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Infection with severe acute respiratory syndrome coronavirus 2 associates with diverse symptoms, which can persist for months. While antiviral antibodies are protective, those targeting interferons and other immune factors are associated with adverse coronavirus disease 2019 (COVID-19) outcomes. Here we discovered that antibodies against specific chemokines were omnipresent post-COVID-19, were associated with favorable disease outcome and negatively correlated with the development of long COVID at 1 yr post-infection. Chemokine antibodies were also present in HIV-1 infection and autoimmune disorders, but they targeted different chemokines compared with COVID-19. Monoclonal antibodies derived from COVID-19 convalescents that bound to the chemokine N-loop impaired cell migration. Given the role of chemokines in orchestrating immune cell trafficking, naturally arising chemokine antibodies may modulate the inflammatory response and thus bear therapeutic potential.

The MS4A gene family encodes 18 tetraspanin‐like proteins, most of which with unknown function. MS4A1 (CD20), MS4A2 (FcεRIβ), MS4A3 (HTm4), and MS4A4A play important roles in immunity, whereas expression and function of other members of the family are unknown. The present investigation was designed to obtain an expression fingerprint of MS4A family members, using bioinformatics analysis of public databases, RT‐PCR, and protein analysis when possible. MS4A3, MS4A4A, MS4A4E, MS4A6A, MS4A7, and MS4A14 were expressed by myeloid cells. MS4A6A and MS4A14 were expressed in circulating monocytes and decreased during monocyte‐to‐Mϕ differentiation in parallel with an increase in MS4A4A expression. Analysis of gene expression regulation revealed a strong induction of MS4A4A, MS4A6A, MS4A7, and MS4A4E by glucocorticoid hormones. Consistently with in vitro findings, MS4A4A and MS4A7 were expressed in tissue Mϕs from COVID‐19 and rheumatoid arthritis patients. Interestingly, MS4A3, selectively expressed in myeloid precursors, was found to be a marker of immature circulating neutrophils, a cellular population associated to COVID‐19 severe disease. The results reported here show that members of the MS4A family are differentially expressed and regulated during myelomonocytic differentiation, and call for assessment of their functional role and value as therapeutic targets. MS4A tetraspan molecules are differentially expressed and regulated in myeloid cells under normal and pathological conditions.

The humoral arm of innate immunity includes diverse molecules with antibody-like functions, some of which serve as disease severity biomarkers in coronavirus disease 2019 (COVID-19). The present study was designed to conduct a systematic investigation of the interaction of human humoral fluid-phase pattern recognition molecules (PRMs) with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Of 12 PRMs tested, the long pentraxin 3 (PTX3) and mannose-binding lectin (MBL) bound the viral nucleocapsid and spike proteins, respectively. MBL bound trimeric spike protein, including that of variants of concern (VoC), in a glycan-dependent manner and inhibited SARS-CoV-2 in three in vitro models. Moreover, after binding to spike protein, MBL activated the lectin pathway of complement activation. Based on retention of glycosylation sites and modeling, MBL was predicted to recognize the Omicron VoC. Genetic polymorphisms at the MBL2locus were associated with disease severity. These results suggest that selected humoral fluid-phase PRMs can play an important role in resistance to, and pathogenesis of, COVID-19, a finding with translational implications.

Long pentraxin 3 (PTX3) is an essential component of humoral innate immunity, involved in resistance to selected pathogens and in the regulation of inflammation1–3. The present study was designed to assess the presence and significance of PTX3 in Coronavirus Disease 2019 (COVID-19)4–7. RNA-sequencing analysis of peripheral blood mononuclear cells, single-cell bioinformatics analysis and immunohistochemistry of lung autopsy samples revealed that myelomonocytic cells and endothelial cells express high levels of PTX3 in patients with COVID-19. Increased plasma concentrations of PTX3 were detected in 96 patients with COVID-19. PTX3 emerged as a strong independent predictor of 28-d mortality in multivariable analysis, better than conventional markers of inflammation, in hospitalized patients with COVID-19. The prognostic significance of PTX3 abundance for mortality was confirmed in a second independent cohort (54 patients). Thus, circulating and lung myelomonocytic cells and endothelial cells are a major source of PTX3, and PTX3 plasma concentration can serve as an independent strong prognostic indicator of short-term mortality in COVID-19.

The plasma membrane tetraspan molecule MS4A4A is selectively expressed by macrophage-lineage cells, but its function is unknown. Here we report that MS4A4A was restricted to murine and human mononuclear phagocytes and was induced during monocyte-to-macrophage differentiation in the presence of interleukin 4 or dexamethasone. Human MS4A4A was co-expressed with M2/M2-like molecules in subsets of normal tissue-resident macrophages, infiltrating macrophages from inflamed synovium and tumor-associated macrophages. MS4A4A interacted and colocalized with the β-glucan receptor dectin-1 in lipid rafts. In response to dectin-1 ligands, Ms4a4a-deficient macrophages showed defective signaling and defective production of effector molecules. In experimental models of tumor progression and metastasis, Ms4a4a deficiency in macrophages had no impact on primary tumor growth, but was essential for dectin-1-mediated activation of macrophages and natural killer (NK) cell–mediated metastasis control. Thus, MS4A4A is a tetraspan molecule selectively expressed in macrophages during differentiation and polarization, essential for dectin-1-dependent activation of NK cell–mediated resistance to metastasis. The membrane tetraspan protein MS4A4A is expressed on tissue-resident and tumor-associated macrophages. Locati and colleagues show that MS4A4A colocalizes with the β-glucan receptor dectin-1 to enhance cytokine and reactive oxygen species production and to enhance NK cell–mediated control of tumor metastasis.

