Binding of pentraxins to different nuclear structures: C-reactive protein binds to small nuclear ribonucleoprotein particles, serum amyloid P component binds to chromatin and nucleoli.

Binding of the human pentraxin plasma proteins, C-reactive protein (CRP) and serum amyloid P component (SAP), to the nuclei of human cells was studied using whole acute phase serum as the source of the proteins and confocal immunofluorescence microscopy. CRP and SAP clearly bound to distinct, different structures. Double staining with MoAbs to the Sm D and Sm B/B' components of small nuclear ribonucleoproteins confirmed that CRP bound exclusively to these particles. As expected, SAP bound to chromatin and, in addition, binding to the nucleolus was observed for the first time. These interactions demonstrated under relatively physiological conditions, with native pentraxins unseparated from serum and with nuclear constituents in situ, are likely to be of functional importance in vivo.     

Studies of the structure and binding properties of hamster female protein.

We report here the characterization of hamster female protein (FP), a member of the pentraxin family of plasma proteins, as a molecule composed of glycosylated subunits of 25,655 MW containing a single intrachain disulphide bridge. In the presence of EDTA the subunits are non-covalently associated as pentamers of mass approximately 128,000 MW, and in the presence of calcium they aggregate further, probably to form decamers. This pentamer-decamer transition at physiological ionic strength has not been described in other pentraxins. As previously reported, FP shares the capacity of C-reactive protein (CRP) in other species to bind phosphocholine and we show here that it also resembles human CRP in binding only weakly to agarose, to human AA amyloid fibrils in vitro, and to mouse AA amyloid deposits in vivo. It thus differs markedly from human and mouse serum amyloid P component (SAP) but it is nevertheless deposited in hamster AA amyloid in vivo and clearly is the hamster counterpart of SAP in other species. These results illustrate the subtle diversity among members of the otherwise conserved pentraxin family of vertebrate plasma proteins.     

巨噬细胞LPS相关模式识别受体的研究进展

脓毒症(sepsis)是由各种致病微生物或其毒素引起的全身炎症反应综合征(systemic inflammatory response syndrome, SIRS),是严重感染、重度创伤、大手术后和休克常见的并发症,进一步发展可导致脓毒性休克、急性呼吸窘迫综合征(acute respiratory distress syndrome, ARDS)和多器官功能障碍综合征(multi-organ dysfunction syndrome,MODS)等致命性并发症.     

Isolation and characterisation of pentraxin-like serum proteins from the common carp Cyprinus carpio

There is increasing economic and ecological interest in the development of assays for the early detection of infection, disease activity and environmental stress in marine and freshwater animals. In humans the serum pentraxin C-reactive protein (CRP) is universally used as a clinical indicator of inflammation and underlying infection. As a first step towards assessing the potential of an immunoassay for CRP in fish, we have isolated and characterised common carp Cyprinus carpio CRP and a highly specific and sensitive anti-carp CRP polyclonal antibody has been raised. The results show levels of CRP in healthy fish similar to those found in healthy humans. A protein of unknown function, which displays the characteristic calcium-dependent phosphate monoester binding exhibited by CRP and some similarity to the known fish pentraxin sequences, has also been identified.     

C-reactive protein and the biology of disease

The C-reactive protein (CRP) is a plasma protein of hepatic origin, belonging to pentraxin family and forms a major component of any inflammatory reaction. A key component of the innate immunity pathway, the concentration of CRP may rapidly increase to levels more than 1,000-folds above normal values as a consequence to tissue injury or infection. Although functioning as a classical mediator of innate immunity, it functions via interaction of components of both humoral and cellular effector systems of inflammation. Initially considered as an acute-phase marker in tissue injury, infection and inflammation, it now has a distinct status of a disease marker in cardiovascular diseases and is well known of its clinical and pathological significance. The present torrent of studies in a large number of diseases and associated conditions has highly elucidated the role of CRP as a therapeutic and research reagent. In this review, we focus our attention to role of CRP in health and disease. The future prospect of this review lies in the applicability of CRP as a molecule in understanding and monitoring of the biology of disease.     

Hamster female protein binding to chromatin, histones and DNA.

