Nadia Polentarutti

Role of IL-6 and its soluble receptor in induction of chemokines and leukocyte recruitment.

IL-6-/- mice showed impaired leukocyte accumulation in subcutaneous air pouches. Defective leukocyte accumulation was not due to a reduced migratory capacity of IL-6-/- leukocytes and was associated with a reduced in situ production of chemokines. These observations led to a reexamination of the interaction of IL-6 with endothelial cells (EC). EC express only the gp130 signal transducing chain and not the subunit-specific IL-6R and are therefore unresponsive to IL-6. However, EC are responsive to a combination of IL-6 and soluble IL-6R as measured by the activation of STAT3, chemokine expression, and augmentation of ICAM-1. Activation by IL-6-IL-6R complexes was inhibited by an IL-6 receptor antagonist and potentiated by a superagonist. Hence, in vivo and in vitro evidence supports the concept that the IL-6 system plays an unexpected positive role in local inflammatory reactions by amplifying leukocyte recruitment.

Autocrine Production of IL-10 Mediates Defective IL-12 Production and NF-κB Activation in Tumor-Associated Macrophages

IL-12 is a central cytokine in the activation of inflammation and immunity and in the generation of Th1-type responses. Tumor-associated macrophages (TAM) from mouse and human tumors showed defective production of IL-12. Defective IL-12 production was associated with lack of p50/p65 NF-κB activation. TAM produced increased amounts of the immunosuppressive cytokine IL-10. Abs against IL-10 restored the defective capacity of TAM to produce IL-12. Our data suggest that during tumor growth an IL-10-dependent pathway of diversion of macrophage function can be activated into the tumor microenvironment and results in the promotion of the IL-10+ IL-12− phenotype of TAM. Blocking IL-10, as well as other immunosuppressive cytokines present in the tumor microenvironment, such as TGF-β, may complement therapeutic strategies aimed at activating type I antitumor immune responses.

Inhibition of monocyte chemotactic protein-1 synthesis by statins

The beneficial effects of statins on the reduction of cardiovascular events has been partly attributed to their anti-inflammatory properties. In the complex of the different pathogenetic events leading to atherosclerosis, recent data suggest a central role of monocyte chemotactic protein-1 (MCP-1), because mice knock-out for MCP-1 or its receptor CC-chemokine receptor 2 were considerably resistant to plaque formation. In this study we investigated the effect of different statins on in vitro and in vivo production of MCP-1. Lovastatin and simvastatin caused a dose-dependent inhibition of MCP-1 production in peripheral blood mononuclear cells exposed to lipopolysaccharide or inactivated Streptococcus hemoliticus and in human endothelial cells exposed to interleukin-1β. The addition of mevalonate overrode the inhibitory effect of statins indicating that mevalonate-derived products are important for chemokine production. The in vivo anti-inflammatory effect of statins was investigated using the mouse air-pouch model of local inflammation. Lovastatin and pravastatin were orally administered to mice according to a treatment schedule that significantly inhibited the hepatic 3-hydroxy-3-methylglutaryl coenzyme A reductase activity without affecting total blood cholesterol. At the dose of 10 mg/kg, lovastatin and pravastatin reduced by approximately 50% the lipopolysaccharide-induced leukocytes recruitment and the exudate MCP-1 production. In conclusion, statins, by inhibiting mevalonate-derived products, reduced both in vitro and in vivo the production of chemokines involved in leukocyte migration, and this effect is unrelated to their cholesterol-lowering action.

Toll-like receptors: a growing family of immune receptors that are differentially expressed and regulated by different leukocytes.