AbstractObjective: We studied the impact of chlorinated agents exposure on exhaled breath condensate (EBC) biomarkers in cleaners.Methods: Malondialdehyde (MDA), 4-hydroxynonenal (4-HNE), nitrites (), nitrates (), pH, hydrogen peroxide (H2O2) and ammonium () were tested in EBC of 40 cleaners and 40 non-exposed controls. Pentraxin-3 (PTX3) and soluble type II receptor of IL-1 (sIL-1RII) were analyzed also in plasma.Results: Levels of MDA-EBC, 4-HNE-EBC and -EBC were higher, while pH-EBC values were lower, in cleaners. MDA-EBC was associated with 4-HNE-EBC, -EBC and pH. 4-HNE-EBC correlated with PTX3.Conclusion: Professional exposure to chlorinated agents increases EBC biomarkers of oxidative stress and inflammation.

Although mesenchymal stromal cells (MSCs) can promote wound healing in different clinical settings, the underlying mechanism of MSC-mediated tissue repair has yet to be determined. Because a nonredundant role of pentraxin 3 (PTX3) in tissue repair and remodeling has been recently described, here we sought to determine whether MSC-derived PTX3 might play a role in wound healing. Using a murine model of skin repair, we found that Ptx3-deficient (Ptx3−/−) MSCs delayed wound closure and reduced granulation tissue formation compared with wt MSCs. At day 2, confocal microscopy revealed a dramatic reduction in green fluorescent protein (GFP)-expressing Ptx3−/−MSCs recruited to the wound, where they appeared to be not only poorly organized in bundles but also scattered in the extracellular matrix. These findings were further confirmed by quantitative biochemical analysis of GFP content in wound extracts. Furthermore, Ptx3−/−MSC-treated skins displayed increased levels of fibrin and lower levels of D-dimer, suggesting delayed fibrin-rich matrix remodeling compared with control skins. Consistently, both pericellular fibrinolysis and migration through fibrin were found to be severely affected in Ptx3−/−MSCs. Overall, our findings identify an essential role of MSC-derived PTX3 in wound repair underscoring the beneficial potential of MSC-based therapy in the management of intractable wounds.

Pentraxin 3 (PTX3) is a fluid-phase pattern recognition molecule and a key component of the humoral arm of innate immunity. In four different models of tissue damage in mice, PTX3 deficiency was associated with increased fibrin deposition and persistence, and thicker clots, followed by increased collagen deposition, when compared with controls. Ptx3-deficient macrophages showed defective pericellular fibrinolysis in vitro. PTX3-bound fibrinogen/fibrin and plasminogen at acidic pH and increased plasmin-mediated fibrinolysis. The second exon-encoded N-terminal domain of PTX3 recapitulated the activity of the intact molecule. Thus, a prototypic component of humoral innate immunity, PTX3, plays a nonredundant role in the orchestration of tissue repair and remodeling. Tissue acidification resulting from metabolic adaptation during tissue repair sets PTX3 in a tissue remodeling and repair mode, suggesting that matrix and microbial recognition are common, ancestral features of the humoral arm of innate immunity.

Progression of arterial involvement is often observed in patients with Takayasu arteritis (TA) thought to be in remission. This reflects the failure of currently used biomarkers and activity criteria to detect smouldering inflammation occurring within arterial wall. Pentraxin-3 (PTX3) is a soluble pattern recognition receptor produced at sites of inflammation and could reveal systemic as well as localized inflammatory processes. We verified whether the blood concentrations of PTX3 and of C-reactive protein (CRP) in patients with Takayasu arteritis (TA) might reflect vascular wall involvement, as assessed by signal enhancement after contrast media administration, and the progression of arterial involvement. A cross-sectional single-centre study was carried out on 42 patients with TA that comprised assessment of PTX3, of CRP and erythrocyte sedimentation velocity (ESR). In total, 20 healthy controls and 20 patients with Systemic Lupus Erythematous (SLE) served as controls. Vascular imaging was carried out by magnetic resonance angiography, doppler ultrasonography and computed tomography angiography. Patients with TA and SLE had higher plasmatic PTX3 and CRP concentrations than healthy controls (P= 0.009 and 0.017, respectively). PTX3 levels did not correlate with those of CRP. Patients with active systemic TA had significantly higher concentrations of CRP but similar levels of PTX3 than patients with quiescent disease. In contrast, patients with vascular inflammation detectable at imaging had higher PTX3 concentrations (P= 0.016) than those in which vessel inflammation was not evident, while CRP levels were similar. The concentration of PTX3 but not that of CRP was significantly higher in TA patients with worsening arterial lesions that were not receiving antagonists of tumor necrosis factor-α or interleukin-6. Arterial inflammation and progression of vascular involvement influence plasma PTX3 levels in TA, while levels of CRP accurately reflect the burden of systemic inflammation. These results support the contention that PTX3 reflects different aspects of inflammation than CRP and might represent a biomarker of actual arteritis in TA.

The prototypic long pentraxin PTX3, a soluble pattern recognition receptor, plays an important role in innate defense against selected pathogens by favoring their elimination and the initiation of protective responses. PTX3 has notably beneficial effects in mice infected with Aspergillus fumigatus and Pseudomonas aeruginosa. Cystic fibrosis (CF), a severe inherited autosomal recessive disease, is characterized by recurrent lung infections, especially by these two pathogens. We thus hypothesized that the status of PTX3 may be altered in CF patients. Level and integrity of PTX3 were analyzed in the sputum samples from 51 CF patients and 7 patients with chronic obstructive pulmonary disease (COPD). The levels of PTX3 were increased in serums from CF patients, but low in their respiratory secretions. PTX3 concentrations in sputum samples were dramatically lower in CF patients than in COPD patients. The low concentration of PTX3 resulted from a proteolysis cleavage by elastase and A. fumigatus proteases. Interestingly, the N-ter domain of PTX3, involved in protection against A. fumigatus, is preferentially degraded by these proteases. These results indicate that the selective proteolysis of PTX3 in the CF lung may explain, in part, the recurrent lung infections by PTX3-sensitive pathogens in CF patients.

