Soluble Pattern Recognition Receptor Research Articles

A Sweet Response to a Sour Situation: The Role of Soluble Pattern Recognition Receptors in the Innate Immune Response to Invasive Aspergillus fumigatus Infections.

Aspergillus spp. infect around 11,000,000 patients, resulting in about 600,000 deaths per year, but these numbers are on the rise due to the emergence of antifungal-resistant strains and a lack of sensitive diagnostic tests [1]. It is increasingly acknowledged that soluble pattern recognition receptors (PRRs), such as the complement component C1q, the collectins (MBL, SP, and CL-11), PTX3, and the ficolins (ficolin-1, 2, 3 and A), are important within anti-Aspergillus immunity [2]. Moreover, studies have highlighted that they may be used as a possible alternative to current antifungal drugs or used in combination to increase efficacy [3]. Binding of pathogen-associated molecular patterns (PAMPs) on the pathogen surface by soluble PRRs often results in opsonisation. This enhances interactions with membrane-associated PRRs on phagocytes, such as the important β-glucan receptor Dectin-1, Toll-like receptors (TLRs), complement receptors (CR1), and Fc receptors; ultimately augmenting phagocytosis, which is essential in controlling the infection [2]. Alternatively, opsonins can promote fungal damage directly or further promote opsonisation by C3b deposition via activation of the conserved complement system [4]. There are three main arms of the complement system, which are the classical, alternative, and lectin pathways. C1q primarily activates the classical antibody-mediated pathway, whereas MBL, CL-11, and the ficolins are known to activate the lectin complement pathway via activation of the mannose-binding lectin-associated serine proteases (MASPs). However, SP-A and SP-D are not involved in complement activation, and the role of CL-11 in Aspergillus immunity is yet to be explored. Furthermore, PTX3 can interact with complement activators and inhibitory components to modulate all three pathways [5]. The role of each of these PRRs in anti-Aspergillus immunity will be discussed further.

High glucose concentration-induced expression of pentraxin-3 in a rat model of continuous peritoneal dialysis.

Continuous exposure to peritoneal dialysis fluids (PDFs) is associated with pathological responses such as persistent micro-inflammation, which leads to ultrafiltration failure. Pentraxin-3 (PTX3), a multifunctional soluble pattern recognition receptor, is produced at sites of inflammation by a wide range of cell types. This study investigates the in vivo expression of PTX3 in the peritoneal membrane of a rat continuous peritoneal dialysis (PD) model, as well as the effect of high glucose on the in vitro expression of PTX3. The expression of PTX3 was analyzed using RT-PCR, real-time PCR, immunohistochemistry and western blotting in a PD rat model receiving saline or conventional PDF containing 3.86% glucose for 8 weeks. The effects of high glucose on the expression of PTX3 were examined in cultured rat peritoneal mesothelial cells (RPMCs), mouse macrophage-like cells, and mouse fibroblasts. In a rat model of PD, eight-week instillation of the conventional PDF produced increased submesothelial thickening, followed by substantially enhanced PTX3 protein levels in the submesothelial layer of peritoneal membrane. PTX3 was detected in peritoneal mesothelial cells, macrophages and fibroblasts in the thickened submesothelial area. Glucose was found to induce PTX3 protein expression in RPMCs as well as macrophage-like cells and fibroblasts. Continuous exposure to conventional PDF induces PTX3 expression in the peritoneal membrane of rats. High glucose may be involved in the mechanism of PDF-induced local micro-inflammation in the peritoneum.

In silico Analysis Revealed High-risk Single Nucleotide Polymorphisms in Human Pentraxin-3 Gene and their Impact on Innate Immune Response against Microbial Pathogens.

