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C-reactive protein (CRP) is a member of the pentraxin family of proteins. These proteins are highly conserved over the course of evolution being present as far back as 250 million years ago. Mammalian pentraxins are characterized by the presence of five identical non-covalently linked subunits. Each subunit has a structurally conserved site for calcium-dependent ligand binding. The biological activities of the pentraxins established over many years include the ability to mediate opsonization for phagocytosis and complement activation. Pentraxins have an important role in protection from infection from pathogenic bacteria, and regulation of the inflammatory response. It was recognized early on that some of these functions are mediated by activation of the classical complement pathway through C1q. However, experimental evidence suggested that cellular receptors for pentraxins also play a role in phagocytosis. More recent experimental evidence indicates a direct link between pentraxins and Fc receptors. The Fc receptors were first identified as the major receptors for immunoglobulins. The avidity of the interaction between IgG complexes and Fc receptors is greatly enhanced when multivalent ligands interact with the IgG binding sites and activation of signaling pathways requires Fc receptor crosslinking. Human pentraxins bind and activate human and mouse IgG receptors, FcγRI and FcγRII, and the human IgA receptor, FcαRI. The affinities of the interactions between Fc receptors and pentraxins in solution and on cell surfaces are similar to antibody binding to low affinity Fc receptors. Crystallographic and mutagenesis studies have defined the structural features of these interactions and determined the stoichiometry of binding as one-to-one. Pentraxin aggregation or binding to multivalent ligands increases the avidity of binding and results in activation of these receptors for phagocytosis and cytokine synthesis. This review will discuss the structural and functional characteristics of pentraxin Fc receptor interactions and their implications for host defense and inflammation.

Pentraxins are a family of secreted pattern recognition proteins containing a homologous pentraxin (PTX) domain of approximately 200 amino acids (Fig. 1). Members of the family, such as C-reactive protein (CRP) (also called PTX1) and serum amyloid P component (SAP) (also called PTX2) containing only a PTX domain, are termed short chain pentraxins, while others, such as pentraxin member 3 (PTX3), and neuronal pentraxins 1 and 2, have an additional N-terminal domain preceding the PTX domain and are referred as long chain pentraxins. The structure and function of short chain pentraxins have been studied more extensively compared to the members of long chain pentraxins (1-5). Both CRP and SAP are part of acute phase proteins expressed by hepatocytes in response to inflammatory cytokine stimulation during infections (6-8). CRP recognizes phosphorylcholine moieties associated with microbial polysaccharides or necrotic cells in a calcium-dependent manner (9, 10). In addition, CRP also recognizes some nuclear antigens, such as small nuclear ribonucleoprotein and chromatin subunits (11-15). SAP is known to recognize phosphorylethanolamine (PE) and other microbial associated molecules as well as nuclear antigens in a calcium-dependent manner (11, 16-18). Both CRP and SAP can opsonize microbial pathogens or apoptotic cells and effectively clear them through complement activation (19-22). Fig. 1 Domain structure of pentraxins. The structure of a pentraxin domain displays a conserved β-sandwich fold with two opposing β-sheets each consisting of 7 β-strands (2, 23, 24) (Fig. 2A). A three turn α-helix from residue 168-176 of CRP, termed ridge helix, is situated on top of the β-sandwich. Both CRP and SAP subunits display a similar donut-shaped pentameric structure, with all five ridge helices on one face and microbial ligand binding sites on the opposite face. Calcium ions form part of the phosphorylcholine-binding site (23). The face with ridge helices is shown to contain C1q-binding sites by mutational analysis (25). Fig. 2 Structural recognition between pentraxins and Fc receptors Immunoglobulin Fc receptors (FcRs) are broadly expressed on hematopoietic cells and are important for antibody-mediated humoral and cellular immunity. Binding of immune complexes to Fc receptors activates effector cells leading to phagocytosis, endocytosis of IgG-opsonized particles, the release of inflammatory mediators, and antibody-dependent cellular cytotoxicity (ADCC) (26-28). The most well studied Fc receptors are FcγR and neonatal FcR (FcRn) for immunoglobulin G (IgG), FceR for IgE, and FcαR for IgA. Fc receptors belong to the immunoglobulin superfamily, except for the FcRn and FceRII, which are structurally related to class I major histocompatibility antigens and C-type lectins, respectively. FcγRI, FceRI, and FcαRI display higher binding affinities to their cognate immunoglobulins than the low affinity FcγRII and FcγRIII. In addition to the affinity variations among the receptors, Fcγ receptors also display distinct IgG subtype specificities. For example, FcγRIII binds IgG1 and IgG3 better than IgG2 and IgG4 (29). All Fcγ receptors are type I transmembrane or GPI-anchored glycoproteins consisting of two or three C2-type immunoglobulin-like domains. All Fcγ receptors show a high degree of sequence identity in their extracellular portion (50-96%) but differ significantly in their cytoplasmic domains (28). Activating Fcγ receptors either contain or associate with a common signaling γ-chain that contains the immunoreceptor tyrosine-based activation motifs (ITAMs), while the inhibitory FcγRIIB contains the immunoreceptor tyrosine-based inhibitory motif (ITIM) in its cytoplasmic tail. Several structures of the extracellular portion of Fc receptors have been published (30-34). They can be divided into two structural groups depending on their relative orientation between D1 and D2 domains (Fig. 2B). Group I receptors include all Fcγ receptors and FceRI, and group II includes FcαRI. The D1-D2 domain juxtaposition of group I receptors is opposite to that of group II, which displays natural killer cell immunoglobulin-like receptor (KIR) domain juxtaposition. The antibody-binding affinities of Fc receptors vary substantially between 10-5 and 10-9 M (26, 27, 29). Among them, FceRI and FcγRI display high affinity IgE and IgG binding, respectively, and are presumed preoccupied with their antibodies in circulation. Others, including FcγRII, FcγRIII, and FcαRI, display intermediate and low binding affinities toward their antibodies and thus can only recognize antigen-bound immune complexes. Despite differences in antibody affinities, both the high affinity FceRI and the lower affinity FcγRs have been shown to recognize their antibody Fc domains in similar structure modes in which one Fc receptor interacts with both heavy chains of Fc in the lower hinge region, between CH1 and CH2 domains in IgG or between Ce2 and Ce3 in IgE, near the conserved Fc glycosylation site (35-39). Aside from their well-established functions in antibody-mediated immune activations, Fc receptors were not known to recognize soluble innate pentraxins until recently.