Hamster female protein (FP) is a member of the family of proteins known as pentraxins which share amino acid sequence homology, cyclic pentameric structure and calcium-dependent binding to ligands. Other members of this family include C-reactive protein (CRP) and serum amyloid P component (SAP), and most species synthesize both CRP and SAP. FP is unusual in that it is apparently the only pentraxin produced in hamsters, it is under hormonal control and it shares binding characteristics with both CRP and SAP. CRP has been defined and isolated by its calcium-dependent binding to pneumococcal C-polysaccharide via phosphocholine (PC) residues. SAP has been isolated by calcium-dependent binding to agarose. FP binds to both PC and agarose. Recently, both SAP and CRP have been found to bind to chromatin in a calcium-dependent manner and involvement of these proteins in the clearance of nuclear material has been proposed. In this paper we test whether FP shares the ability to bind to chromatin and histones, and compare its relative avidities for these ligands. Similar to CRP, FP bound to histones H1 and H2A, and chromatin. FP shared with SAP the ability to bind to DNA. However, FP binding was inhibited by PC for all ligands, whereas SAP binding was not. FP and SAP also failed to compete with each other for binding to DNA. By cross-inhibition FP bound much less well to PC than CRP, but was a very effective inhibitor of CRP binding to H2A. These studies demonstrate that chromatin and histone binding are conserved among these pentraxins. The role of the proposed PC binding site in these binding reactions is discussed.     

Changes in serum concentration of a serum amyloid P-like pentraxin in Atlantic salmon, Salmo salar L., during infection and inflammation

A serum amyloid P-like pentraxin has been isolated from Atlantic salmon. Salmo salar L., based on its calcium dependent binding to agarose. The subunits of approximately 37 kDa were all glycosylated and when covalently linked together formed a pentamer with disulphide bonds between all subunits. A specific rabbit antiserum raised against the pentameric form was used to follow changes in serum levels of the salmon pentraxin during infection with Aeromonas salar and inflammation induced by Escherichia coli LPS or killed A. salmonicida. The salmon pentraxin level in normal serum was in the range approximately 50-300 microg/ml. Unlike pentraxins from other species, the salmon pentraxin showed only moderate but significant increases or decreases in response to E. coli LPS or A. salmonicida infection, respectively. Although pentraxins and pentraxin-like proteins are evolutionary conserved, not all pentraxins are acute phase responders suggesting that their most ancestral function(s) are not related to acute phase induction.     

脂多糖模式识别受体的研究进展

G-细菌的脂多糖（LPS）是重要的病原体相关模式分子。PAMPsd均可被动物作为低外来分子进行识别,LPS能激发机体细胞因子IL-1、TNF-α等活性分子的合成,对感染具有十分重要的作用。LPS是通过什么受体怎样将信号传玫免疫细胞并启动免疫反应的,人们一直都不十分清楚,近年来,一种名为Toll蛋白的发现,使人们对机体识别LPS机制的认识向前跨进了一大步,本文试对该模式识别受体的研究进展做一综述。     

Maternal Plasma and Amniotic Fluid LBP,Pentraxin 3, Resistin,and IGFBP-3: Biomarkers of Microbial Invasion of Amniotic Cavity and/or Intra-amniotic Inflammation in Women with Preterm Premature Rupture of Membranes

BackgroundWe sought to determine whether lipopolysaccharide binding protein (LBP), pentraxin 3, resistin, and insulin-like growth factor binding protein (IGFBP)-3 in plasma and amniotic fluid (AF) can predict microbial invasion of the amniotic cavity (MIAC), intra-amniotic inflammation (IAI), and microbial-associated IAI in women with preterm premature rupture of membranes (PPROM).MethodsThis was a retrospective cohort study involving 168 singleton pregnant women with PPROM. AF obtained via amniocentesis was cultured and assayed for interleukin (IL)-6 to define IAI and for IL-8 to compare with AF biomarkers. Plasma samples were collected at the time of amniocentesis, and C-reactive protein (CRP) levels in serum were compared with plasma biomarkers. The stored plasma and AF samples were assayed for LBP, pentraxin 3 (PTX3), resistin, and IGFBP-3 by ELISA.ResultsMultivariate logistic regression analysis revealed that: 1) elevated plasma and AF levels of LBP were independently associated with increased risks of MIAC, IAI, and microbial-associated IAI; 2) elevated AF, but not plasma, PTX3, and resistin levels were independently associated with increased risks of MIAC, IAI, and microbial-associated IAI; 3) decreased IGFBP-3 levels in the plasma were independently associated with only IAI, whereas those in the AF were associated with only microbial-associated IAI. Among the tested biomarkers, AF PTX3 and resistin had the highest predictive performance for MIAC, IAI, and microbial-associated IAI (area under the curves [AUC] = 0.85–0.95), which is similar to the performance of AF IL-8. The AUCs of the plasma LBP and IGFBP-3 were similar to that of serum CRP with respect to IAI.ConclusionMaternal plasma LBP and IGFBP-3 are potential biomarkers for the non-invasive identification of IAI in women with PPROM, with a similar accuracy to the serum CRP level. AF LBP, PTX3, resistin, and IGFBP-3 may be involved in the intra-amniotic inflammatory responses in PPROM complicated by MIAC.     