Pentraxin-3 (PTX-3) protein is an evolutionary conserved protein that acts as a soluble pattern-recognition receptor for pathogens and plays important role in innate immune response. It recognizes various pathogens by interacting with extracellular moieties such as glactomannan of conidia (Aspergillus fumigatus), lipopolysaccharide of Pseudomonas aeruginosa, Streptococcus pneumonia and Salmonella typhimurium. Thus, PTX-3 protein helps to clear these pathogens by activating downstream innate immune process. In this study, computational methods were used to analyze various non-synonymous single nucleotide polymorphisms (nsSNPs) in PTX-3 gene. Three different databases were used to retrieve SNP data sets followed by seven different in silico algorithms to screen nsSNPs in PTX-3 gene. Sequence homology based approach was used to identify nsSNPs. Conservation profile of PTX-3 protein amino acid residues were predicted by ConSurf web server. In total, 10 high-risk nsSNPs were identified in pentraxin-domain of PTX-3 gene. Out of these 10 high-risk nsSNPs, 4 were present in the conserved structural and functional residues of the pentraxin-domain, hence, selected for structural analyses. The results showed alteration in the putative structure of pentraxin-domain. Prediction of protein–protein interactions analysis showed association of PTX-3 protein with C1q component of complement pathway. Different functional and structural residues along with various putative phosphorylation sites and evolutionary relationship were also predicted for PTX-3 protein. This is the first extensive computational analyses of pentraxin protein family with nsSNPs and will serve as a valuable resource for future population based studies.

Inflammation and its role in age-related macular degeneration.

Inflammation is a cellular response to factors that challenge the homeostasis of cells and tissues. Cell-associated and soluble pattern-recognition receptors, e.g. Toll-like receptors, inflammasome receptors, and complement components initiate complex cellular cascades by recognizing or sensing different pathogen and damage-associated molecular patterns, respectively. Cytokines and chemokines represent alarm messages for leukocytes and once activated, these cells travel long distances to targeted inflamed tissues. Although it is a crucial survival mechanism, prolonged inflammation is detrimental and participates in numerous chronic age-related diseases. This article will review the onset of inflammation and link its functions to the pathogenesis of age-related macular degeneration (AMD), which is the leading cause of severe vision loss in aged individuals in the developed countries. In this progressive disease, degeneration of the retinal pigment epithelium (RPE) results in the death of photoreceptors, leading to a loss of central vision. The RPE is prone to oxidative stress, a factor that together with deteriorating functionality, e.g. decreased intracellular recycling and degradation due to attenuated heterophagy/autophagy, induces inflammation. In the early phases, accumulation of intracellular lipofuscin in the RPE and extracellular drusen between RPE cells and Bruch’s membrane can be clinically detected. Subsequently, in dry (atrophic) AMD there is geographic atrophy with discrete areas of RPE loss whereas in the wet (exudative) form there is neovascularization penetrating from the choroid to retinal layers. Elevations in levels of local and systemic biomarkers indicate that chronic inflammation is involved in the pathogenesis of both disease forms.

Oxidation-specific epitopes are major targets of innate immunity in atherothrombosis.

Atherosclerosis is a chronic inflammatory disease of the vascular wall that results from disturbed lipoprotein metabolism and increased oxidative stress. A major consequence of this is lipid peroxidation, which generates a number of breakdown products of membrane lipids that form so called oxidation-specific epitopes (OSE). OSE have been documented in oxidized lipoproteins and on the surface of dying cells and circulating microparticles, and their ability to trigger robust pro-inflammatory and pro-thrombotic responses has been demonstrated extensively. Recent studies have identified specific OSE as major targets of both cellular and soluble pattern recognition receptors of the innate immune system, including innate natural IgM antibodies. This allows the immune system to identify metabolic waste and mediate important physiological house keeping functions, e.g. by promoting the removal of cellular debris and by neutralizing oxidized molecules. Indeed, innate B1 cells and B1 cell derived natural IgM with specificity for OSE have been shown to protect mice from the development of atherosclerotic lesions. Moreover, OSE-specific natural IgM antibodies bind and neutralize the pro-inflammatory and pro-thrombotic effects of OSE, and low levels of OSE-specific IgM are associated with an increased risk for myocardial infarction. Understanding the molecular components and mechanisms involved in this process, will help identify individuals with increased risk for atherothrombosis and indicate novel points for therapeutic intervention.