C-reactive protein (CRP) is an important biomarker for inflammatory diseases. However, its role in inflammation beyond complement-mediated pathogen clearance remains poorly defined. We identified the major IgA receptor, FcαRI, as a ligand for pentraxins. CRP recognized FcαRI both in solution and on cells, and the pentraxin binding site on the receptor appears distinct from that recognized by IgA. Further competitive binding and mutational analysis showed that FcαRI bound to the effector face of CRP in a region overlapping with complement C1q and Fcγ receptor (FcγR) binding sites. CRP cross-linking of FcαRI resulted in extracellular signal-regulated kinase (ERK) phosphorylation, cytokine production, and degranulation in FcαRI-transfected RBL cells. In neutrophils, CRP induced FcαRI surface expression, phagocytosis, and TNF-α secretion. The ability of CRP to activate FcαRI defines a function for pentraxins in inflammatory responses involving neutrophils and macrophages. It also highlights the innate aspect of otherwise humoral immunity-associated antibody receptors.

The classical pentraxins, serum amyloid P component (SAP) and C-reactive protein (CRP), are major acute phase reactants in mouse and man. As shown in this paper, pentraxins recognize various FcγRs, and SAP opsonization activates FcγR-mediated phagocytosis and cytokine secretion. The receptor binding sites for SAP and IgG overlap resulting in competition of IgG binding to FcγR as well as inhibition of immune complex-mediated phagocytosis by soluble pentraxins. The classical pentraxins, serum amyloid P component (SAP) and C-reactive protein (CRP), are major acute phase reactants in mouse and man. It is shown that pentraxins recognize various FcγRs and SAP opsonization activates FcγR-mediated phagocytosis and cytokine secretion. The receptor binding sites for SAP and IgG overlap, resulting in competition of IgG binding to FcγR as well as inhibition of immune complex-mediated phagocytosis by soluble pentraxins. Pentraxins are a family of ancient innate immune mediators conserved throughout evolution. The classical pentraxins include serum amyloid P component (SAP) and C-reactive protein, which are two of the acute-phase proteins synthesized in response to infection1,2. Both recognize microbial pathogens and activate the classical complement pathway through C1q (refs 3 and 4). More recently, members of the pentraxin family were found to interact with cell-surface Fcγ receptors (FcγR) and activate leukocyte-mediated phagocytosis5,6,7,8. Here we describe the structural mechanism for pentraxin’s binding to FcγR and its functional activation of FcγR-mediated phagocytosis and cytokine secretion. The complex structure between human SAP and FcγRIIa reveals a diagonally bound receptor on each SAP pentamer with both D1 and D2 domains of the receptor contacting the ridge helices from two SAP subunits. The 1:1 stoichiometry between SAP and FcγRIIa infers the requirement for multivalent pathogen binding for receptor aggregation. Mutational and binding studies show that pentraxins are diverse in their binding specificity for FcγR isoforms but conserved in their recognition structure. The shared binding site for SAP and IgG results in competition for FcγR binding and the inhibition of immune-complex-mediated phagocytosis by soluble pentraxins. These results establish antibody-like functions for pentraxins in the FcγR pathway, suggest an evolutionary overlap between the innate and adaptive immune systems, and have new therapeutic implications for autoimmune diseases.

