predominant heat/stress-stimulated transcriptional activator of heat shock protein genes as a repressor of certain cytokine genes, including TNF-α and IL-1β. We previously showed that exposing macrophages to febrile-range temperature (FRT; 39.5°C) activates HSF-1 to a DNA binding form that does not activate heat shock protein gene transcription, but apparently represses TNF-α and IL-1β transcription. Prewarming macrophages to 39.5°C for 30 min prior to stimulation with bacterial lipopolysaccharide (LPS) does not change the induction of TNF-α transcription, but markedly reduces its duration. This raised the question of how TNF-α transcription could occur at all in the presence of activated HSF-1. We used RAW 264.7 cells to test the hypothesis that macrophage activation triggers a transient reversal of HSF-1-mediated repression, thereby allowing induction of TNF-α transcription. Electrophoretic mobility shift assays revealed that LPS triggers a transient inactivation of HSF-1 that temporally correlates with TNF-α transcription and was associated with a transient increase in HSF-1 molecular weight, a decrease in its pI, and appearance of HSF-1 phosphorylating activity. The serine/threonine phosphatase inhibitor, calyculin A, blocked the inhibitory affect of FRT on LPS-induced TNF-α generation and prevented the re-activation of HSF-1. We propose that LPS stimulation of FRT-exposed macrophages stimulates a sequential phosphorylation and dephosphorylation of HSF-1, causing a cycle of inactivation and re-activation of HSF-1 repressor activity that allows a temporally-limited period of gene transcription.
TLR4 Asp299Gly and Thr399Ile polymorphisms are very rare in the Chinese population
Authors: Jingqing Hang  , Wei Zhou  , Hongxi Zhang  , Bixiong Sun  , Helian Dai  , Li Su  , Christiani David C.
Published: June 2007

Endotoxin is an important component of bio-aerosols that contribute to airway inflammation and airflow obstruction. Toll-like receptor 4 (TLR4) mediates the host response to bacterial lipopolysaccharide (LPS), and the Asp299Gly and Thr399Ile polymorphisms have been associated with the development of respiratory diseases. We hypothesized that TLR4 polymorphisms may be associated with a decline of lung function in cotton workers who were exposed continuously to endotoxin. We investigated these two polymorphisms in our 20-year longitudinal cohort of Han Chinese cotton and silk textile workers. The two polymorphisms were genotyped using TaqMan and PCR-restriction fragment length polymorphism methods. No homozygous or heterozygous variant genotypes of the Asp299Gly and Thr339Ile polymorphisms were detected in 491 samples of this population. Our results suggest that the TLR4 Asp299Gly and Thr339Ile polymorphisms are very rare in the Chinese population.
Interferon response induced by Toll-like receptor signaling
Authors: Takeuchi Osamu , Hemmi Hiroaki , Akira Shizuo
Published: June 2007

Toll-like receptors (TLRs) are essential for the recognition of distinct pathogen-associated molecular patterns (PAMPs). Activation of TLRs induces intracellular signaling pathways which lead to the production of pro-inflammatory cytokines, chemokines, and interferon (IFN)-inducible genes. TIR domain containing adaptor molecules in turn determine the signaling specificity of the response. Recent studies demonstrated that serine/threonine kinases IKK-i/TBK1 are critical for the regulation of IFN-β as well as IFN-inducible genes. In response to lipopolysaccharide (LPS), transfection of poly(I:C) and viral infection, embryonic fibroblasts (MEFs) derived from TBK1-deficient ( TBK1— /—) mice show impaired production of IFN-inducible genes, but not pro-inflammatory cytokines. Although IKK-i— /— mice show normal production of these genes, MEFs from IKK-i/ TBK1-doubly deficient mice were completely defective in the induction of IFN-β as well as IFN-inducible genes in response to poly(I:C) stimulation. Activation of IFN-regulatory factor (IRF) 3 in response to LPS and poly(I:C) was abolished in IKK-i/TBK1 doubly deficient cells. Interestingly, intracellular transduction of poly(I:C) initiates activation of IFN response in a TLR3-independent manner. These observations demonstrate that IKK-i/TBK1 signaling is essential for both TLR3-dependent and TLR3-independent viral and dsRNA-induced IFN responses.
Endotoxin signaling in human macrophages: signaling via an alternate mechanism
Authors: Sacre Sandra M. , Andreakos Evangelos , Feldmann Mark , Foxwell Brian M.
Published: June 2007