The clearance of cell remnants and the regeneration of the injured muscle depend on soluble pattern recognition receptor PTX3.

The signals causing the resolution of muscle inflammation are only partially characterized. The long pentraxin PTX3, which modulates leukocyte recruitment and activation, could contribute. We analysed the expression of ptx3 after muscle injury and verified whether hematopoietic precursors are a source of the protein. The kinetics of regeneration and leukocytes infiltration, the accumulation of cell remnants and anti-histidyl-t-RNA synthetase autoantibodies were compared in wild-type and ptx3-deficient mice. Ptx3 expression was up-regulated three-five days after injury and restricted to the extracellular matrix. Cellular debris and leukocytes persisted in the muscle of ptx3-deficient mice for a long time after wild-type animals had healed. ptx3-deficient macrophages expressed receptors involved in apoptotic cell clearance and engulfed dead cells in vitro. Accumulation of cell debris in a pro-inflammatory microenvironment was not sufficient to elicit autoantibodies. PTX3 generated in response to muscle injury prompts the clearance of debris and the termination of the inflammatory response.

Ficolins Promote Fungal Clearance in vivo and Modulate the Inflammatory Cytokine Response in Host Defense against Aspergillus fumigatus

Aspergillus fumigatus is an opportunistic fungal pathogen that causes severe invasive infections in immunocompromised patients. Innate immunity plays a major role in protection against A. fumigatus. The ficolins are a family of soluble pattern recognition receptors that are capable of activating the lectin pathway of complement. Previous in vitro studies reported that ficolins bind to A. fumigatus, but their part in host defense against fungal infections in vivo is unknown. In this study, we used ficolin-deficient mice to investigate the role of ficolins during lung infection with A. fumigatus. Ficolin knockout mice showed significantly higher fungal loads in the lungs 24 h postinfection compared to wild-type mice. The delayed clearance of A. fumigatus in ficolin knockout mice could not be attributed to a compromised recruitment of inflammatory cells. However, it was revealed that ficolin knockout mice exhibited a decreased production of proinflammatory cytokines in the lungs compared to wild-type mice following A. fumigatus infection. The impaired clearance and cytokine production in ficolin knockout mice was independent of complement, as shown by equivalent levels of A. fumigatus-mediated complement activation in ficolin knockout mice and wild-type mice. In conclusion, this study demonstrates that ficolins are important in initial innate host defense against A. fumigatus infections in vivo.

Crucial role of the interplay of the MAPKAP kinases 2 and 3 for LPS-induced inflammation and their relevance for liver pathogenesis

The liver plays a pivotal role in innate and adaptive immunity as it is frequently exposed to pathogens reaching the liver via the blood flow. It is important for the production of acute phase proteins, among others soluble pattern-recognition receptors and components of the complement system, which are important constituents of innate immunity. Moreover, the liver harbors the largest pool of resident tissue macrophages. They play a key role for the elimination of opsonized immune complexes by phagocytosis and they are important sources of inflammatory cytokines, which induce acute phase protein production in hepatocytes and further trigger innate immunity and subsequent formation of adaptive immunity.

TNF up-regulates Pentraxin3 expression in human airway smooth muscle cells via JNK and ERK1/2 MAPK pathways.