C-reactive protein (CRP) is an acute phase reactant with roles in innate host defense, clearance of damaged cells, and regulation of the inflammatory response. These activities of CRP depend on ligand recognition, complement activation, and binding to FcγR. CRP binds to phosphocholine in the Streptococcus pneumoniae cell wall and provides innate defense against pneumococcal infection. These studies examine the effect of this early innate defense molecule on the development of Abs and protective immunity to S. pneumoniae. Dendritic cells (DC) initiate and direct the adaptive immune response by integrating innate stimuli with cytokine synthesis and Ag presentation. We hypothesized that CRP would direct uptake of S. pneumoniae to FcγR on DC and enhance Ag presentation. CRP opsonization of the R36a strain of S. pneumoniae increased the uptake of bacteria by DC. DC pulsed with untreated or CRP-opsonized R36a were transferred into recipient mice, and Ab responses were measured. In mice challenged with free R36a, CRP opsonization resulted in higher secondary and memory IgG responses to both phosphocholine and pneumococcal surface protein A. Furthermore, mice immunized with DC that had been pulsed with CRP-opsonized R36a showed increased resistance to intranasal infection with virulent S. pneumoniae. The effects of CRP on Ag uptake, Ab responses, and protection from infection all required FcR γ-chain expression on DC. The results indicate that innate recognition by CRP enhances effective uptake and presentation of bacterial Ags through FcγR on DC and stimulates protective adaptive immunity.

C-reactive protein (CRP) is the prototypic marker of inflammation and a strong predictor of cardiovascular events in humans. There are questions regarding the validity of the biological effects reported for CRP, in spite of adherence to rigorous control measures minimizing endotoxin [lipopolysaccharide (LPS)] contamination in these in vitro studies. In this study, we addressed the key question of endotoxin contamination in CRP preparations using Toll-like receptor 4 (TLR4) knockdown endothelial cells. Human aortic endothelial cells (HAECs) transfected with prevalidated TLR4 small interfering RNA (siRNA) and scrambled siRNA controls were challenged with pleural fluid-derived CRP or LPS for 12-16 h. Secreted interleukin-6 (IL-6), IL-1beta, IL-8, and plasminogen activator inhibitor-1 (PAI-1) levels and endothelial Nitric oxide synthase (eNOS) activity were determined. TLR4 knockdown in HAECs significantly decreased LPS-induced IL-1beta, IL-6, and IL-8, whereas the stimulatory effects of CRP were similar in both scrambled control and TLR4 knockdown cells. Furthermore, CRP significantly stimulated PAI-1 levels in both control and TLR4-transfected cells and inhibited eNOS activity, whereas LPS effects were negated in TLR4-transfected cells. The data presented cogently demonstrate and further confirm that the biological effects of CRP on HAECs are independent of LPS and thus are attributable to native protein per se. This is the first study to positively authenticate the significance of earlier in vitro reports on CRP biological effects.