Lipopolysaccharide (LPS) signals through Toll-like receptors (TLRs) in the course of sepsis, resulting in the release of inflammatory factors. In cell lines and murine models, parts of the signaling pathways involved have been elucidated with MyD88, Mal/TIRAP and IKK2 playing an important role in the induction of NF-κB. By focusing on primary human cells, we have shown that there are fundamental signaling differences between human and murine macrophages and between cells of myeloid and non-myeloid origins. In primary human cells, there are no available knockouts so we employed the use of dominant negatives to investigate the signaling cascades. We show that in primary human macrophages MyD88, Mal/TIRAP and IKK2-independent alternative pathways activate NF-κB and induce the expression of inflammatory cytokines, whereas in non-myeloid synovial fibroblasts MyD88 and/or Mal/TIRAP are essential adaptors for LPS signaling.
Mechanisms of TLR9 activation
Authors: Latz Eicke , Visintin Alberto , Espevik Terje , Golenbock Douglas T.
Published: June 2007

Non-methylated CpG-motifs in bacterial or viral DNA are recognized by TLR9 as foreign. The activation of TLR9 by microbial DNA or synthetic oligonucleotides based on these motifs leads to the induction of innate immune responses. We have compared the subcellular localization of fluorescent versions of TLR9 and TLR4 and found that TLR9 is expressed in the endoplasmic reticulum while TLR4 is expressed on the plasma membrane. Fluorescently tagged bacterial DNA or CpG-DNA was observed to traffic to a tubular lysosomal compartment in human pDCs. In stimulated cells, TLR9 translocated to CpG-DNA or microbial DNA containing structures in the endosome, where TLR9 binds to DNA and initiates signaling.
Regulatory role of Toll-like receptor 2 during infection with Trypanosoma cruzi
Authors: Ropert Catherine , Tostes Gazzinelli Ricardo
Published: June 2007

Previous studies have emphasized the role of Toll-like receptors (TLRs) and myeloid differentiation factor 88 (MyD88) during infection with protozoan parasites. TLR2 was shown to be important for induction of cytokine synthesis by macrophages exposed to the purified glycosylphosphatidylinositol (GPI)-anchored mucin-like glycoproteins of Trypanosoma cruzi trypomastigotes (tGPIm). On the other hand, MyD88 —/— mice, but not TLR2 —/— mice, showed impaired cytokine production and resistance to infection with T. cruzi parasites. Here we evaluate the importance of MyD88 and TLR2 in MAPK activation and cytokine synthesis by macrophages exposed to live T. cruzi parasites and compared to tGPIm. The absence of MAPK phosphorylation in TLR2- and MyD88-deficient macrophages exposed to tGPIm correlated with the incapacity to induce cytokine release in these cells. In contrast, activation of MAPK and synthesis of pro-inflammatory cytokines were not abrogated in TLR2-deficient macrophages exposed to live T. cruzi parasites. We also showed that pretreatment with tGPIm significantly reduces cytokine release by macrophages in response to T. cruzi in a TLR2-dependent manner. Consistently, TLR2-/- mice were shown to produce enhanced levels of cytokines upon in vivo challenge with T. cruzi parasites. Together, these results suggest the involvement of additional TLR(s) in the pro-inflammatory response of macrophages to whole parasites, and that, in vivo, TLR2 may have a predominant immunoregulatory role during acute infection with T. cruzi parasites .
Deacylation and palmitoylation of lipid A by Salmonellae outer membrane enzymes modulate host signaling through Toll-like receptor 4
Authors: Kawasaki Kiyoshi , Ernst Robert K. , Miller Samuel I.
Published: June 2007

