Diana Boraschi

From Monocytes to M1/M2 Macrophages: Phenotypical vs. Functional Differentiation

Studies on monocyte and macrophage biology and differentiation have revealed the pleiotropic activities of these cells. Macrophages are tissue sentinels that maintain tissue integrity by eliminating/repairing damaged cells and matrices. In this M2-like mode, they can also promote tumor growth. Conversely, M1-like macrophages are key effector cells for the elimination of pathogens, virally infected, and cancer cells. Macrophage differentiation from monocytes occurs in the tissue in concomitance with the acquisition of a functional phenotype that depends on microenvironmental signals, thereby accounting for the many and apparently opposed macrophage functions. Many questions arise. When monocytes differentiate into macrophages in a tissue (concomitantly adopting a specific functional program, M1 or M2), do they all die during the inflammatory reaction, or do some of them survive? Do those that survive become quiescent tissue macrophages, able to react as naïve cells to a new challenge? Or, do monocyte-derived tissue macrophages conserve a “memory” of their past inflammatory activation? This review will address some of these important questions under the general framework of the role of monocytes and macrophages in the initiation, development, resolution, and chronicization of inflammation.

IL-1 family nomenclature

To the Editor: Newly cloned interleukin 1 (IL-1) family members1–3 were originally given an IL-1 family (IL-1F) designation4, but as functions have now been elucidated for several of these5,6, we propose that each now be assigned an individual interleukin designation. IL-1F6, IL-1F8 and IL-1F9 are encoded by distinct genes but use the same receptor complex (IL-1Rrp2 and AcP), are proinflammatory and deliver nearly identical signals7–12. We propose these be designated IL-36α, IL-36β and IL-36γ, respectively. IL-1F5 also binds to IL-1Rrp2 but antagonizes those cytokines in a manner analogous to that used by IL-1Ra to antagonize IL-1α and IL-1β7–9. We propose that IL-1F5 be renamed IL-36Ra (for ‘receptor antagonist’). In the IL-1 nomenclature, IL-1Ra is used for the natural product, whereas IL-1ra is used for the recombinant product; therefore, IL-36Ra is appropriate for natural IL-1F5. IL-1F7 produces anti-inflammatory effects by suppressing innate immune responses; it does this by decreasing the production of inflammatory cytokines induced by Toll-like receptor agonists as well as that of IL-1 and tumor necrosis factor13,14. We propose this IL-1 family member be renamed IL-37. IL-1F7 has various splice forms1,2,15,16, of which IL-1F7b is the most studied. We propose that IL-1F7a, IL-1F7b and so on be renamed IL-37a, IL-37b and so on. The one remaining IL-1 family member, for which no function has yet been demonstrated, is IL-1F10; however, as evidence of its properties remains limited, we suggest that it retain its IL-1F designation until a function is clearly identified, although it might be prudent to reserve the designation IL-38 for this eventuality.

IL-37: a new anti-inflammatory cytokine of the IL-1 family

The IL-1 family of cytokines encompasses eleven proteins that each share a similar β-barrel structure and bind to Ig-like receptors. Some of the IL-1-like cytokines have been well characterised, and play key roles in the development and regulation of inflammation. Indeed, IL-1α (IL-1F1), IL-1β (IL-1F2), and IL-18 (IL-1F4) are well-known inflammatory cytokines active in the initiation of the inflammatory reaction and in driving Th1 and Th17 inflammatory responses. In contrast, IL-1 receptor antagonist (IL-1Ra, IL-1F3) and the receptor antagonist binding to IL-1Rrp2 (IL-36Ra, IL-1F5) reduce inflammation by blocking the binding of the agonist receptor ligands. In the case of IL-37 (IL-1F7), of which five different splice variants have been described, less is known of its function, and identification of the components of a heterodimeric receptor complex remains unclear. Some studies suggest that IL-37 binds to the α chain of the IL-18 receptor in a non-competitive fashion, and this may explain some of the disparate biological effects that have been reported for mice deficient in the IL-18R. The biological properties of IL-37 are mainly those of down-regulating inflammation, as assessed in models where human IL-37 is expressed in mice. In this review, an overview of the role of IL-37 in the regulation of inflammation is presented. The finding that IL-37 also locates to the nucleus, as do IL-1α and IL-33, for receptor-independent organ/tissue-specific regulation of inflammation is also reviewed.

Problems and challenges in the development and validation of human cell-based assays to determine nanoparticle-induced immunomodulatory effects

BackgroundWith the increasing use of nanomaterials, the need for methods and assays to examine their immunosafety is becoming urgent, in particular for nanomaterials that are deliberately administered to human subjects (as in the case of nanomedicines). To obtain reliable results, standardised in vitro immunotoxicological tests should be used to determine the effects of engineered nanoparticles on human immune responses. However, before assays can be standardised, it is important that suitable methods are established and validated.ResultsIn a collaborative work between European laboratories, existing immunological and toxicological in vitro assays were tested and compared for their suitability to test effects of nanoparticles on immune responses. The prototypical nanoparticles used were metal (oxide) particles, either custom-generated by wet synthesis or commercially available as powders. Several problems and challenges were encountered during assay validation, ranging from particle agglomeration in biological media and optical interference with assay systems, to chemical immunotoxicity of solvents and contamination with endotoxin.ConclusionThe problems that were encountered in the immunological assay systems used in this study, such as chemical or endotoxin contamination and optical interference caused by the dense material, significantly affected the data obtained. These problems have to be solved to enable the development of reliable assays for the assessment of nano-immunosafety.