Streptococcus pneumoniae is the most common organism responsible for community acquired pneumonia and meningitis. In pneumococcal pneumonia, a strong local inflammatory cytokine response reduces the frequency of bacteremia and increases survival. The initiation of this cytokine response by innate recognition of bacterial cell wall components through TLR has been described, but the role of soluble innate mediators has received limited attention. C-reactive protein (CRP) is an acute phase protein that binds phosphocholine residues on S. pneumoniae cell walls. CRP interacts with phagocytic cells through FcγRI and FcγRII and activates the classical complement pathway. CRP is protective in mouse pneumococcal bacteremia by increasing complement-dependent clearance and killing of bacteria. We studied the cytokine response of PBMC stimulated with CRP-opsonized S. pneumoniae to determine the effect of CRP interaction with FcγR. CRP dramatically increased the production of TNF-α and IL-1β in response to S. pneumoniae. These increases were blocked by phosphocholine, which inhibits CRP binding to S. pneumoniae, by inhibitors of FcγR signaling, and by mAb to FcγRI and FcγRII. A mutated rCRP with decreased FcγR binding had a decreased ability to stimulate TNF-α release, compared with wild-type CRP. Individuals who were homozygous for the R-131 allele of FcγRIIA, which has a higher affinity for CRP, showed higher responses to CRP-opsonized bacteria than did individuals homozygous for the H-131 allele, further implicating this receptor. The results indicate that CRP recognition of S. pneumoniae and binding to FcγR may enhance the early protective cytokine response to infection.

Human C-reactive protein (CRP) is a classical, acute phase serum protein synthesized by the liver in response to infection, inflammation, or trauma. CRP binds to microbial antigens and damaged cells, opsonizes particles for phagocytosis and regulates the inflammatory response by the induction of cytokine synthesis. These activities of CRP depend on its ability to activate complement and to bind to Fcgamma receptors (FcgammaR). The goal of this study was to elucidate amino acid residues important for the interaction of CRP with human FcgammaRI (CD64) and FcgammaRIIa (CD32). Several mutations of the CRP structure were studied based on the published crystal structure of CRP. Mutant and wild-type recombinant CRP molecules were expressed in the baculovirus system and their interactions with FcgammaR and C1q were determined. A previous study by our laboratory identified an amino acid position, Leu(176), critical for CRP binding to FcgammaRI and work by others (Agrawal, A., Shrive, A. K., Greenhough, T. J., and Volanakis, J. E. (2001) J. Immunol. 166, 3998-4004) determined several residues important for C1q binding. The amino acid residues important to CRP binding to FcgammaRIIa were previously unknown. This study newly identifies residues Thr(173) and Asn(186) as important for the binding of CRP to FcgammaRIIa and FcgammaRI. Lys(114), like Leu(176), was implicated in binding to FcgammaRI, but not FcgammaRIIa. Single mutations at amino acid positions Lys(114), Asp(169), Thr(173), Tyr(175), and Leu(176) affected C1q binding to CRP. These results further identify amino acids involved in the binding sites on CRP for FcgammaRI, FcgammaRIIa, and C1q and indicate that these sites are overlapping.

C-reactive protein (CRP) is an acute-phase serum protein and a member of the pentraxin protein family. Its host defense functions predate the adaptive immune system by millions of years. Our current understanding of CRP interactions with complement and with Fcgamma receptors (FcgammaR) have led to an increased appreciation of the regulatory role of CRP in inflammation and autoimmunity. This review outlines the role of CRP in infection, inflammation, and autoimmune disease. We provide a description of recent studies, which suggest that CRP acts through FcgammaR to reduce inflammation and protect from certain autoimmune diseases. A general description of the proposed function of CRP is provided as a framework for future investigation.

C-reactive protein (CRP) is a component of the acute phase response to infection, inflammation, and trauma. A major activity of acute phase proteins is to limit the inflammatory response. It has been demonstrated that CRP protects mice from lethal doses of LPS. In the mouse, CRP binds to the regulatory receptor, FcγRIIb, and to the γ-chain-associated receptor, FcγRI. The goal ofthis study was to determine whether FcγRs are necessary for the protective effect of CRP. The ability of CRP to protect mice from a lethal dose of LPS was confirmed using injections of 500 and 250 μg of CRP at 0 and 12 h. CRP treatment of FcγRIIb-deficient mice increased mortality after LPS challenge and increased serum levels of TNF and IL-12 in response to LPS. CRP did not protect FcR γ-chain-deficient mice from LPS-induced mortality. Treatment of normal mice, but not γ-chain-deficient mice, with CRP increased IL-10 levels following LPS injection. In vitro, in the presence of LPS, CRP enhanced IL-10 synthesis and inhibited IL-12 synthesis by bone marrow macrophages from normal, but not γ-chain-deficient mice. The protective effect of CRP appears to be mediated by binding to FcγRI and FcγRII resulting in enhanced secretion of the anti-inflammatory cytokine IL-10 and the down-regulation of IL-12. These results suggest that CRP can alter the cytokine profile of mouse macrophages by acting through FcγR leading to a down-regulation of the inflammatory response.