The Salmonella typhimurium virulence gene products, PhoP/PhoQ sense host micro-environments to regulate the expression of a lipid A 3- O-deacylase, PagL, and a lipid A palmitoyltransferase, PagP. Therefore, deacylation and/or palmitoylation of lipid A could occur in Salmonellae adapted to host environments. The acylation state of lipid A can alter host recognition and signaling by Toll-like receptor (TLR) 4, and may play an important role in host defenses against Salmonellae infection. Deacylated lipid A, deacylated and palmitoylated lipid A, palmitoylated lipid A, and unmodified lipid A species were purified, and the activity was examined using cell lines expressing recombinant human or mouse TLR4. Compared with unmodified lipid A, the modified lipid A species are 10—100-fold less active. These results suggest that PagL and PagP modify lipid A to reduce TLR4-signaling as part of Salmonellae adaptation to the host environment.
Invited review: The dynamics of LPS recognition: complex orchestration of multiple receptors
Authors: Triantafilou Martha , Triantafilou Kathy
Published: June 2007

The molecular mechanisms that have been designed to protect the host from invading pathogens are responsible for sepsis, an often fatal response of the immune system against microbial pathogens. In the past few years, intense research in the field of innate immunity has identified a plethora of pattern recognition receptors that are responsible for bacterial-induced activation. Recognition of bacterial lipopolysaccharide seems to involve a complex orchestration of protein—protein interactions that eventually leads to cellular activation. In this review, we attempt to unravel the dynamic interactions that occur among the different receptors involved and dictate the outcome of the innate immune response.
Evolution and integration of innate immune recognition systems: the Toll-like receptors
Authors: Takeda Kiyoshi
Published: June 2007

Toll-like receptors (TLRs) recognize specific components of micro-organisms and trigger the activation of innate immunity. TLR-mediated signaling pathways are now rapidly being elucidated. TLR signaling originates from the cytoplasmic Toll/IL-1 receptor (TIR) domain, which is conserved among all TLRs. Furthermore, TIR domain-containing adaptors, such as MyD88, TRIF, TIRAP and TRAM, play essential roles in TLR signaling. MyD88 is essential for inflammatory cytokine production via all TLRs, whereas TRIF is involved in TLR3- and TLR4-mediated MyD88-independent induction of IFN-β. Thus, innate immunity represents a skilful system that senses microbial invasion and initiates appropriate immune responses.
Innate immunity of the human newborn: distinct cytokine responses to LPS and other Toll-like receptor agonists
Authors: Levy Ofer
Published: June 2007

Human newborns are at increased risk of microbial invasion and display diminished responses to many vaccines indicating a functional immaturity of the immune system at birth. Such altered immune reactivity may reflect the demands of in utero existence, including the need to avoid potentially harmful inflammatory immune reactions. Despite normal basal expression of Toll-like receptors and membrane CD14, innate immune responses of neonatal mononuclear cells to lipopolysaccharide are characterized by markedly reduced release of the pro-inflammatory Th1-polarizing cytokines TNF-α and interferon-γ with relative preservation of anti-inflammatory Th2-polarizing cytokines. Differences between newborns and adults with respect to TLR-induced TNF-α release extend to a range of TLR agonists, including bacterial lipopeptides, and are due to differences in soluble factors present in blood plasma. Soluble factors in neonatal blood plasma suppress TLR-induced TNF-α release from monocytes and efforts to identify and characterize these inhibitors are on-going. Such altered immunity to TLR agonists is likely to alter both innate and adaptive immune responses in newborns profoundly. Definition of the mechanisms underlying distinct neonatal immunity promises to identify novel ways to prevent and treat infection in this relatively high-risk population.
Monomeric endotoxin:protein complexes are essential for TLR4-dependent cell activation
Authors: Gioannini T.L. , Teghanemt A. , Zhang DeS. , Levis E.N. , Weiss J.P.
Published: June 2007