Pentraxins are a family of evolutionarily conserved multifunctional pattern-recognition proteins characterized by a cyclic multimeric structure. Based on the primary structure of the subunit, the pentraxins are divided into two groups: short pentraxins and long pentraxins. C-reactive protein (CRP) and serum amyloid P-component (SAP) are the two short pentraxins. The prototype protein of the long pentraxin group is pentraxin 3 (PTX3). CRP and SAP are produced primarily in the liver in response to IL-6, while PTX3 is produced by a variety of tissues and cells and in particular by innate immunity cells in response to proinflammatory signals and Toll-like receptor (TLR) engagement. PTX3 interacts with several ligands, including growth factors, extracellular matrix components and selected pathogens, playing a role in complement activation and facilitating pathogen recognition by phagocytes, acting as a predecessor of antibodies. In addition, PTX3 is essential in female fertility by acting as a nodal point for the assembly of the cumulus oophorus hyaluronanrich extracellular matrix. Here we will concisely review the general properties of PTX3 in the context of the pentraxin superfamily and discuss recent data suggesting that PTX3 plays a cardiovascular protective effect. PTX3 may represent a new marker in vascular pathology which correlates with the risk of developing vascular events.

Pentraxin 3 (PTX3) is a soluble pattern recognition molecule playing a nonredundant role in resistance against Aspergillus fumigatus. The present study was designed to investigate the molecular pathways involved in the opsonic activity of PTX3. The PTX3 N-terminal domain was responsible for conidia recognition, but the full-length molecule was necessary for opsonic activity. The PTX3-dependent pathway of enhanced neutrophil phagocytic activity involved complement activation via the alternative pathway; Fcγ receptor (FcγR) IIA/CD32 recognition of PTX3-sensitized conidia and complement receptor 3 (CR3) activation; and CR3 and CD32 localization to the phagocytic cup. Gene targeted mice (ptx3, FcR common γ chain, C3, C1q) validated the in vivo relevance of the pathway. In particular, the protective activity of exogenous PTX3 against A fumigatuswas abolished in FcR common γ chain-deficient mice. Thus, the opsonic and antifungal activity of PTX3 is at the crossroad between complement, complement receptor 3-, and FcγR-mediated recognition. Because short pentraxins (eg, C-reactive protein) interact with complement and FcγR, the present results may have general significance for the mode of action of these components of the humoral arm of innate immunity.

Pentraxin 3 (PTX3) is a soluble pattern recognition molecule playing a nonredundant role in resistance against Aspergillus fumigatus. The present study was designed to investigate the molecular pathways involved in the opsonic activity of PTX3. The PTX3 N-terminal domain was responsible for conidia recognition, but the full-length molecule was necessary for opsonic activity. The PTX3-dependent pathway of enhanced neutrophil phagocytic activity involved complement activation via the alternative pathway; Fcγ receptor (FcγR) IIA/CD32 recognition of PTX3-sensitized conidia and complement receptor 3 (CR3) activation; and CR3 and CD32 localization to the phagocytic cup. Gene targeted mice (ptx3, FcR common γ chain, C3, C1q) validated the in vivo relevance of the pathway. In particular, the protective activity of exogenous PTX3 against A fumigatus was abolished in FcR common γ chain-deficient mice. Thus, the opsonic and antifungal activity of PTX3 is at the crossroad between complement, complement receptor 3-, and FcγR-mediated recognition. Because short pentraxins (eg, C-reactive protein) interact with complement and FcγR, the present results may have general significance for the mode of action of these components of the humoral arm of innate immunity.

ObjectivePentraxin 3 PTX3, a key component of innate immunity, is a strong marker of disease severity in coronary artery disease CAD. The aim of this study was to compare levels of serum PTX3 in CAD patients with and without inflammatory rheumatic disease IRD and in healthy controls.MethodsWe examined 69 patients with IRD CADIRD group and 53 patients without IRD CADnonIRD referred to coronary artery bypass grafting, and 30 healthy controls.ResultsThe mean ± SD serum PTX3 level in the CADIRD group was 1.96 ± 0.98 ngml; this was statistically significantly higher than that of the CADnonIRD 1.41 ± 0.74 ngml and healthy control 1.21 ± 0.59 ngml groups. In contrast to most other IRDs, serum PTX3 levels were relatively low in patients with systemic lupus erythematosus SLE and other systemic connective tissue diseases. In sex and ageadjusted analysis, IRD, acute coronary syndromes, and low alcohol intake were associated with higher serum PTX3 levels.ConclusionCAD patients with IRD had higher mean serum PTX3 levels than patients without IRD and healthy controls. In addition, acute coronary syndromes and low alcohol intake independently predicted higher serum PTX3 levels. Higher serum PTX3 levels in IRD may be related to the higher cardiovascular risk of IRD patients. Circulating PTX3 could likely be used as a biomarker for severity of cardiovascular disease in IRDs; its importance, however, might be limited in SLE and related disorders.

Pentraxins are a family of evolutionarily conserved multifunctional pattern-recognition proteins characterized by a cyclic multimeric structure. Based on the primary structure of the subunit, the pentraxins are divided into two groups: short pentraxins and long pentraxins. C-reactive protein (CRP) and serum amyloid P-component (SAP) are the two short pentraxins. The prototype protein of the long pentraxin group is pentraxin 3 (PTX3). CRP and SAP are produced primarily in the liver in response to IL-6, while PTX3 is produced by a variety of tissues and cells and in particular, by innate immunity cells in response to proinflammatory signals and Toll-like receptor (TLR) engagement. PTX3 interacts with several ligands, including growth factors, extracellular matrix components and selected pathogens, playing a role in complement activation and facilitating pathogen recognition by phagocytes, acting as a predecessor of antibodies. In addition, PTX3 is essential in female fertility by acting as a nodal point for the assembly of the cumulus oophorus hyaluronan-rich extracellular matrix. Thus, the prototypic long pentraxin PTX3 is a multifunctional soluble pattern recognition receptor acting as a nonredundant component of the humoral arm of innate immunity and involved in tuning inflammation, in matrix deposition and female fertility.