Serum amyloid P component (SAP) is a member of the pentraxin family of proteins. These proteins are characterized by cyclic pentameric structure, calcium-dependent ligand binding, and frequent regulation as acute-phase serum proteins. SAP is the serum precursor of the P component of amyloid. It binds to a broad group of molecules, including autoantigens, through a pattern recognition binding site. The related pentraxin, C-reactive protein (CRP), is a strong acute-phase reactant in man and an opsonin. We previously determined that the binding of CRP to leukocytes occurs through Fc receptors for IgG (FcγR). We now report that SAP also binds to FcγR and opsonizes particles for phagocytosis by human polymorphonuclear leukocytes (PMN). Specific, saturable binding of SAP to FcγRI, FcγRIIa, and FcγRIIIb expressed on transfected COS cells was detected using SAP-biotin and PE-streptavidin. Zymosan was used to test the functional consequences of SAP and CRP binding to FcγR. Both SAP and CRP bound to zymosan and enhanced its uptake by PMN. This enhanced phagocytosis was abrogated by treatment of PMN with wortmannin, a phosphatidylinositol-3 kinase inhibitor, or with piceatannol, a Syk inhibitor, consistent with uptake through FcγR. Treatment of PMN with phosphatidylinositol-specific phospholipase C to remove FcγRIIIb also decreased phagocytosis of SAP-opsonized zymosan, but not CRP-opsonized zymosan. These results suggest that SAP may function in host defense. In addition, as SAP binds to chromatin, a major immunogen in systemic lupus erythematosus, it may provide a clearance mechanism for this Ag through FcγR bearing cells.

The pentraxins, serum amyloid P component (SAP) and C-reactive protein (CRP) are acute-phase serum proteins in mice and humans, respectively. Although SAP binds to DNA and chromatin and affects clearance of these autoantigens, no specific receptor for SAP has been identified. CRP is an opsonin, and we have shown that it binds to FcγR. Mice deficient in FcγR were used to assess the role of these receptors in phagocytosis by pentraxins using zymosan as a ligand. Phagocytosis of zymosan by bone marrow macrophages (BMM) was enhanced by opsonization with SAP or CRP. BMM from mice deficient in all three FcγR or in γ-chain ingested unopsonized zymosan, but phagocytosis of SAP- or CRP-opsonized zymosan was not enhanced. SAP binding to BMM from γ-chain-deficient mice was also greatly reduced, indicating little or no binding of SAP to FcγRII. SAP and CRP opsonized zymosan for phagocytosis by BMM from mice deficient in FcγRII or FcγRIII. SAP, but not CRP, opsonized zymosan for uptake by neutrophils that express only low levels of FcγRI. Together these results indicate that FcγRI and FcγRIII are receptors for SAP in the mouse. Opsonization of zymosan by CRP is mediated through FcγRI. Pentraxins are major proteins of the innate immune system and arose earlier in evolution than Igs. The use of FcγR by the pentraxins links innate and adaptive immunity and may have important consequences for processing, presentation, and clearance of the self-Ags to which these proteins bind.

Human C-reactive protein (CRP) is an acute phase protein that binds to receptors on human and mouse leukocytes. We have recently determined that the high and low affinity receptors for CRP on human leukocytes are Fc gamma RIIa and Fc gamma RI, respectively. Previous work by others suggested that CRP receptors on mouse macrophages are distinct from Fc gamma R. We have taken advantage of the availability of mice deficient in one or more Fc gamma R to reexamine the role of Fc gamma R in CRP binding to mouse leukocytes. Three strains of Fc gamma R-deficient mice were examined: gamma-chain-deficient mice that lack Fc gamma RI and Fc gamma RIII, Fc gamma RII-deficient mice, and mice deficient in both gamma-chain and Fc gamma RII that lack all Fc gamma R. No binding of CRP was detected to leukocytes from double-deficient mice, indicating that Fc gamma R are required for CRP binding. CRP binding to leukocytes from gamma-chain-deficient and Fc gamma RII-deficient mice was reduced compared with binding to leukocytes from wild-type mice. Further analysis of CRP binding to macrophages, neutrophils, and lymphocytes provides direct evidence that Fc gamma RIIb1, Fc gamma RIIb2, and Fc gamma RI are the receptors for CRP on mouse leukocytes. These findings may have important implications in understanding the physiological function of CRP.