BackgroundLong pentraxin 3 (PTX3) is a novel candidate marker for inflammation in many chronic diseases. As a soluble pattern recognition receptor, PTX3 is involved in amplification of inflammatory reactions and regulation of innate immunity. Previously, we demonstrate that human airway smooth muscle cells (HASMC) express constitutively PTX3 and upon TNF stimulation. However, very little is known about the mechanism governing its expression in HASMC. We sought to investigate the mechanism governing TNF induced PTX3 expression in primary HASMC.MethodsHASMC were stimulated with TNF in the presence of transcriptional inhibitor actinomycin D (ActD) or MAPKs pharmacological inhibitors. PTX3 mRNA and protein expression were analyzed by Real-time RT-PCR and ELISA, respectively. PTX3 promoter activity was determined using luciferase assay.ResultsPTX3 mRNA and protein are expressed constitutively by HASMC and significantly up-regulated by TNF. TNF-induced PTX3 mRNA and protein release in HASMC were inhibited by transcriptional inhibitor actinomycin D. TNF induced significantly PTX3 promoter activation in HASMC. MAPK JNK and ERK1/2 specific inhibitors (SP600125 and UO126), but not p38, significantly down regulates TNF induced PTX3 promoter activity and protein release in HASMC. Finally, TNF mediated PTX3 promoter activity in HASMC was abolished upon mutation of NF-κβ and AP1 binding sites.ConclusionsOur data suggest that TNF induced PTX3 in HASMC at least via a transcriptional mechanism that involved MAPK (JNK and ERK1/2), NF-κβ and AP1 pathways. These results rise the possibility that HASMC derived PTX3 may participate in immune regulation in the airways.

Antibacterial effect of porcine PTX3 against Streptococcus suis type 2 infection

Pentraxin 3 (PTX3), a soluble pattern recognition receptor, plays an important role in innate immunity and has been implicated to be a candidate resistance gene against Streptococcus suis 2 infection. To discover the antibacterial effect of porcine PTX3 against S. suis 2, the 42-kDa PTX3 protein was expressed by Chinese hamster ovary cells (CHO), and an additional eukaryotic expression vector pVAX-ptx3 was constructed. The expressed porcine PTX3 mediated a range of antibacterial activities including increasing phagocytic capacity of primary porcine alveolar macrophages (PAM) against S. suis 2 and inhibiting adhesion of S. suis 2 to human epidermoid cancer cells (Hep-2). In mouse model, pre-intramuscular injecting with pVAX-ptx3 reduced mortality and reduced bacteria loads in blood, spleen, lung and brain compared with that of control group during 2–12 h following intraperitoneal injection (i.p.) with S. suis 2. Meanwhile, the expressions of IL-6 and IL-8 in blood were increased in pVAX-ptx3 group, whereas no obvious changes about IL-10. In piglet model, bacteria load in blood of pVAX-ptx3 group was significantly lower than that of control group after i.p. with S. suis 2, correspondingly, expression of IL-6 and IL-8 were significantly increased in pVAX-ptx3 group. In contrast, white blood cell (WBC) and neutrophil cell (NEU) count of peripheral blood in pVAX-ptx3 group were lower than that of control group. These studies described a novel antibacterial role for porcine PTX3 against S. suis 2 both in vitro and in vivo and suggested that porcine PTX3 may be a potential biological agent against S. suis 2 in pig and be used for the clinical prevention and treatment of streptococcosis caused by S. suis 2.

Genetic variation in PTX3 and plasma levels associated with hepatocellular carcinoma in patients with HCV.

Hepatitis C virus (HCV) is the main cause of chronic liver disease, cirrhosis and hepatocellular carcinoma (HCC) worldwide. The risk to develop HCC increases with the severity of liver inflammation and fibrosis. Long pentraxin 3 (PTX3) is a soluble pattern-recognition receptor produced by phagocytes and nonimmune cells at sites of inflammation or injury. The aim of the present study was to determine the association of PTX3 polymorphisms and its plasma levels with HCC occurrence among patients with HCV. Samples from 524 patients with chronic hepatitis C were evaluated in this study. Two polymorphisms (rs1840680 and rs2305619) in the PTX3 gene were determined by real-time PCR. PTX3 plasma levels were measured by Enzyme-linked Immunosorbent Assay (ELISA). Our data show a significant association between PTX3 polymorphisms and HCC occurrence in univariate and multivariate analysis (P = 0.024). Patients with HCC had higher PTX3 plasma levels compared to individuals with mild or severe fibrosis (P < 0.0001 and P = 0.002, respectively). In addition, PTX3 rs2305619 polymorphism and plasma levels were correlated with Child-Pugh scores B and C in HCC individuals. PTX3 seems to be a risk factor for HCC occurrence in chronic hepatitis C. This is the first study that evaluates PTX3 in the context of hepatitis C.