The long pentraxin 3 (PTX3) is member of a complex superfamily of multifunctional proteins characterized by a cyclic multimeric structure. PTX3 is highly conserved in evolution and is produced by innate‐immunity cells in response to proinflammatory signals and Toll‐like receptor engagement. PTX3 plays complex, nonredundant functions in vivo, acting as a predecessor of antibodies, recognizing microbes, activating complement, facilitating pathogen recognition by phagocytes, and hence, playing a nonredundant role in resistance against selected pathogens. In addition, PTX3 is essential in female fertility by acting as a nodal point for the assembly of the cumulus oophorus hyaluronan‐rich extracellular matrix. Thus, the prototypic long pentraxin PTX3 is a multifunctional, soluble pattern recognition receptor acting as a nonredundant component of the humoral arm of innate immunity and involved in matrix deposition and female fertility.

The protopypic long pentraxin 3 (PTX3) is a unique, humoral pattern‐recognition receptor, which plays a nonredundant function in innate resistance to pathogens. Dendritic cells (DC) of myelomonocytic origin, but not plasmacytoid DC, are a major source of PTX3 in response to Toll‐like receptor (TLR) engagment. The present study was designed to explore the regulation of PTX3 production in DC. PTX3 production was induced by TLR ligands, CD40 ligand, and interleukin (IL)‐1β and was suppressed by dexamethasone, 1α, 25‐dihydroxivitamin D3, and prostaglandin E2. It was unexpected that lipopolysaccharide (LPS)‐stimulated PTX3 production was enhanced by IL‐10 and inhibited by IL‐4 and interferon‐γ (IFN‐γ). Enhancement of PTX3 production by IL‐10 was also evident when Pam3Cys‐Ser‐(Lys)4.3HCl, a TLR2‐TLR1 agonist, polyionisicpolycytidylic acid, a TLR3 agonist, and IL‐1β were used as stimuli. The effect of IL‐10 was blocked by an anti‐IL‐10 monoclonal antibody (mAb) or an anti‐IL‐10 receptor α mAb, which also reduced the LPS‐induced production. Thus, production of PTX3 in DC is subjected to a distinct regulatory network, with inhibition by IFN‐γ and enhancement by IL‐10. The amplification by IL‐10 of production of a nonredundant component of fluid‐phase innate immunity mirrors the IL‐10 stimulatory function on B cells in adaptive immunity. As PTX3 is also an extracellular matrix component, IL‐10‐enhanced PTX3 production may play a role in orchestration of tissue remodeling in chronic inflammation.

To identify the molecular basis underlying the functions of tumor-associated macrophages (TAMs), we characterized the gene expression profile of TAMs isolated from a murine fibrosarcoma in comparison with peritoneal macrophages (PECs) and myeloid suppressor cells (MSCs), using a cDNA microarray technology. Among the differentially expressed genes, 15 genes relevant to inflammation and immunity were validated by real-time polymerase chain reaction (PCR) and protein production. Resting TAMs showed a characteristic gene expression pattern with higher expression of genes coding for the immunosuppressive cytokine IL-10, phagocytosis-related receptors/molecules (Msr2 and C1q), and inflammatory chemokines (CCL2 and CCL5) as expected, as well as, unexpectedly, IFN-inducible chemokines (CXCL9, CXCL10, CXCL16). Immunohistology confirmed and extended the in vitro analysis by showing that TAMs express M2-associated molecules (eg, IL-10 and MGL1), as well as CCL2, CCL5, CXCL9, CXCL10, and CXCL16, but no appreciable NOS2. Lipopolysaccharide (LPS)-mediated activation of TAMs resulted in defective expression of several proinflammatory cytokines (eg, IL-1β, IL-6, TNF-α) and chemokines (eg, CCL3), as opposed to a strong up-regulation of immunosuppressive cytokines (IL-10, TGFβ) and IFN-inducible chemokines (CCL5, CXCL9, CXCL10, CXCL16). Thus, profiling of TAMs from a murine sarcoma revealed unexpected expression of IFN-inducible chemokines, associated with an M2 phenotype (IL-10high, IL-12low), and divergent regulation of the NF-κB versus the IRF-3/STAT1 pathway.

To identify the molecular basis underlying the functions of tumor-associated macrophages (TAMs), we characterized the gene expression profile of TAMs isolated from a murine fibrosarcoma in comparison with peritoneal macrophages (PECs) and myeloid suppressor cells (MSCs), using a cDNA microarray technology. Among the differentially expressed genes, 15 genes relevant to inflammation and immunity were validated by real-time polymerase chain reaction (PCR) and protein production. Resting TAMs showed a characteristic gene expression pattern with higher expression of genes coding for the immunosuppressive cytokine IL-10, phagocytosis-related receptors/molecules (Msr2 and C1q), and inflammatory chemokines (CCL2 and CCL5) as expected, as well as, unexpectedly, IFN-inducible chemokines (CXCL9, CXCL10, CXCL16). Immunohistology confirmed and extended the in vitro analysis by showing that TAMs express M2-associated molecules (eg, IL-10 and MGL1), as well as CCL2, CCL5, CXCL9, CXCL10, and CXCL16, but no appreciable NOS2. Lipopolysaccharide (LPS)-mediated activation of TAMs resulted in defective expression of several proinflammatory cytokines (eg, IL-1β, IL-6, TNF-α) and chemokines (eg, CCL3), as opposed to a strong up-regulation of immunosuppressive cytokines (IL-10, TGFβ) and IFN-inducible chemokines (CCL5, CXCL9, CXCL10, CXCL16). Thus, profiling of TAMs from a murine sarcoma revealed unexpected expression of IFN-inducible chemokines, associated with an M2 phenotype (IL-10high, IL-12low), and divergent regulation of the NF-κB versus the IRF-3/STAT1 pathway.