C-reactive protein (CRP) is an acute-phase serum protein and a mediator of innate immunity. CRP binds to microbial polysaccharides and to ligands exposed on damaged cells. Binding of CRP to these substrates activates the classical complement pathway leading to their uptake by phagocytic cells. Complement activation by CRP is restricted to C1, C4, C2 and C3 with little consumption of C5-9. Surface bound CRP reduces deposition of and generation of C5b-9 by the alternative pathway and deposition of C3b and lysis by the lectin pathway. These activities of CRP are the result of recruitment of factor H resulting in regulation of C3b on bacteria or erythrocytes. Evidence is presented for direct binding of H to CRP. H binding to CRP or C3b immobilized on microtiter wells was demonstrated by ELISA. Attachment of CRP to a surface was required for H binding. H binding to CRP was not inhibited by EDTA or phosphocholine, which inhibit ligand binding, but was inhibited by a 13 amino acid CRP peptide. The peptide sequence was identical to the region of CRP that showed the best alignment to H binding peptides from Streptococcus pyogenes (M6) and Neisseria gonorrhoeae (Por1A). The results suggest that CRP bound to a surface provides secondary binding sites for H resulting in greater regulation of alternative pathway amplification and C5 convertases. Complement activation by CRP may help limit the inflammatory response by providing opsonization with minimal generation of C5a and C5b-9.

C-reactive protein (CRP) is an acute phase serum protein that shares several functions with immunoglobulin (Ig)G including complement activation and binding to receptors on monocytes and neutrophils. The identity of the receptor for CRP has been the target of extensive research. We previously determined that CRP binds to the high affinity receptor for IgG, FcgammaRI (CD64). However, this interaction could not account for the majority of binding of CRP to neutrophils or monocytic cells. We now determine that CRP also interacts with FcgammaRIIa (CD32), the low affinity receptor for IgG on monocytes and neutrophils. COS-7 cells were transfected with a construct containing the human FcgammaRIIA cDNA. CRP binding and the presence of CD32 were detected by mAb and analyzed by two-color flow cytometry. Cells expressing CD32 bound CRP in a dose-dependent and saturable manner consistent with receptor binding. CRP bound to transfectants and K-562 cells with similar kinetics, and in both cases binding was completely inhibited by aggregated IgG. On monocytic cell lines, treatment with Bt(2)cAMP increased FcgammaRII expression and enhanced CRP binding. CRP also specifically precipitated FcgammaRI and FcgammaRII from the monocytic cell line, THP-1. It is suggested that the major receptor for CRP on phagocytic cells is FcgammaRII.

Severe injury remains a leading cause of death and morbidity in patients under 40, with the number of annual trauma-related deaths approaching 160,000 in the United States. Patients who survive the initial trauma and post-traumatic resuscitation are at risk for immune dysregulation, which contributes to late mortality and accounts for approximately 20% of deaths after traumatic injury. This post-traumatic immunosuppressed state has been attributed to over-expression of anti-inflammatory mediators in an effort to restore host homeostasis. We measured a panel of monocyte markers and cytokines in 50 severely injured trauma patients for 3 days following admission. We made the novel observation that the subpopulation of monocytes expressing high levels of CD14 and CD16 was expanded in the majority of patients. These cells also expressed CD163 consistent with differentiation into alternatively activated macrophages with potential regulatory or wound-healing activity. We examined factors in trauma plasma that may contribute to the generation and activation of these cells. The percentage of CD14(high)CD16(+) monocytes after trauma correlated strongly with plasma C-reactive protein (CRP) transforming growth factor-β (TGF-β), and macrophage colony-stimulating factor (M-CSF) levels. We demonstrate a role for TGF-β and M-CSF, but not CRP in generating these cells using monocytes from healthy volunteers incubated with plasma from trauma patients. CD16 is a receptor for CRP and IgG, and we showed that monocytes differentiated to the CD14(high)CD16(+) phenotype produced anti-inflammatory cytokines in response to acute phase concentrations of CRP. The role of these cells in immunosuppression following trauma is an area of ongoing investigation.