Potent TLR4-dependent cell activation by Gram-negative bacterial endotoxin depends on sequential endotoxin—protein and protein—protein interactions with LBP, CD14, MD-2 and TLR4. LBP and CD14 combine, in an albumin-dependent fashion, to extract single endotoxin molecules from purified endotoxin aggregates (Eagg) or the bacterial outer membrane and form monomeric endotoxin:CD14 complexes that are the preferred presentation of endotoxin for transfer to MD-2. Endotoxin in endotoxin:CD14 is readily transferred to MD-2, again in an albumin-dependent manner, to form monomeric endotoxin:MD-2 complex. This monomeric endotoxin:protein complex (endotoxin:MD-2) activates TLR4 at picomolar concentrations, independently of albumin, and is, therefore, the apparent ligand in endotoxin-dependent TLR4 activation. Tetra-, penta-, and hexa-acylated forms of meningococcal endotoxin (LOS) react similarly with LBP, CD14, and MD-2 to form endotoxin:MD-2 complexes. However, tetra- and penta-acylated LOS:MD-2 complexes are less potent TLR4 agonists than hexa-acylated LOS:MD-2. This is mirrored in the reduced activity of tetra-, penta- versus hexa-acylated LOS aggregates (LOS agg) + LBP toward cells containing mCD14, MD-2, and TLR4. Therefore, changes in agonist potency of under-acylated meninigococcal LOS are determined by differences in properties of monomeric endotoxin:MD-2.
Differential maturation of murine bone-marrow derived dendritic cells with lipopolysaccharide and tumor necrosis factor-α
Authors: Efron Philip A. , Tsujimoto Hironori , Bahjat Frances R. , Ungaro Ricardo , Debernardis Justin , Tannahill Cynthia , Baker Henry V. , Edwards Carl K. , Moldawer Lyle L.
Published: June 2007

Dendritic cells (DCs) play a key role in the interface between the innate and acquired immune systems. In response to both exogenous as well as endogenous signals, DCs undergo a programmed maturation to become an efficient, antigen-presenting cell. Yet little is known regarding the differential responses by endogenous versus exogenous stimuli on DC maturation. In the present report, we have compared the phenotypic, functional, and genome-wide expression responses associated with maturation by bone marrow derived DCs to either an endogenous danger signal, tumor necrosis factor-α (TNF-α), or a microbial product, bacterial lipopolysaccharide (LPS). Examination of the cell surface expression of DCs as well as cytokine production demonstrated that patterns of DC maturation varied dramatically depending upon the stimulus. Whereas LPS was highly effective in terms of inducing phenotypic and functional maturation, TNF-α exposure produced a phenotypically distinct DC. Gene expression patterns in DCs 6 and 24 h after LPS and TNF-α exposure revealed that these activation signals produce fundamentally different genomic responses. Supervised analysis revealed that the expression of 929 probe sets discriminated among the treatment groups, and the patterns of gene expression in TNF-α stimulated DCs were more similar to unstimulated cells at both 6 and 24 h post-stimulation than to LPS-stimulated cells at the same time points. These findings reveal that DCs are capable of a varying phenotypic response to different antigens and endogenous signals.
Remodeling of Helicobacter pylori lipopolysaccharide
Authors: Tran An X. , Stead Christopher M. , Trent M. Stephen
Published: June 2007

Modification of the lipid A domain of lipopolysaccharide (LPS) has been reported to contribute to the virulence and pathogenesis of various Gram-negative bacteria. The Kdo (3-deoxy-D- manno-octulosonic acid)—lipid A domain of Helicobacter pylori LPS shows several differences to that of Escherichia coli. It has fewer acyl chains, a reduced number of phosphate groups, much lower immunobiological activity, and only a single Kdo sugar is attached to the disaccharide backbone. However, H. pylori synthesizes a minor lipid A species resembling that of E. coli, which is both bis-phosphorylated and hexa-acylated suggesting that the major species results from the action of specific modifying enzymes. This work describes two enzymes, a lipid A phosphatase and a phosphoethanolamine transferase, involved in the periplasmic modification of the 1-position of H. pylori lipid A. Furthermore, we report a novel Kdo trimming enzyme that requires prior removal of the 1-phosphate group for enzymatic activity. Discovery of the enzymatic machinery involved in the remodeling of H. pylori LPS will help unravel the importance of these modifications in H. pylori pathogenesis.
Interactions between LPS and lung surfactant proteins
Authors: Chaby Richard , Garcia-Verdugo Ignacio , Espinassous Quentin , Augusto Luis A.
Published: June 2007