Targeting C-Type Lectin Receptors for Cancer Immunity.

C-type lectin receptors (CLRs) are a large family of soluble and trans-membrane pattern recognition receptors that are widely and primarily expressed on myeloid cells. CLRs are important for cell–cell communication and host defense against pathogens through the recognition of specific carbohydrate structures. Similar to a family of Toll-like receptors, CLRs signaling are involved in the various steps for initiation of innate immune responses and promote secretion of soluble factors such as cytokines and interferons. Moreover, CLRs contribute to endocytosis and antigen presentation, thereby fine-tune adaptive immune responses. In addition, there may also be a direct activation of acquired immunity. On the other hand, glycans, such as mannose structures, Lewis-type antigens, or GalNAc are components of tumor antigens and ligate CLRs, leading to immunoregulation. Therefore, agonists or antagonists of CLRs signaling are potential therapeutic reagents for cancer immunotherapy. We aim to overview the current knowledge of CLRs signaling and the application of their ligands on tumor-associating immune response.

Long-Pentraxin 3 Derivative as a Small-Molecule FGF Trap for Cancer Therapy

The fibroblast growth factor (FGF)/FGF receptor (FGFR) system plays a crucial role in cancer by affecting tumor growth, angiogenesis, drug resistance, and escape from anti-angiogenic anti-vascular endothelial growth factor therapy. The soluble pattern recognition receptor long-pentraxin 3 (PTX3) acts as a multi-FGF antagonist. Here we demonstrate that human PTX3 overexpression in transgenic mice driven by the Tie2 promoter inhibits tumor growth, angiogenesis, and metastasis in heterotopic, orthotopic, and autochthonous FGF-dependent tumor models. Using pharmacophore modeling of the interaction of a minimal PTX3-derived FGF-binding pentapeptide with FGF2, we identified a small-molecule chemical (NSC12) that acts as an extracellular FGF trap with significant implications in cancer therapy.

Interaction of human mannose-binding lectin (MBL) with Yersinia enterocolitica lipopolysaccharide

The lipopolysaccharide (LPS) is involved in the interaction between Gram-negative pathogenic bacteria and host. Mannose-binding lectin (MBL), complement-activating soluble pattern-recognition receptor targets microbial glycoconjugates, including LPS. We studied its interactions with a set of Yersinia enterocolitica O:3 LPS mutants. The wild-type strain LPS consists of lipid A (LA) substituted with an inner core oligosaccharide (IC) which in turn is substituted either with the O-specific polysaccharide (OPS) or the outer core hexasaccharide (OC), and sometimes also with the enterobacterial common antigen (ECA). The LPS mutants produced truncated LPS, missing OPS, OC or both, or, in addition, different IC constituents or ECA. MBL bound to LA-IC, LA-IC-OPS and LA-IC-ECA but not LA-IC-OC structures. Moreover, LA-IC substitution with both OPS and ECA prevented the lectin binding. Sequential truncation of the IC heptoses demonstrated that the MBL targets the IC heptose region. Furthermore, microbial growth temperature influenced MBL binding; binding was stronger to bacteria grown at room temperature (22°C) than to bacteria grown at 37°C. In conclusion, our results demonstrate that MBL can interact with Y. enterocolitica LPS, however, the in vivo significance of that interaction remains to be elucidated.

C-reactive protein directly suppresses Th1 cell differentiation and alleviates experimental autoimmune encephalomyelitis.