The long pentraxin PTX3 is a soluble pattern recognition receptor produced by monocytes and endothelial cells that plays a nonredundant role in inflammation. Several pathologic conditions are characterized by local production of both PTX3 and the angiogenic fibroblast growth factor-2 (FGF2). Here, solid-phase binding assays demonstrated that PTX3 binds with high affinity to FGF2 but not to a panel of cytokines and growth factors, including FGF1, FGF4, and FGF8. Accordingly, PTX3 prevented 125I-FGF2 binding to endothelial cell receptors, leading to specific inhibition of FGF2-induced proliferation. PTX3 hampered also the motogenic activity exerted by endogenous FGF2 on a wounded endothelial cell monolayer. Moreover, PTX3 cDNA transduction in FGF2-transformed endothelial cells inhibited their autocrine FGF2-dependent proliferation and morphogenesis in vitro and their capacity to generate vascular lesions when injected in nude mice. Finally, PTX3 suppressed neovascularization triggered by FGF2 in the chick embryo chorioallantoic membrane with no effect on physiologic angiogenesis. In contrast, the short pentraxin C-reactive protein was a poor FGF2 ligand/antagonist. These results establish the selective binding of a member of the pentraxin superfamily to a growth factor. PTX3/FGF2 interaction may modulate angiogenesis in various physiopathologic conditions driven by inflammation, innate immunity, and/or neoplastic transformation.

The long pentraxin PTX3 is a soluble pattern recognition receptor produced by monocytes and endothelial cells that plays a nonredundant role in inflammation. Several pathologic conditions are characterized by local production of both PTX3 and the angiogenic fibroblast growth factor-2 (FGF2). Here, solid-phase binding assays demonstrated that PTX3 binds with high affinity to FGF2 but not to a panel of cytokines and growth factors, including FGF1, FGF4, and FGF8. Accordingly, PTX3 prevented 125I-FGF2 binding to endothelial cell receptors, leading to specific inhibition of FGF2-induced proliferation. PTX3 hampered also the motogenic activity exerted by endogenous FGF2 on a wounded endothelial cell monolayer. Moreover, PTX3 cDNA transduction in FGF2-transformed endothelial cells inhibited their autocrine FGF2-dependent proliferation and morphogenesis in vitro and their capacity to generate vascular lesions when injected in nude mice. Finally, PTX3 suppressed neovascularization triggered by FGF2 in the chick embryo chorioallantoic membrane with no effect on physiologic angiogenesis. In contrast, the short pentraxin C-reactive protein was a poor FGF2 ligand/antagonist. These results establish the selective binding of a member of the pentraxin superfamily to a growth factor. PTX3/FGF2 interaction may modulate angiogenesis in various physiopathologic conditions driven by inflammation, innate immunity, and/or neoplastic transformation.

Pentraxin 3 (PTX3) is a recently characterized member of the pentraxin family of acute-phase proteins produced during inflammation. Classical short pentraxins, C-reactive protein, and serum amyloid P component can bind to C1q and thereby activate the classical complement pathway. Since PTX3 can also bind C1q, the present study was designed to define the interaction between PTX3 and C1q and to examine the functional consequences of this interaction. A dose-dependent binding of both C1q and the C1 complex to PTX3 was observed. Experiments with recombinant globular head domains of human C1q A, B, and C chains indicated that C1q interacts with PTX3 via its globular head region. Binding of C1q to immobilized PTX3 induced activation of the classical complement pathway as assessed by C4 deposition. Furthermore, PTX3 enhanced C1q binding and complement activation on apoptotic cells. However, in the fluid-phase, pre-incubation of PTX3 with C1q resulted in inhibition of complement activation by blocking the interaction of C1q with immunoglobulins. These results indicate that PTX3 can both inhibit and activate the classical complement pathway by binding C1q, depending on the way it is presented. PTX3 may therefore be involved in the regulation of the innate immune response.

Pentraxins are acute-phase proteins produced in vivo during inflammatory reactions. Classical short pentraxins, C-reactive protein, and serum amyloid P component are generated in the liver in response to interleukin (IL)–6. The long pentraxin PTX3 is produced in tissues under the control of primary proinflammatory signals, such as lipopolysaccharide, IL-1ß, and tumor necrosis factor-a, which also promote maturation of dendritic cells (DCs). Cell death commonly occurs during inflammatory reactions. In this study, it is shown that PTX3 specifically binds to dying cells. The binding was dose dependent and saturable. Recognition was restricted to extranuclear membrane domains and to a chronological window after UV irradiation or after CD95 cross-linking–induced or spontaneous cell death in vitro. PTX3 bound to necrotic cells to a lesser extent. Human DCs failed to internalize dying cells in the presence of PTX3, while they took up normally soluble or inert particulate substrates. These results suggest that PTX3 sequesters cell remnants from antigen-presenting cells, possibly contributing to preventing the onset of autoimmune reactions in inflamed tissues.

Pentraxins are acute-phase proteins produced in vivo during inflammatory reactions. Classical short pentraxins, C-reactive protein, and serum amyloid P component are generated in the liver in response to interleukin (IL)–6. The long pentraxin PTX3 is produced in tissues under the control of primary proinflammatory signals, such as lipopolysaccharide, IL-1β, and tumor necrosis factor-α, which also promote maturation of dendritic cells (DCs). Cell death commonly occurs during inflammatory reactions. In this study, it is shown that PTX3 specifically binds to dying cells. The binding was dose dependent and saturable. Recognition was restricted to extranuclear membrane domains and to a chronological window after UV irradiation or after CD95 cross-linking–induced or spontaneous cell death in vitro. PTX3 bound to necrotic cells to a lesser extent. Human DCs failed to internalize dying cells in the presence of PTX3, while they took up normally soluble or inert particulate substrates. These results suggest that PTX3 sequesters cell remnants from antigen-presenting cells, possibly contributing to preventing the onset of autoimmune reactions in inflamed tissues.

Three malignant tumors (3LL carcinoma, mFS6 sarcoma, and B16 F1melanoma) were transplanted in mice with congenital (beige) or acquired (antiasialo GM1‐treated) defects of natural killer cell (NK) activity. The macrophage content of the neoplastic tissues was not influenced by host NK activity levels. These data suggest that NK cell‐mediated resistance does not play an appreciable role in the regulation of the levels of tumor‐associated macrophages in established malignancy.