C-reactive protein (CRP) binds to chromatin, histones, and small nuclear ribonucleoproteins (snRNPs) through a phosphocholine (PC)-inhibitable, calcium-dependent binding site. snRNPs process pre-mRNA to mature mRNA and are composed of small uridine-rich RNAs (designated U1, U2, U5 and U4/U6) and associated proteins. We have shown that CRP binds to snRNPs in intact cells and to the U1 snRNP-specific 70 K protein in cell extracts. To determine whether CRP bound to other snRNP proteins, snRNPs were purified from rabbit thymus extract and CRP binding was assessed by blotting. CRP bound to a protein with the same mobility as Sm-D as well as to the 70 K protein. CRP specifically bound to and precipitated a fusion protein containing full-length Sm-D, confirming the binding of CRP to Sm-D. Binding was inhibited by PC and by EDTA. Binding studies using deletion mutants of the Sm-D fusion protein revealed that CRP binding was mediated by the C-terminal region of Sm-D, a region which binds autoantibodies and is proposed to bind to RNA. A comparison of the peptide regions on different autoantigens suggests that there is a shared motif to which CRP binds.

Ischemia/reperfusion injury and organ allograft rejection both entail excessive cell and tissue destruction. A number of innate immune proteins, including the pentraxins, participate in the removal of this potentially inflammatory and autoimmunogenic material. The classical pentraxins, C-reactive protein (CRP) and serum amyloid P component (SAP) are serum opsonins, which bind to damaged membranes and nuclear autoantigens. Understanding the role of pentraxins in inflammation has been advanced by the recent identification and structural analysis of their receptor interactions.The ligand-binding, complement-activating and opsonic properties of pentraxins have been known for some time. However, the establishment of Fcγ receptors as the primary receptors for pentraxins is a recent finding with important implications for CRP and SAP functions. The crystal structure of SAP in complex with FcγRIIa was recently solved, leading to new insights into function and new opportunities for pentraxin-based therapeutics. In addition, new approaches to inhibit CRP synthesis or binding are being developed based on clinical associations between CRP levels and cardiovascular disease.This review will summarize data on the function of pentraxins in clearance of injured tissue and cells and discuss the implications of this clearance pathway for autoimmunity and ischemia/reperfusion injury.

C-reactive protein (CRP) is the prototypical acute phase serum protein, rising rapidly in response to inflammation. CRP binds to phosphocholine (PC) and related molecules on microorganisms and plays an important role in host defense. However, a more important role may be the binding of CRP to PC in damaged membranes. CRP increases clearance of apoptotic cells, binds to nuclear antigens and by masking autoantigens from the immune system or enhancing their clearance, CRP may prevent autoimmunity. CRP binds to both the stimulatory receptors, FcgammaRI and FcgammaRIIa, increasing phagocytosis and the release of inflammatory cytokines; and to the inhibitory receptor, FcgammaRIIb, blocking activating signals. We have shown that, in two animal models of systemic lupus erythematosus (SLE), the (NZB x NZW)F1 mouse and the MRL/lpr mouse, a single injection of CRP before onset of proteinuria delayed disease development and late treatment reversed proteinuria. Thus, in these models, CRP plays an anti-inflammatory role.

Objective—In addition to being a cardiovascular risk marker, recent studies support a role for CRP in atherothrombosis. Several investigators have reported that CRP binds to Fcgamma receptors on leukocytes. The aim of the study is to determine the processing of CRP by human aortic endothelial cells (HAECs).Methods and Results—Binding studies were performed by incubation of HAECs with biotinylated CRP (B-CRP, 25 to 200 μg/mL) for 30 to 180 minutes at 4°C. B-CRP binding was quantitated using streptavidin-fluorescein isothiocyanate followed by flow cytometry. Saturable binding of CRP was obtained at 60 minutes with a CRP concentration between 100 to 150 μg/mL and Kd of 88 nM. CRP binding was inhibited by 10× cold CRP (58%). CRP (100 μg/mL) significantly upregulated surface expression of Fcgamma receptors, CD32, as well as CD64 on HAECs (PConclusions—We demonstrate that CRP mediates its biological effects on HAECs via binding and internalization through Fcgamma receptors, CD32 and CD64.