Elevated plasma levels of the long pentraxin, pentraxin 3, in severe dengue virus infections

C-reactive protein is one of the most widely used indicators of the response of acute-phase proteins. The measurement of C-reactive protein in dengue, however, is clinically not useful, because of marginally elevated levels and absent association with disease severity. The prototypic long pentraxin, pentraxin 3, is an acute phase protein that is structurally related but distinct from C-reactive protein which has proven to correlate with the severity of bacterial infection in critically ill patients. The potential involvement of pentraxin 3 in dengue and its aptitude to predict more severe disease or poor clinical outcome has not been studied previously. We therefore measured pentraxin 3 plasma levels in 44 dengue virus infected patients. Pentraxin 3 levels were strikingly higher when compared to C-reactive protein levels, with highest pentraxin 3 values observed in the first 7 days after the onset of symptoms. Median pentraxin 3 levels at admission and peak levels during follow up were higher in patients suffering from dengue shock syndrome (at admission: 119.3 ng/ml [interquartile range 61.8--188.7], peak values during follow up: 147.9 ng/ml [interquartile range 85.7--204.3]) compared to levels found in patients with dengue fever and dengue hemorrhagic fever (at admission: 59.0 ng/ml [interquartile range 28.6--100.3], P=0.040; peak values during follow up: 80.8 ng/ml [interquartile range 36.1--168.1], P=0.020). Our results indicate that pentraxin 3 seems to be a marker of infection better than C-reactive protein in dengue. The role of pentraxin 3 in the pathogenesis of dengue and its potential as an early prognostic indicator of disease severity needs further assessment.     

PTX3 as a paradigm for the interaction of pentraxins with the Complement system

Pentraxins are highly conserved components of the humoral arm of innate immunity. They include the short pentraxins C reactive protein (CRP) and serum amyloid P component (SAP), and the long pentraxin PTX3. These are soluble pattern-recognition molecules that are present in the blood and body fluids, and share the ability to recognize pathogens and promote their disposal. CRP and SAP are produced systemically in the liver while PTX3 is produced locally in a number of tissues, macrophages and neutrophils being major sources of this long pentraxin. Pentraxins interact with components of the classical and lectin pathways of Complement as well as with Complement regulators. In particular, PTX3 recognizes C1q, factor H, MBL and ficolins, where these interactions amplify the repertoire of microbial recognition and effector functions of the Complement system. The complex interaction of pentraxins with the Complement system at different levels has broad implications for host defence and regulation of inflammation.     

Isolation and characterization of pharmaceutical grade human pentraxins, serum amyloid P component and C-reactive protein, for clinical use

The human pentraxin proteins, serum amyloid P component (SAP) and C-reactive protein (CRP) are important in routine clinical diagnosis, SAP for systemic amyloidosis and CRP for monitoring the non-specific acute phase response. They are also targets for novel therapies currently in development but their roles in health and disease are controversial. Thus, both for clinical use and to rigorously elucidate their functions, structurally and functionally intact, pharmaceutical grade preparations of the natural, authentic proteins are required. We report here the production from normal human donor plasma and the characterization of the first such preparations. Importantly, we demonstrate that, contrary to reports using recombinant proteins and less well characterized preparations, neither CRP nor SAP stimulate the release by human peripheral blood mononuclear cells in vitro of any TNFα, IL-6 or IL-8, nor does SAP cause release of IL-1β or IL-10. Furthermore neither of our preparations was pro-inflammatory in mice in vivo.     