After penetration into the lower airways, bacterial lipopolysaccharide (LPS) interacts with alveolar cells in a fluid environment consisting of pulmonary surfactant, a lipid—protein complex which prevents alveolar collapsing and participates in lung defense. The two hydrophilic surfactant components SP-A and SP-D are proteins with collagen-like and lectin domains (collectins) able to interact with carbohydrate-containing ligands present on microbial membranes, and with defined regions of LPS. This explains their capacity to damage the bacterial envelope and induce an antimicrobial effect. In addition, they modulate LPS-induced production of pro-inflammatory mediators in leukocytes by interaction with LPS or with leukocyte receptors. A third surfactant component, SP-C, is a small, highly hydrophobic lipopeptide which interacts with lipid A and reduces LPS-induced effects in macrophages and splenocyte cultures. The interaction of the different SPs with CD14 might explain their ability to modulate some LPS responses. Although the alveolar fluid contains other antiLPS and antimicrobial agents, SPs are the most abundant proteins which might contribute to protect the lung epithelium and reduce the incidence of LPS-induced lung injury. The presence of the surfactant collectins SP-A and SP-D in non-pulmonary tissues, such as the female genital tract, extends their field of action to other mucosal surfaces.
A designed TLR4/MD-2 complex to capture LPS
Authors: Brandl Katharina , Glück Thomas , Hartmann Pia , Salzberger Bernd , Falk Werner
Published: June 2007

The family of Toll-like receptors (TLRs) is involved in the defense of an organism to microbial attack. TLR4-induced signaling is involved in infectious diseases, chronic inflammatory diseases and sepsis; therefore, we aimed at modulating TLR4-signaling via ligand-binding soluble receptors. Because recognition of microbial structures shows some species-specific traits, we specifically selected the mouse model for later in vivo studies. We first prepared the N-terminally Flag-tagged mouse (m) recombinant (r) soluble (s) fusion proteins mrsTLR4-IgGFc (T4Fc) and mrsMD-2 in Drosophila melanogaster Schneider 2 (S2) cells. The function of these molecules was tested by inhibition of synthesis of pro-inflammatory cytokines after stimulation of mouse macrophage RAW 264.7 cells with purified lipopolysaccharide (LPS). T4Fc alone had no inhibitory activity; however, a T4Fc/MD-2 complex blocked LPS activity. By analogy with `cytokine traps', we then prepared a designer molecule (LPS-Trap) by fusing MD-2 to the C-terminus of soluble TLR4 via a flexible linker. LPS-Trap significantly inhibited TNF production by LPS-stimulated RAW 264.7 cells. Thus, the T4Fc/MD-2 complex as well as the LPS-Trap blocked LPS activity in vitro and might thus represent a new therapeutic option in sepsis by neutralization of TLR4-activating ligands.
Evolution and integration of innate immune systems from fruit flies to man: lessons and questions
Authors: Martinelli Cosimo , Reichhart Jean-Marc
Published: June 2007

Despite broad differences in morphology, ecology and behavior, the fruit fly Drosophila melanogaster and humans show a remarkably high degree of conservation for many molecular, cellular, and developmental aspects of their biology. During the last decade, similarities have also been discovered in some of the mechanisms regulating their innate immune system. These parallels regard mainly the Toll-like receptor family and the intracellular signaling pathways involved in the control of the immune response. However, if the overall similarities are important, the detailed pathogen recognition mechanisms differ significantly between fly and humans, highlighting a complicated evolutionary history of the metazoan innate defenses. In this review, we will discuss the main similarities and differences between the two types of organisms. We hope that this current knowledge will be used as a starting point for a more comprehensive view of innate immunity within the broad variety of metazoan phyla.
Effects of perineural capsaicin treatment of the abdominal vagus on endotoxin fever and on a non-febrile thermoregulatory event
Authors: Petervari Erika , Garami Andras , Pakai Eszter , Szekely Miklos
Published: June 2007