Human C-reactive protein (CRP) is a serum-soluble pattern recognition receptor that serves as a marker of inflammation and directly contributes to innate immunity. In this study, we show that human CRP also directly contributes to adaptive immunity, that is, native CRP binds specifically to human Jurkat T cells and to mouse naive CD4(+) T cells and modulates their Th1 and Th2 responses. In vitro both exogenously added (purified) and endogenously expressed (via transfection) human CRP inhibited Th1 differentiation and augmented Th2 differentiation of naive CD4(+) T cells. In vivo for human CRP transgenic compared with wild-type mice, a lesser proportion of the T cells recovered from the spleens of healthy animals were Th1 cells. Moreover, in both CRP transgenic mice and in wild-type mice treated with human CRP, during myelin oligodendrocyte glycoprotein peptide-induced experimental autoimmune encephalomyelitis both the Th1 cell response and disease severity were inhibited. These pattern recognition-independent actions of CRP directly on T cells highlights the potential for this soluble pattern recognition receptor to act as a tonic regulator of immunity, shaping global adaptive immune responses during both homeostasis and disease.

A long pentraxin-3-derived pentapeptide for the therapy of FGF8b-driven steroid hormone-regulated cancers.

Fibroblast growth factor-8b (FGF8b) affects the epithelial/stromal compartments of steroid hormone-regulated tumors by exerting an autocrine activity on cancer cells and a paracrine pro-angiogenic function, thus contributing to tumor progression. The FGF8b/FGF receptor (FGFR) system may therefore represent a target for the treatment of steroid hormone-regulated tumors. The soluble pattern recognition receptor long pentraxin-3 (PTX3) binds various FGFs, including FGF2 and FGF8b, thus inhibiting the angiogenic and tumorigenic activity of androgen-regulated tumor cells. Nevertheless, the complex/proteinaceous structure of PTX3 hampers its pharmacological exploitation. In this context, the acetylated pentapeptide Ac-ARPCA-NH2 (ARPCA), corresponding to the N-terminal amino acid sequence PTX3(100-104), was identified as a minimal FGF2-binding peptide able to antagonize the biological activity of FGF2. Here, we demonstrate that ARPCA binds FGF8b and inhibits its capacity to form FGFR1-mediated ternary complexes with heparan sulphate proteoglycans. As a FGF8b antagonist, ARPCA inhibits FGFR1 activation and signalling in endothelial cells, hampering the angiogenic activity exerted in vitro and in vivo by FGF8b. Also, ARPCA suppresses the angiogenic and tumorigenic potential of prototypic androgen/FGF8b-dependent Shionogi 115 mammary carcinoma cells and of androgen/FGF8b/FGF2-dependent TRAMP-C2 prostate cancer cells. In conclusion, ARPCA represents a novel FGF8b antagonist with translational implications for the therapy of steroid hormone-regulated tumors.

Human lectins and their roles in viral infections.

Innate recognition of virus proteins is an important component of the immune response to viral pathogens. A component of this immune recognition is the family of lectins; pattern recognition receptors (PRRs) that recognise viral pathogen-associated molecular patterns (PAMPs) including viral glycoproteins. In this review we discuss the contribution of soluble and membrane-associated PRRs to immunity against virus pathogens, and the potential role of these molecules in facilitating virus replication. These processes are illustrated with examples of viruses including human immunodeficiency virus (HIV), hepatitis C virus (HCV) and Ebola virus (EBOV). We focus on the structure, function and genetics of the well-characterised C-type lectin mannose-binding lectin, the ficolins, and the membrane-bound CD209 proteins expressed on dendritic cells. The potential for lectin-based antiviral therapies is also discussed.

MBL2 deficiency is associated with higher genomic bacterial loads during meningococcemia in young children

Mannose binding lectin (MBL2) is a soluble pattern recognition receptor that is key to generating innate immune responses to invasive infection, including against the cardinal Gram-negative bacterium Neisseria meningitidis. Individuals homozygous or heterozygous for any of three variant alleles of MBL2 (O/O or A/O genotypes) have deficient concentrations of MBL2 in circulating blood, but previous studies linking MBL deficiency to susceptibility to meningococcal disease have not revealed a consistent association. We genotyped 741 patients with microbiologically – proven meningococcal disease and correlated MBL2 genotype with plasma bacterial load of N. meningitidis with blood samples taken during hospital admission. We show that individuals with genotypes compatible with MBL2 deficiency have higher measurable levels of bacterial plasma genomic load with the greatest effect seen in children <2 years of age. However, the overall impact of this is minor, because there was no evidence that such genotypes are more common in children with meningococcal disease compared with uninfected cohorts. The findings suggest that MBL2 supports innate immune defence against meningococcal disease in the early months of life, before acquired immunity is sufficiently robust for effective natural protection.