Isolation and partial characterization of a pentraxin-like protein with complement-fixing activity from snapper (Pagrus auratus,Sparidae) serum

Pentraxin-like molecules have been isolated from a number of fish species. However, little is known about the function of these proteins in the teleosts. In this study we report the isolation and characterization of a pentraxin-like molecule from the serum of snapper (Pagrus auratus) that has the ability to activate complement. This pentraxin-like protein was isolated from serum by calcium-dependent binding to agarose. SDS-PAGE analysis demonstrated an oligomeric protein of approximately 200k Da consisting of non-covalently bound subunits of 26 and 23 kDa. Protein sequencing revealed significant (50%) sequence identity with pentraxins from both Atlantic salmon (S. salar) and rainbow trout (O. mykiss). However, polyclonal antibodies raised against snapper pentraxin did not recognise salmon or trout pentraxin in Western blot analysis. Following LPS injection, snapper pentraxin levels increased 2-fold before gradually returning to basal levels. Most significantly, the isolated pentraxin initiated complement-mediated lysis of ligand-coated sheep erythrocytes in a dose-dependent fashion. In view of the similarity between the known fish pentraxins, and their similarity to mammalian serum amyloid P-components we conclude that the isolated protein may be a snapper pentraxin homologue.     

Production and characterisation of mouse monoclonal antibodies reactive with the lipopolysaccharide core of Pseudomonas aeruginosa.

Monoclonal antibodies (MAbs) to the core antigen region of lipopolysaccharide (LPS) of Pseudomonas aeruginosa were produced from mice immunised with whole cells of heat-killed rough mutants of Pseudomonas aeruginosa expressing partial or complete core LPS. MAbs were screened in an enzyme-linked immunosorbent assay (ELISA) against three different antigen cocktails: S-form LPS from three P. aeruginosa strains, R-form LPS from six P. aeruginosa strains and, as a negative control, R-form LPS from Salmonella typhimurium and Escherichia coli. Selected MAbs were subsequently screened against a range of extracted LPS and whole cells from both reference strains and clinical isolates of P. aeruginosa. The antibodies were also screened in ELISA against whole-cell antigens from other Pseudomonas spp. as well as strains of Haemophilus influenzae, Neisseria subflava and Staphylococcus aureus. Five MAbs reacting with the core component of P. aeruginosa LPS were finally selected. Two of these, MAbs 360.7 and 304.1.4, were particularly reactive in immunoblots against unsubstituted core LPS, including that from O-antigenic serotypes of P. aeruginosa. The MAbs also reacted with some of the other Pseudomonas spp., but not with P. cepacia or Xanthomonas (Pseudomonas) maltophilia. Cross-reactivity with whole cells from other bacterial species was minimal or not observed. Reactivity of MAbs with some Staph. aureus strains was observed, and binding to the protein A component was implicated. The reactivity of the MAbs was investigated further by flow cytometry and immunogold electronmicroscopy. The suitability of the MAbs for an immunological assay for detection of P. aeruginosa in respiratory secretions from CF patients is discussed.     

Lipopolysaccharide binding protein-deficient mice have a normal defense against pulmonary mycobacterial infection

Lipopolysaccharide (LPS) binding protein (LBP) facilitates the transfer of LPS of Gram-negative bacteria to the pattern recognition receptor CD14, resulting in activation of immunocompetent cells. LBP can also facilitate the binding of lipoarabinomannan, a major cell wall component of mycobacteria, to immune cells. To determine the role of LBP in the immune response to pulmonary Mycobacterium tuberculosis infection, LBP gene-deficient (-/-) and normal wild-type (WT) mice were intranasally infected with M. tuberculosis. LBP-/- mice displayed a similar survival and mycobacterial outgrowth in lungs and liver, although they demonstrated a reduced lymphocyte recruitment and activation during the early stages of infection. The clearance of pulmonary infection with the non-pathogenic M. smegmatis was also unaltered in LBP-/- mice. These data suggest that LBP does not contribute to an effective host response in M. tuberculosis infection.     

Inflammatory Markers in the Second Trimester Prior to Clinical Onset of Preeclampsia, Intrauterine Growth Restriction, and Spontaneous Preterm Birth

Low-grade inflammation has been associated with pregnancy complications including preeclampsia (PE), intrauterine growth restriction (IUGR), and spontaneous preterm birth (SPB). In an unmatched, nested case–control study, we assessed the possible predictive association of maternal C-reactive protein (CRP), interferon-γ-inducible protein 10 (IP-10), and soluble urokinase plasminogen activator receptor (suPAR) in second trimester plasma samples in relation to later development of PE (n?=?29), IUGR (n?=?53), and SPB (n?=?9). Inflammatory marker levels in these groups were compared to normotensive healthy pregnant controls (n?=?127). We found no statistically significant difference in CRP, IP-10, or suPAR in second trimester plasma samples from pregnant women with later PE, IUGR, and SPB when compared to normotensive healthy controls. Second trimester plasma samples of CRP, IP-10, and suPAR cannot be used as a prognostic marker for PE, IUGR, and SPB.     