Toll is a Drosophila gene essential for ontogenesis and antimicrobial resistance. Several hortologues of Toll have been identified and cloned in vertebrates, namely Toll‐like receptors (TLR). Human TLR are a growing family of molecules involved in innate immunity. TLR are structurally characterized by a cytoplasmic Toll/interleukin‐1R (TIR) domain and by extracellular leucine‐rich repeats. TLR characterized so far activate the My D88/IRAK signaling cascade, which bifurcates and leads to NF‐κB and c‐Jun/ATF2/TCF activation. Genetic, gene transfer, and dominant‐negative approaches have involved TLR family members (TLR2 and TLR4) in lipopolysaccharide recognition and signaling. Accumulating evidence suggests that some TLR molecules are also involved in signaling receptor complexes that recognize components of gram‐positive bacteria and mycobacteria. However, the definitive role of other TLR is still lacking. A systematic approach has been used to determine whether different human leukocyte populations selectively or specifically expressed TLR mRNA. Based on expression pattern, TLR can be classified as ubiquitous (TLR1), restricted (TLR2, TLR4, and TLR5), and specific (TLR3). Expression and regulation of distinct though overlapping ligand recognition patterns may underlie the existence of a numerous, seemingly redundant, TLR family. Alternately, the expression of a TLR in a single cell type may indicate a specific role for this molecule in a restricted setting. J. Leukoc. Biol. 67: 450–456; 2000.

Increased Susceptibility to Colitis-Associated Cancer of Mice Lacking TIR8, an Inhibitory Member of the Interleukin-1 Receptor Family

TIR8 (also known as SIGIRR) is a member of the interleukin-1/Toll-like receptor family with inhibitory activity on inflammatory reactions and high expression in intestinal mucosa. Here, we report that Tir8-deficient mice exhibited a dramatic intestinal inflammation in response to dextran sulfate sodium salt (DSS) administration in terms of weight loss, intestinal bleeding, and mortality and showed increased susceptibility to carcinogenesis in response to azoxymethane and DSS. Increased susceptibility to colitis-associated cancer was associated to increased permeability and local production of prostaglandin E(2), proinflammatory cytokines, and chemokines. Thus, these results are consistent with the hypothesis that TIR8, by negatively regulating intestinal inflammation, plays a nonredundant role in the control of the protumor activity of chronic inflammation in the gut.

Regulation of PTX3, a key component of humoral innate immunity in human dendritic cells: stimulation by IL-10 and inhibition by IFN-γ

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 Pam3 Cys‐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.

MCP.-1 and CCR2 in HIV infection: regulation of agonist and receptor expression

Monocyte chemotactic protein‐1 (MCP‐1) interacts with the chemokine receptor CCR2. Two CCR2 cDNAs have been described. Sequence analysis as well as Northern blotting and RNase protection with different probes revealed that the CCR2 gene is expressed in activated natural killer (NK) cells and mononuclear phagocytes as a predominant long transcript (3.4 kb) consisting of CCR2B followed by a novel sequence (X), corresponding to an intron in the genome, and by a CCR2A specific portion. The predominant long transcript is polyadenylated and present in the cytoplasm. We found that bacterial products and cytokines affect CCR2 expression. Interleukin‐2 (IL‐2) augmented CCR2 mRNA in monocytes and NK cells. The augmented migratory capacity of Reactivated versus resting NK cells was associated with increased CCR2 transcript levels. Lipopolysaccharide (LPS) and other microbial agents caused a rapid and drastic reduction of CCR2 mRNA levels. The rate of nuclear transcription of CCR2 was not affected by LPS, whereas the mRNA half life was reduced. These results suggest that regulation of receptor expression, in addition to agonist production, is probably a crucial point in the regulation of the chemokine system. Down‐regulation of chemokine receptor expression may play a role in the modulation of HIV infection in macrophages by LPS. Levels of MCP‐1 were markedly elevated in the cerebrospinal fluid (CSF) but not in blood of HIV‐infected patients with cytomegalovirus (CMV) encephalitis. The CSF levels of MCP‐1 in CMV encephalitis were markedly higher than those found in the CSF of HIV‐infected patients with or without unrelated neurological diseases. IL‐8, the prototype of C‐X‐C chemokines and RANTES and macrophage inflammatory protein‐1α (C‐C chemokines) were not substantially increased in the liquor of CMV encephalitis patients. High levels of MCP‐1 may underlie monocyte recruitment and tissue damage in CMV encephalitis and may represent a rapid and useful tool in the diagnostic armamentarium for neurological disorders associated with HIV. J. Leukoc. Biol. 62: 30–33; 1997.