Systemic pentraxin-3 levels reflect vascular enhancement and progression in Takayasu arteritis.

IntroductionProgression 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.MethodsA 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.ResultsPatients 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.ConclusionsArterial 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.

Abstract 178: Stromal expression of long Pentraxin-3 impairs tumor growth and metastasis

Abstract Long Pentraxin-3 (PTX3) is a soluble pattern recognition receptor expressed by endothelial and immune cells in inflammatory contexts. We have previously demonstrated that PTX3 binds to different members of the FGF family, thus inhibiting their biological activity. The FGF/FGFR system strongly contributes to cancer progression by inducing tumor growth and neovascularization. To date, recombinant PTX3 protein or PTX3-overexpressing tumor cell lines have been exploited to assess the antitumor effects of this natural FGF trap. Here we generated C57BL/6 transgenic mice expressing human (h)PTX3 under the control of the endothelial specific Tie2/Tek transcription regulatory sequences. These animals were used to investigate the impact of PTX3 overexpression by the host stroma on tumor growth, vascularization and metastasis. Transgenic Tie2-hPTX3 mice were generated by cloning the hPTX3 cDNA into the late-generation, self-inactivating lentiviral vector Tie2p/e to obtain the Tie2-hPTX3 lentiviral transfer vectors that were injected into fertilized oocytes. Expression of the transgene was confirmed by RT-PCR and western blot analyses of different organs from Tie2-hPTX3 mice that showed increased levels of circulating PTX3 (80-180 ng/ml) when compared to wild type (wt) animals (&amp;lt;1.8 ng/ml). Also, histological analysis confirmed the perivascular accumulation of hPTX3 in transgenic animals. To assess the anti-angiogenic activity of endothelium-derived hPTX3, we performed ex vivo aorta ring and in vivo matrigel plug assays. Both assays revealed a significant inhibition of FGF2-driven angiogenesis in Tie2-hPTX3 mice that maintained their responsiveness to VEGF-A, thus confirming the specificity of the effect. Next, different syngeneic FGF-dependent tumor cell lines, including TRAMP-C2 prostate carcinoma, B16-F10 melanoma and Lewis Lung carcinoma cells, were subcutaneously injected in Tie2-hPTX3 mice. Notably, the growth of all tumor grafts was significantly reduced in Tie2-hPTX3 mice when compared to wt animals. Also, histological analysis of TRAMP-C2 tumors grown in Tie2-hPTX3 mice showed a strong perivascular expression of hPTX3 and a significant reduction of FGFR1 phosphorylation. This was paralleled by a significant decrease of tumor vascularity and tumor cell proliferation whereas no difference in tumor growth was observed for TRAMP-C2 grafts expressing a constitutively activated form of FGFR1. Finally, B16-F10 melanoma and M5076 ovarian sarcoma cells showed a dramatic decrease of their capacity to form experimental metastases in the lung and liver, respectively, after intravenous injection in Tie2-hPTX3 mice. PTX3 is a natural FGF ligand trap. Our findings demonstrate for the first time that the production of PTX3 by the host stroma may exert a dramatic impact on tumor growth, vascularization and metastasis with potential exploitation for the therapy of FGF-dependent tumors. Citation Format: Arianna Giacomini, Emanuela Di Salle, Daniela Coltrini, Mirella Belleri, Marco Presta, Roberto Ronca. Stromal expression of long Pentraxin-3 impairs tumor growth and metastasis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 178. doi:10.1158/1538-7445.AM2014-178