Reactivity of anti-human C-reactive protein (CRP) and serum amyloid P component (SAP) monoclonal antibodies with limulin and pentraxins of other species.

Limulus polyphemus C-reactive protein (CRP) (limulin) has approximately 30% amino acid sequence homology and shares at least one idiotypic determinant associated with ligand-binding activity with human CRP (hCRP); limulin also shares amino acid sequence homology and lectin activity with human serum amyloid P component (hSAP). In the present study panels of 14 anti-hCRP monoclonal antibodies (mAb) directed to distinct hCRP epitopes and 11 anti-hSAP mAb directed to distinct epitopes of hSAP were tested for reactivity with limulin and pentraxins of other species including rabbit CRP (raCRP), rat CRP and hamster female protein (FP) by ELISA and Western blot analyses. None of the anti-human pentraxin mAb showed strong cross-reactivity with limulin; only five mAb reacted with limulin at all, and cross-reactivities of these mAb with the other pentraxins, when present, also were weak. Cross-reactivity of limulin with hCRP and hSAP was similar, and in light of comparable amino acid sequence homology, suggests this molecule can be considered the limulus SAP as well as the limulus CRP. Several anti-hCRP mAb cross-reacted strongly with rabbit CRP and rat CRP; a few anti-hSAP cross-reacted strongly with FP; and weak cross-reactions were observed between hCRP and hSAP, but cross-reactivities between the pentraxins generally were limited and weak. A rabbit polyclonal antibody raised to highly conserved limulin peptide 141-156 and strongly reactive with limulin reacted weakly with hCRP and raCRP but failed to react with rat CRP, hSAP or FP. These studies emphasize a limited but distinct antigenic similarity between limulin, hCRP and other pentraxins, and identify mAb reactive with potential regions of shared structure and/or function between pentraxins of different species.     

The role of long pentraxin 3, a new inflammatory mediator in inflammatory responses]

Pentraxin 3 (PTX3) is suggested to play important roles in the innate resistance against pathogens, regulation of inflammatory reactions, and clearance of apoptotic cells. PTX3 is the first long pentraxin identified. Long pentraxin shares a C-terminal pentraxin domain with the classical short pentraxin (C-reactive protein, serum amyloid P), but holds an unrelated N-terminal domain that is unique to the long pentraxin. While the short pentraxin is produced only in the liver, PTX3 is made by diverse types of cells, prominently endothelial cells and macrophage, in response to inflammatory signals. Unlike the short pentraxin, the expression of PTX3 in multiple types of tissue cells implies a mechanism for local amplification of innate resistance at the site of infection and inflammation. PTX3 plasma levels are very low in normal subjects but are rapidly increased by inflammatory conditions resulting from a wide range of diseased states, from infection to autoimmune and degenerative disorders. Critically ill patients show elevated circulating levels of PTX3 which are determined by the severity of the disease. Clinical evidence has demonstrated that the elevated PTX3 levels might be a useful early and sensitive marker for severely ill patients. Further studies will definitely be needed to deepen our understanding of PTX3.     

Pentraxin 3 Increase is Much Less Pronounced Than C-Reactive Protein Increase After Surgical Procedures

Pentraxin 3 is an acute phase marker that belongs to the same protein family as C-reactive protein (CRP). The aim of this study was to compare the acute phase reactions of pentraxin 3 and CRP in humans. High sensitivity CRP and pentraxin 3 were analyzed in blood samples from orthopedic surgery (n = 29) and coronary bypass patients (n = 21). The samples were collected prior to surgery and 4 and 30 days after surgery, respectively. Both CRP and pentraxin 3 were significantly increased at day 4. Median pentraxin 3 values increased from 4,021 to 7,459 pg/mL in the orthopedic group and from 4,637 pg/mL to 10,419 pg/mL in the coronary bypass group while CRP increased from 6.3 mg/L to 151.6 mg/L and from 5.7 mg/L to 176.3 mg/L in the same groups. Pentraxin 3 shows a much smaller increment in humans in comparison with CRP.