Following perineural capsaicin pretreatment of the main trunks of the abdominal vagus of rats, the first and the second phases of the polyphasic febrile response to intravenous lipopolysaccharide were unaltered, while the third phase of fever course (peak at 5 h) was attenuated. In rats desensitized by intraperitoneal (i.p.) capsaicin (i.e. abdominal non-systemic desensitization), mainly the first but not the later fever phases were reduced. The postprandial hyperthermia to intragastric injection of BaSO4 suspension was attenuated by either i.p. or perineural capsaicin treatment. It is concluded that, in contrast to the accepted model of postprandial hyperthermia, which is mediated by capsaicin-sensitive fibers of the abdominal vagus, in the early phase of polyphasic fever the vagal afferent nerves appear to play no role. The influence of i.p. capsaicin-desensitization on this initiating fever phase is independent of the vagus, and a capsaicin-induced alteration of endotoxin action in the liver, prior to vagal nerve endings, is more likely. The late febrile phase is probably influenced by efferent vagal fibers, which might be damaged more easily by perineural than i.p. capsaicin treatment.
Role of surfactant protein D (SP-D) in innate immunity in the gastric mucosa: evidence of interaction with Helicobacter pylori lipopolysaccharide
Authors: Moran Anthony P. , Khamri Wafa , Walker Marjorie M. , Thursz Mark R.
Published: June 2007

Surfactant protein D (SP-D) is a collagenous glycoprotein, a collectin, which functions as a pathogen-associated molecular pattern (PAMP) recognition receptor in the innate immune response. Although originally identified in the lung as a component of surfactant, SP-D also occurs in the gastric mucosa at the luminal surface and within gastric pits of mucus-secreting cells. Infection with the gastroduodenal pathogen Helicobacter pylori up-regulates expression of SP-D in human patients with gastritis, and its influence on colonization has been demonstrated in a Helicobacter SP-D-deficient (SP-D—/ —) mouse model. SP-D binds and agglutinates H. pylori cells in a lectin-specific manner, and has been shown to bind H. pylori lipopolysaccharide. Furthermore, evidence indicates that H. pylori varies LPS O-chain structure to evade SP-D binding which is speculated aids persistence of this chronic infection.
Review: Structural features and structural variability of the lipopolysaccharide of Yersinia pestis, the cause of plague
Authors: Knirel Yuriy A. , Dentovskaya Svetlana V. , Senchenkova Sof'ya N. , Shaikhutdinova Rima Z. , Kocharova Nina A. , Anisimov Andrey P.
Published: June 2007

Data on the structure and temperature-dependent variations of the lipopolysaccharide (LPS) of Yersinia pestis are summarized and compared with data of other enteric bacteria, including other Yersinia spp. A correlation between the LPS structure and properties of the LPS and bacterial cultures as well as the LPS biosynthesis control are briefly discussed.
Toll-like receptor-dependent discrimination of streptococci
Authors: Santos-Sierra Sandra , Golenbock Douglas T. , Henneke Philipp
Published: June 2007

Streptococcus pneumoniae and Streptococcus agalactiae cause distinct infectious diseases in small children. Similarly, these bacteria elicit very different host-cell responses in vitro. Inactivated S. agalactiae by far exceeds S. pneumoniae in the activation of inflammatory cytokines and upstream signaling intermediates such as the MAP kinase JNK. The inflammatory response to both Streptococcus spp. is mediated by MyD88, an essential adapter protein of Toll-like receptors (TLRs), although the specific TLRs that are involved have not been fully resolved. Furthermore, during logarithmic growth, S. pneumoniae releases pneumolysin that interacts with TLR4 whereas S. agalactiae releases diacylated molecules that interact with TLR2/6. Interaction of these soluble bacterial products with their cognate TLRs is critical for limiting bacterial dissemination and and systemic inflammation in mice. This might be due, in part, to TLR-mediated apoptosis induced by these factors. In conclusion related streptococcal species induce specific events in TLR-mediated signal transduction. Comparative analysis of the host-cell response to these bacteria reveals molecules such as JNK as valuable targets for adjunctive sepsis therapy.
Endotoxin-activated cultured neonatal rat cardiomyocytes express functional surface-associated interleukin-1α
Authors: Westphal Elena , Li Chen  , Pilowski Claudia , Koch Susanne , Ebelt Henning , Müller-Werdan Ursula , Werdan Karl , Loppnow Harald
Published: June 2007