To examine the relationship of serum C-reactive protein (CRP) levels to other indicators of disease activity during the course of systemic lupus erythematosus (SLE).In 124 patients serum CRP was measured retrospectively by ELISA and in some instances by radial immunodiffusion. Serum CRP levels were compared to laboratory, clinical, and radiographic assessments of disease activity. In many patients, serial CRP levels were measured over months or years to determine whether elevations of serum CRP reflected apparent changes in other disease activity variables. CRP was also measured in lyophilized aliquots of 24 h urine samples from SLE patients and controls with other renal disorders. Parallel determinations of interleukin 6 (IL-6) were made by ELISA in healthy controls and SLE patients.Of the 124 SLE patients studied, most showed elevations in serum CRP levels in the course of their disease. No inverse or direct correlation was noted between serum CRP and levels of nucleosome antigen or serum IgM or IgG anti-DNA antibody. In patients with renal involvement and proteinuria, CRP was often detected in 24-h urine samples. A strong correlation (p0.001) was noted between CRP and IL-6 levels in healthy subjects, but no correlation was recorded between serum CRP and IL-6 in SLE.Contrary to previous reports, most patients with SLE in our study showed elevations of serum CRP during the course of their illness, and extremely high serum CRP was recorded in some patients. CRP was also found in concentrated urine samples from patients with renal involvement and often paralleled elevated serum levels. In patients, no correlation was seen between CRP serum levels and serum IL-6, whereas a strong correlation between CRP level and IL-6 was recorded in healthy subjects.

C-reactive protein (CRP) is an acute phase serum protein that binds to phosphocholine (PC) on phospholipids and polysaccharides and to protein components of chromatin and small nuclear ribonucleoproteins. Complexes between CRP and ligands activate complement and bind to receptors on phagocytic cells. Although complement is required for CRP-mediated clearance or phagocytosis of ligand-coated erythrocytes, the participation of complement and complement receptors in clearance of soluble CRP complexes has not been examined. We have used PC-conjugated BSA to prepare complexes containing either IgG antibody or CRP. We found similar complement-mediated binding of both types of complexes to human erythrocyte complement receptors (CR1, CD35). We also found that serum deficient in C4A or C4B supported binding of CRP and IgG complexes to erythrocytes. These findings indicate that complexes between CRP and soluble ligands may be cleared by the erythrocyte CR1 pathway described for soluble immune complexes.

Two highly sensitive immunoassays for measuring soluble DNA are described. These methods employ a purified mouse monoclonal anti-DNA antibody in enzyme-linked immunoassays. One assay is an antigen capture method utilizing immobilized antibody. Bound DNA is subsequently detected with biotinylated monoclonal anti-DNA antibody, avidin-coupled horseradish peroxidase and a chromogenic substrate. The limit of detection of the assay was 2 ng/ml of DNA. A competition assay relying on immobilized heat-denatured DNA was also designed. In this assay the solution to be analyzed is mixed in equal proportions with monoclonal anti-DNA antibody and the mixture is incubated in the DNA-containing microtiter plates. The inhibition of antibody binding as detected with peroxidase-conjugated anti-mouse IgG was proportional to the concentration of DNA in the sample. The competition assay was tested for its ability to detect DNA in human plasma. The detection limit of DNA in plasma was approximately 150 ng/ml. The assays were compared in their ability to detect various size fragments of DNA. The competition assay was capable of detecting DNA fragments as small as 30 base pairs. In contrast, the capture assay failed to detect low molecular weight fragments up to 150 base pairs although its sensitivity for undigested DNA was comparable to the competition assay. The assay may be of use in the rapid quantitation of low levels of DNA, especially low molecular weight DNA and may also be useful in measuring DNA in human plasma.

In the studies described here we have attempted to evaluate the hypothesis that CRP may function in host defense using two systems in which CRP in the presence of C appears to have opsonic properties. In the first, CRP and C were found to stimulate ingestion of erythrocytes by human monocyte or mouse macrophages in vitro, and to alter clearance patterns in vivo. In the second, we have studied opsonization of S. pneumoniae by CRP and C. Experiments with human neutrophils indicate that although CRP and C can enhance opsonization of S. pneumoniae, this effect is more pronounced in the absence of antibody. In vivo CRP was found to protect mice against intravenous infection with S. pneumoniae.