Two major changes in RNA metabolism occured in macrophages expressing tumoricidal activity: a down regulation of total RNA synthesis and an imbalanced accumulation of ribosomal RNA precursors with double stranded secondary structure. The aim of this study was to investigate a possible role of endogenous ds rRNA precursors in the activation of the dsRNA dependent protein kinase. Using a cell free transcription system purified rRNA precursors were obtained from the murine rRNA gene. The results indicate that rRNA precursors are potent activators of the dsPK. The effects are dose dependent and are not affected by treatment of rRNA precursors with proteinase K or RNase A. The treatment with RNase V1, specific for dsRNA, completely abrogates the activity of transcripts. The results suggest that endogenous RNA, namely rRNA, could control dsPK activation and it can be speculated that dsPK activation may be involved in the control of tumoricidal activity by macrophages.

Pentraxins, which include C reactive protein (CRP) and serum amyloid P component (SAP), are prototypic acute phase reactants that serve as indicators of inflammatory reactions. Here we report genomic and cDNA cloning of mouse ptx3 (mptx3), a member of the pentraxin gene family and characterize its extrahepatic expression in vitro and in vivo. mptx3 is organized into three exons on chromosome 3: the first (43 aa) and second exon (175 aa) code for the signal peptide and for a protein portion with no high similarity to known sequences, the third (203 aa) for a domain related to classical pentraxins, which contains the “pentraxin family signature.” Analysis of the N terminal portion predicts a predominantly α helical structure, while the pentraxin domain of ptx3 is accommodated comfortably in the tertiary structure fold of SAP. Normal and transformed fibroblasts, undifferentiated and differentiated myoblasts, normal endothelial cells, and mononuclear phagocytes express mptx3 mRNA and release the protein in vitro on exposure to interleukin-1β (IL-1β) and tumor necrosis factor (TIMF)α. mptx3 was induced by bacterial lipopolysaccharide in vivo in a variety of organs and, most strongly, in the vascular endothelium of skeletal muscle and heart. Thus, mptx3 shows a distinct pattern of in vivo expression indicative of a significant role in cardiovascular and inflammatory pathology.

The pentraxins C-reactive protein (CRP) and serum amyloid P component (SAP) are acute-phase proteins produced by liver epithelial cells. PTX3 was recently cloned as an interleukin-1 (IL-1)-inducible gene in endothelial cells, with structural similarities to pentraxins in the C-terminal half of the molecule. The present study was designed to investigate the expression of PTX3 in the human leukocyte populations. Human peripheral blood mononuclear cells exposed to lipopolysaccharide (LPS) or IL-1β expressed significant levels of PTX3 mRNA. Tumor necrosis factor-α (TNF-α) was a lesseffective inducer of PTX3, whereas IL-6, monocyte chemotactic protein-1, macrophage colony-stimulating factor, granulocyte-macrophage colony-stimulating factor, and interferon-γ were inactive. Among leukocytes, only monocytes exposed to inflammatory cytokines or LPS expressed the PTX3 transcript, which was undetectable in resting or stimulated polymorphonuclear cells, T or B lymphocytes, and natural killer cells. PTX3 mRNA was also inducible in in vitro monocyte-derived macrophages, in tumor-associated macrophages, and in the myelomonocytic cell lines HL60, U937, and THP1, but not in GF08, with the latter possibly representative of earlier stages of myelomonocytic differentiation. T- and B-cell lines had no detectable PTX3. Inhibition of transcription by actinomycin D blocked induction of PTX3 in monocytes and nuclear run-on analysis showed that LPS induces the expression of the PTX3 gene at the transcriptional level in isolated monocytes. Cycloheximide had no effect on PTX3 induction in U937 cells, but was inhibitory on monocytes exposed to LPS or IL-1β. Monoclonal antibody against TNF and the IL-1 receptor antagonist did not inhibit induction of PTX3 in monocytes by LPS, thus excluding these cytokines as secondary stimulators of PTX3. IL-4, but not dexamethasone or transforming growth factor-β, inhibited PTX3 expression in monocytes. Using a PTX3-specific antiserum, release of PTX3 protein was demonstrated for the first time in stimulated monocytes as well as in endothelial and fibroblastic cells. Thus, PTX3, unlike the classical pentraxins CRP and SAP, is expressed and released by cells of the monocyte-macrophage lineage exposed to inflammatory signals.

PTX3 is a prototypic long pentraxin consisting of a C-terminal 203-amino acid pentraxin-like domain coupled with an N-terminal 178-amino acid unrelated portion. The present study was designed to characterize the structure and ligand binding properties of human PTX3, in comparison with the classical pentraxins C-reactive protein and serum amyloid P component. Sequencing of Chinese hamster ovary cell-expressed PTX3 revealed that the mature secreted protein starts at residue 18 (Glu). Lectin binding and treatment with N-glycosidase F showed that PTX3 is N-glycosylated, sugars accounting for 5 kDa of the monomer mass (45 kDa). Circular dichroism analysis indicated that the protein consists predominantly of β-sheets with a minor α-helical component. While in gel filtration the protein is eluted with a molecular mass of ≅900 kDa, gel electrophoresis using nondenaturing, nonreducing conditions revealed that PTX3 forms multimers predominantly of 440 kDa apparent molecular mass, corresponding to decamers, and that disulfide bonds are required for multimer formation. The ligand binding properties of PTX3 were then examined. As predicted based on modeling, inductive coupled plasma/atomic emission spectroscopy showed that PTX3 does not have coordinated Ca2+. Unlike the classical pentraxins CRP and SAP, PTX3 did not bind phosphoethanolamine, phosphocholine, or high pyruvate agarose. PTX3 in solution, bound to immobilized C1q, but not C1s, and, reciprocally, C1q bound to immobilized PTX3. Binding of PTX3 to C1q is specific and saturable with a Kd7.4 × 10−8mas determined by solid phase binding assay. The Chinese hamster ovary cell-expressed pentraxin domain bound C1q when multimerized. Thus, as predicted on the basis of computer modeling, the prototypic long pentraxin PTX3 forms multimers, which differ from those formed by classical pentraxins in terms of protomer composition and requirement for disulfide bonds, and does not recognize CRP/SAP ligands. The capacity to bind C1q, mediated by the pentraxin domain, is consistent with the view that PTX3, produced in tissues by endothelial cells or macrophages in response to interleukin-1 and tumor necrosis factor, may act as a local regulator of innate immunity.