Interleukin-1 (IL-1) is a potent regulator of cardiovascular proliferation, apoptosis, contraction or production of inflammatory mediators. Thus, we investigated expression and function of IL-1 in cultured neonatal rat heart cells upon endotoxin stimulation. We show that cultured neonatal rat cardiomyocytes expressed IL—1α and IL—1β mRNA. The cells expressed functional cell-associated IL—1 activity and a specific anti-IL—1α-antibody inhibited the activity. Biologically active IL—1α was present at the cell surface of the cardiomyocytes, as indicated in co-culture experiments. Immunohistochemistry showed IL—1α-staining of the neonatal cardiomyocytes. Although the cells also expressed IL—1β mRNA, we did not detect IL—1β in the supernatants of cultured cardiomyocytes by ELISA or in immunohistochemical staining. Furthermore, neonatal and adult rat heart tissues expressed IL—1α mRNA, whereas fetal, but not adult, human cardiac tissues expressed detectable IL—1α mRNA. In contrast, IL-1β mRNA was present in rat and human fetal and adult samples. Furthermore, in patients with dilated or ischemic cardiomyopathy, we measured IL—1β, but not IL—1α, mRNA. These results provide evidence for the presence of functionally active IL—1α on the cell surface of neonatal rat cardiomyocytes and may suggest a differential role of IL—1α in regulation of cellular functions during development, aging and disease in rat and human heart cells.
Hemodynamic effects of titrated norepinephrine in healthy versus endotoxemic sheep
Authors: Lange Matthias , Bröking Katrin , Hucklenbruch Christoph , Ertmer Christian , Van Aken Hugo , Lücke Martin , Bone Hans-Georg , Westphal Martin
Published: June 2007

In patients with sepsis and systemic inflammatory response syndrome, hemodynamic support is often complicated by a vascular hyporesponsiveness to exogenously administered norepinephrine. Although norepinephrine tachyphylaxis represents a significant clinical problem, the relationship between norepinephrine dosages and mean arterial pressure (MAP) in the presence of systemic inflammation is still not fully understood. This study was, therefore, designed as a prospective, controlled laboratory trial to elucidate the hemodynamic response to incremental norepinephrine doses in healthy and endotoxemic sheep. ANOVA demonstrated that a significantly higher mean infusion rate of norepinephrine was needed to increase MAP by 20 mmHg in endotoxemic versus healthy control sheep ( P = 0.007). Whereas the goal-MAP was reached in 100% of healthy controls, it was achieved in only 80% during endotoxemia. Cardiac index increased significantly in healthy, but not in endotoxemic, sheep. Our findings confirm the presence of vascular hyporesponsiveness to norepinephrine in endotoxemia. In addition, this study demonstrates that the presence of systemic inflammation leads to an early hyporesponsiveness against norepinephrine which was caused by a drug-independent mechanism rather than a tachyphylaxis due to long-term administration of norepinephrine.
Transcriptional activation of MMP-13 by periodontal pathogenic LPS requires p38 MAP kinase
Authors: Rossa Carlos , Min Liu  , Bronson Paul , Kirkwood Keith L.
Published: June 2007

Matrix metalloprotease-13 (MMP-13) is induced by pro-inflammatory cytokines and increased expression is associated with a number of pathological conditions such as tumor metastasis, osteoarthritis, rheumatoid arthritis and periodontal diseases. MMP-13 gene regulation and the signal transduction pathways activated in response to bacterial LPS are largely unknown. In these studies, the role of the mitogen-activated protein kinase (MAPK) pathways in the regulation of MMP-13 induced by lipopolysaccharide was investigated. Lipopolysaccharide from Escherichia coli and Actinobacillus actinomycetemcomitans significantly ( P < 0.05) increased MMP-13 steady-state mRNA (average of 27% and 46% increase, respectively) in murine periodontal ligament fibroblasts. MMP-13 mRNA induction was significantly reduced by inhibition of p38 MAP kinase. Immunoblot analysis indicated that p38 signaling was required for LPS-induced MMP-13 expression. Lipopolysaccharide induced proximal promoter reporter (—660/+32 mMMP-13) gene activity required p38 signaling. Collectively, these results indicate that lipopolysaccharide-induced murine MMP-13 is regulated by p38 signaling through a transcriptional mechanism.