Extracellular and intracellular decoys in the tuning of inflammatory cytokines and Toll‐like receptors: the new entry TIR8/SIGIRR

Following the identification of the interleukin (IL)‐1 type II receptor as a prototypic decoy receptor, nonsignaling receptors with decoy functions have been identified for members of the IL‐1/IL‐18,tumor necrosis factor, IL‐10, and IL‐13 receptor families. Moreover, the silent receptor D6 is a promiscuous decoy and scavenger receptor of inflammatory chemokines. The type II IL‐1 decoy receptor also acts as a dominant‐negative molecule. Intracellular pathways of inhibition of IL‐1 and Toll‐like receptor (TLR) signaling have been identified. In particular, recent results suggest that the Toll/IL‐1 receptor (TIR) family member TIR8, also known as single immunoglobulin IL‐1‐related receptor (SIGIRR), is a negative regulator of IL‐1 and TLR signaling. Thus, extracellular and intracellular decoys tune the activation of members of the IL‐1/TLR receptor family.

Inducible expression of the long pentraxin PTX3 in the central nervous system

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. PTX3 is induced by primary proinflammatory signals in various cell types, most prominently macrophages and endothelial cells. Other long pentraxins, such as murine or rat neuronal pentraxin 1 (NP1) and human neuronal pentraxin 2 (NPTX2), are expressed in the central nervous system (CNS). The present study was designed to investigate whether PTX3 is expressed in the brain and to define the structures and cells involved. Intracerebroventricular (i.c.v.), but not i.v., injection of LPS induced high levels of PTX3 mRNA in the mouse brain. In contrast NP1 is constitutively expressed in the murine CNS and is not modulated by LPS administration. I.c.v. IL-1beta was also a potent inducer of PTX3 expression in the CNS, whereas TNFalpha was substantially less effective and IL-6 induced a barely detectable signal. Central administration of LPS and IL-1 induced PTX3 also in the periphery (heart), whereas the reverse did not occur. Expression of PTX3 was also observed in the brain of mice infected with Candida albicans (C. albicans) or Cryptococcus neoformans. (C. neoformans). The kinetics of PTX3 gene induction were consistently different between C. albicans- and C. neoformans-infected mice, according to the diverse outcome of the CNS immune reaction. In situ hybridization revealed that i.c.v. injection of LPS induced a strong PTX3 expression in presumptive glial cells, in the white matter (corpus callosum, fimbria) and meningeal pia mater as well as in dentate gyrus hilus and granule cells. No constitutive expression of PTX3 was detected. Central expression of PTX3 may amplify mechanisms of innate resistance and damage in the CNS. The possibility of a direct interaction of PTX3 with neuronal cells, as suggested for NPTX2, remains to be explored.

The Therapeutic Potential of the Humoral Pattern Recognition Molecule PTX3 in Chronic Lung Infection Caused by Pseudomonas aeruginosa

Chronic lung infections by Pseudomonas aeruginosa strains are a major cause of morbidity and mortality in cystic fibrosis (CF) patients. Although there is no clear evidence for a primary defect in the immune system of CF patients, the host is generally unable to clear P. aeruginosa from the airways. PTX3 is a soluble pattern recognition receptor that plays nonredundant roles in the innate immune response to fungi, bacteria, and viruses. In particular, PTX3 deficiency is associated with increased susceptibility to P. aeruginosa lung infection. To address the potential therapeutic effect of PTX3 in P. aeruginosa lung infection, we established persistent and progressive infections in mice with the RP73 clinical strain RP73 isolated from a CF patient and treated them with recombinant human PTX3. The results indicated that PTX3 has a potential therapeutic effect in P. aeruginosa chronic lung infection by reducing lung colonization, proinflammatory cytokine levels (CXCL1, CXCL2, CCL2, and IL-1β), and leukocyte recruitment in the airways. In models of acute infections and in in vitro assays, the prophagocytic effect of PTX3 was maintained in C1q-deficient mice and was lost in C3- and Fc common γ-chain–deficient mice, suggesting that facilitated recognition and phagocytosis of pathogens through the interplay between complement and FcγRs are involved in the therapeutic effect mediated by PTX3. These data suggested that PTX3 is a potential therapeutic tool in chronic P. aeruginosa lung infections, such as those seen in CF patients.