Ciceri: MolMed SpA: Consultancy.

BACKGROUND:

No relevant conflicts of interest to declare.

Although several studies have shown the capacity of mesenchymal stromal cells (MSCs) to repair and regenerate different tissues, the mechanisms underlying these processes are not understood. Long Pentraxin 3 (PTX3) is a multifunctional protein produced by MSCs and other cell subsets upon activation with inflammatory cytokines. PTX3 is involved in innate immunity, inflammation and extracellular matrix deposition. In the present study we analyzed the potential role of PTX3 in wound repair process induced by MSCs.

Innate immunity represents the first line of defence against pathogens and plays key roles in activation and orientation of the adaptive immune response. The innate immune system comprises both a cellular and a humoral arm. Components of the humoral arm include soluble pattern recognition molecules (PRMs) that recognise pathogen-associated molecular patterns (PAMPs) and initiate the immune response in coordination with the cellular arm, therefore acting as functional ancestors of antibodies. The long pentraxin PTX3 is a prototypic soluble PRM that is produced at sites of infection and inflammation by both somatic and immune cells. Gene targeting of this evolutionarily conserved protein has revealed a nonredundant role in resistance to selected pathogens. Moreover, PTX3 exerts important functions at the cross-road between innate immunity, inflammation, and female fertility. Here, we review the studies on PTX3, with emphasis on pathogen recognition and cross-talk with other components of the innate immune system.

Allogeneic haemopoietic-stem-cell transplantation (HSCT) is the treatment of choice for many malignant and non-malignant disorders. Despite the recent advances in post-transplantat immunosuppressive therapy, Graft-versus-Host Disease (GVHD) still represents the major life-threatening complication, developing in a substantial number of HSCT patients and resulting in poor outcome. The basis of GVHD pathophysiology are still poorly understood and its current diagnosis depends mainly on clinical manifestations and invasive biopsies. Specific biomarkers for GVHD would facilitate the early and accurate recognition of this invalidating disease as well as the monitoring of patient response to adopted anti-GVHD pharmacological treatment. With the aim to explore new reliable markers for predicting and monitoring GVHD course, we focused on pentraxin-3 (PTX-3), an acute-phase protein, that has been shown to play a crucial role in orchestrating inflammatory immune responses.Having obtained an informed consent, we collected plasma samples from 46 patients who received unmanipulated HSCT and from 9 healthy donors (HD) volunteers. After HSCT, 25/46 patients developed skin GVHD (18 acute GVHD and 7 chronic GVHD), while 21/46 never experienced it. Concerning GVHD patients, blood samples were collected at the day of GVHD onset/ flare, before the beginning of GVHD-specific drug therapy. PTX-3 plasma levels were monitored by ELISA assays.Patients who did not develop GVHD after HSCT showed augmented PTX-3 plasma levels (mean=3.3 ng/ml, range=1.1-8.6 ng/ml) if compared to HD (mean=1.2 ng/ml, range=0.3-2.5 ng/ml, p<0.01). Interestingly, we observed a strong increase of PTX-3 plasma levels in patients with acute GVHD (mean=42.2 ng/ml, range=6.7-218.2 ng/ml) or with flair-ups of chronic GVHD (mean=15.8 ng/ml, range=9-44.3 ng/ml). The increase of PTX-3 levels in patients with acute and active chronic GVHD was statistically significant (p<0.01 and p<0.05 respectively) when compared to both HD and HSCT patients without GVHD.These preliminary results suggest that PTX-3 plasma levels increase very rapidly in patients experiencing active GVHD, thus candidating PTX-3 as an easily measurable soluble factor useful to corroborate clinical observations in a disease in which signs and symptoms are often protean. Further studies are needed to clarify if PTX-3 could represent a good diagnostic and/or prognostic factor rapidly indicating therapy responsiveness.No relevant conflicts of interest to declare.

The role of inflammation in the pathogenesis of the athero-thrombosis has become well established over the past decade. In general population, markers of inflammation are associated with cardiovascular events and predictors of future thrombosis are leukocyte (WBC) count and short and long pentraxins such as high-sensitivity C-reactive protein (hs-CRP) and pentraxin 3 (PTX3). In Polycythemia Vera (PV) and Essential Thrombocythemia (ET), WBC count has been related to major vascular complications and there is evidence that this association can be mediated by JAK2V617F mutation. We tested the hypothesis that plasma levels of hs-CRP and PTX3 can be increased in patients with ET and PV following the permanent and constitutive inflammatory state as revealed by leukocytosys and leukocyte activation. This would further support the idea that an underpinning latent chronic inflammation may contribute to the thrombotic propensity of these clonal myeloproliferative disorders.Plasma level concentrations of hs-CRP and PTX3 were tested in the same samples of 48 PV and 52 ET patients (WHO 2008 diagnostic criteria) obtained at different times from diagnosis when patients were receiving aspirin alone (21% of ET and 37% of PV) or cytoreduction in 33% (hydroxyurea in 28%) alone or combined with aspirin in 38% of ET and 27% of PV. Twenty-two percent of ET and PV patients remained untreated and were submitted to phlebotomy according to hematocrit levels. Plasma levels of hs-CRP were measured by a commercially available immunoturbidimetric method (ADVIA 2400 Chemistry CardioPhaseTM, Siemens Healthcare Diagnostics Inc., Tarrytown, NY, US) while PTX3 was measured with a sandwich ELISA based on a monoclonal antibody (mAb) MNB4 (ascites diluted 1:5000 in coating buffer) and rabbit antiserum (Peri et al.: Circulation 2000, Latini et al.: Circulation 2004). The median age of ET and PV patients was 61 years (range: 32-87) and 62 years (range: 33-87), respectively. In 32 normal healthy controls median concentration of hs-CRP was 0.57 mg/L (range: 0.1-2.86). In 52 ET cases median hs-CRP values were not statistically different (median: 0.83 mg/L; range: 0.26-57.69, p=0.43). On the contrary, the 48 PV patients showed a significant increment (p=0.012) of hs-CRP concentration (median 1.31 mg/L; range: 0.15-53.36). The same results were found for PTX3: ET patients exhibited plasma values (median: 1.91; range: 0.8-7.75) not different (p=0.16) from controls (median: 1.74; range: 0.45-2.9) while in PV (median: 2.74; range: 0.33-10.83) the difference was statistically significant (p=0.012). Overall, hs-CRP and PTX3 values above the normal range were found in around a quarter of PV and ET patients. The distribution of clinical and hematological variables by hs-CRP plasma value tertiles (cut-off: <0.65; 0.65-1.88; >1.88) showed a significant correlation with JAK2V617F granulocyte allele burden (p= 0.004) while this correlation was weaker for PTX3 (p= 0.05). Similarly, lower, middle and upper tertiles for PTX3 were calculated (cut-off: <1.68; 16-.68-3.04; >3.04, respectively) and, in contrast to hs-CRP, the highest PTX3 tertile was associated with older age (p=0.001) and the PV phenotype (p=0.02). Both in ET and PV the other variables, including sex, hemoglobin, hematocrit, white blood cell and platelet count as well as treatments, did not show different distribution by hs-CRP and PTX3 tertiles. Vascular complications (32 episodes, calculated in the history, at diagnosis and in follow-up) were equally registered in the tertiles both in ET and PV.The present study is the first to show that both in ET and PV, blood hs-CRP and PTX3 are significantly increased mainly in relation to JAK2V617F mutation and age, and further supports the idea that in these disorders an underpinning latent chronic inflammation is demonstrable. Therefore, it is conceivable that these inflammatory pentraxins may contribute to vascular complications of these clonal disorders. Nevertheless, this preliminary information needs to be validated in a larger study.No relevant conflicts of interest to declare.

The need for antifungal prophylaxis in acute leukemia (AL) is a very debated topic, considering its potential of inducing resistant strains and of impairing detection of Aspergillus antigen as well as cost-efficacy balance. Targeted prophylaxis, focused on patients (pts) with high risk diseases (e.g. AL during induction therapy), seems to be the most promising strategy. Caspofungin is a well-tolerated echinocandin, with a different mechanism of action with respect to azoles and amphotericin B. Pentraxin 3 (PTX3) belongs to the superfamily of pentraxins acute-phase reactants and may play a protective role against Aspergillus spp (Garlanda et al, Nature 2002).To determine the incidence of invasive fungal infections (IFIs) and invasive aspergillosis (IA), the efficacy and safety of caspofungin as primary antifungal prophylaxis during AL induction and to evaluate PTX3 levels as in vivo predictive tool for IFI development.From Jan-07 to Jan-09, the incidence of IFIs and IA during induction was evaluated among all pts with acute lymphoblastic leukemia (ALL) and acute myeloblastic leukemia (AML) registered in NILG (Northern Italy Leukemia Group) protocols. Ninety-three pts entered the prospective ProfilC protocol and received Caspofungin prophylaxis (70 mg i.v. d1, then 50 mg i.v./d till leukemia remission). ProfilC epidemiological data were compared with those observed among 82 pts not assigned to ProfilC and treated according to the standard prophylactic (SP) policy adopted by each center (67 itraconazole, 10 fluconazole, 1 posaconazole, 4 no prophylaxis). PTX3 blood levels could be evaluated in 90 pts.Considering all 175 pts, M/F ratio was 103/72, median age was 50 (range 18-73); AML/ALL ratio was 138/37. Overall 32 IFIs (18.3%) were observed, 10 probable/proven (5.7%) (8 IA, 1 Candida spp and 1 G. capitatum bacteremias) and 22 possible (12.6%) according to EORTC/MSG criteria. IFIs (probable/proven and possible) were more frequent in AML (6.5% and 14.5%) than ALL (2.7% and 5.4%). Probable/proven IAs were not seen in ALL. The incidence of probable/proven or possible IFIs between ProfilC pts and those treated with SP was 7.5% and 9.7% vs 3.7% and 15.9%, respectively. Specifically, probable/proven or possible IAs were 5.4% and 8.6% in ProfilC group and 3.7% and 14.6% in SP group, respectively. These data show a lower, albeit not statistically significantly, incidence of IFI and IA in ProfilC pts. Fifteen out of the 175 (8.6%) pts died (ProfilC: 9.7%, SP: 7.3%, p=ns) with only one death due to IFI (G. capitatum sepsis). None of the 8 pts with IA died. Five pts experienced WHO grade >2 toxicity (3 hepatic, 1 skin, 1 hepatic and renal), 3 receiving Caspofungin and 2 itraconazole. One of the 5 pts died (itraconazole) of hepato-renal failure. A trend towards an association between lower PTX3 levels and IFI development was shown, since 13.8% of pts with PTX3 levels >10 ng/ml had IFI (probable: 3.4%), compared to 27.8% of those with '10 ng/ml (probable 8.2%).Anti-Aspergillus oriented prophylaxis in AL pts was standard policy at most NILG centers. The incidence and letality of probable/proven IFIs and IA were lower than expected and reported by others. Given this epidemiological scenario, the superiority of one drug with respect to others is difficult to demonstrate. Caspofungin was well tolerated during induction and was at least as effective as other prophylactic agents. It may be particularly suited for AL pts with impaired intestinal absorption. High levels of PTX3 (>10 ng/ml) could be protective against IFI/IA, suggesting that PTX3 could be used to identify a very high risk pts population in which an intensified strategy of prophylaxis may be worth testing.Off Label Use: Caspofungin as antifungal prophylaxis. Rossi:Merck Sharp & Dohme: Membership on an entity's Board of Directors or advisory committees.

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