IL-18 cDNA vaccination protects mice from spontaneous lupus-like autoimmune disease

The lupus-like autoimmune syndrome of MRL/Mp-Tnfrsf6lpr (lpr) mice is characterized by progressive lymphadenopathy and autoantibody production, leading to early death from renal failure. Activation of T helper lymphocytes is one of the events in the pathogenesis of the disease in these mice and likely in human systemic lupus erythematosus. Among T helper lymphocytedependent cytokines, IFN-γ plays a pivotal role in the abnormal cell activation and the fatal development of the lpr disease. IL-18, an inducer of IFN-γ in T lymphocytes and natural killer cells, may contribute to the disease because cells from lpr mice are hypersensitive to IL-18 and express high levels of IL-18. To assess the contribution of IL-18 to the pathogenesis in the animal model, in vivo inhibition of IL-18 was attempted. Young lpr mice were vaccinated against autologous IL-18 by repeated administration of a cDNA coding for the murine IL-18 precursor. Vaccinated mice produced autoantibodies to murine IL-18 and exhibited a significant reduction in spontaneous lymphoproliferation and IFN-γ production as well as less glomerulonephritis and renal damage. Moreover, mortality was significantly delayed in anti-IL-18-vaccinated mice. These studies support the concept that IL-18 plays a major role in the pathogenesis of the autoimmune syndrome of lpr mice and that a reduction in IL-18 activity could be a therapeutic strategy in autoimmune diseases.

Role of metalloproteases in the release of the IL-1 type II decoy receptor

The IL-1 type II receptor (decoy RII) is a nonsignaling molecule the only established function of which is to capture IL-1 and prevent it from interacting with signaling receptor. The decoy RII is released in a regulated way from the cell surface. Here, we reported that hydroxamic acid inhibitors of matrix metalloproteases inhibit different pathways of decoy RII release, including the following: (a) the slow (18 h) gene expression-dependent release from monocytes and polymorphonuclear cells exposed to dexamethasone; (b) rapid release (minutes) from myelomonocytic cells exposed to tumor necrosis factor, chemoattractants, or phorbol myristate acetate; (c) phorbol myristate acetate-induced release from decoy RII-transfected fibroblasts and B cells. Inhibition of release was associated with increased surface expression of decoy RII. Inhibitors of other protease classes did not substantially affect release. However, serine protease inhibitors increased the molecular mass of the decoy RII released from polymorphonuclear cells from 45 to 60 kDa. Thus, irrespective of the pathway responsible for release and of the cellular context, matrix metalloproteases, rather than differential splicing, play a key role in production of soluble decoy RII.

Concern-driven integrated approaches to nanomaterial testing and assessment - report of the NanoSafety Cluster Working Group 10

Abstract Bringing together topic-related European Union (EU)-funded projects, the so-called “NanoSafety Cluster” aims at identifying key areas for further research on risk assessment procedures for nanomaterials (NM). The outcome of NanoSafety Cluster Working Group 10, this commentary presents a vision for concern-driven integrated approaches for the (eco-)toxicological testing and assessment (IATA) of NM. Such approaches should start out by determining concerns, i.e., specific information needs for a given NM based on realistic exposure scenarios. Recognised concerns can be addressed in a set of tiers using standardised protocols for NM preparation and testing. Tier 1 includes determining physico-chemical properties, non-testing (e.g., structure–activity relationships) and evaluating existing data. In tier 2, a limited set of in vitro and in vivo tests are performed that can either indicate that the risk of the specific concern is sufficiently known or indicate the need for further testing, including details for such testing. Ecotoxicological testing begins with representative test organisms followed by complex test systems. After each tier, it is evaluated whether the information gained permits assessing the safety of the NM so that further testing can be waived. By effectively exploiting all available information, IATA allow accelerating the risk assessment process and reducing testing costs and animal use (in line with the 3Rs principle implemented in EU Directive 2010/63/EU). Combining material properties, exposure, biokinetics and hazard data, information gained with IATA can be used to recognise groups of NM based upon similar modes of action. Grouping of substances in return should form integral part of the IATA themselves.

The interleukin-1 receptor family

The cytokines IL-1 and IL-18 are key molecules both in the innate and in the adaptive immune response. Their activity is mediated by specific receptors present on the membrane of target cells. It has become apparent that these receptors are members of a larger family of related receptors, most of which are apparently involved in the mechanisms of host defense. Thus, the large Toll/IL-1R (TIR) superfamily encompasses the Ig domain family (IL-1 receptors, IL-18 receptors, and IL-1R-like receptors), the leucine-rich domain family [the Toll-like receptors (TLR) and similar receptors], and a series of TIR domain-containing intracellular adapter molecules. The TIR superfamily is defined by a common intracellular TIR domain, involved in the initiation of signaling. A group of TIR domain-containing adapters (MyD88, TIRAP, TRIF, and TRAM) are differentially recruited to the Toll/IL-1 receptors, contributing to the specificity of signaling. Recent studies have also begun to unravel the mechanisms of negative regulation of the Toll/IL-1 receptors. The orphan receptor TIR8/SIGIRR, a member of TIR superfamily, while unable to initiate signaling, can negatively modulate the TIR-mediated responses. Other negative regulators of the Toll/IL-1R family include T1/ST2, some soluble forms of TLR, and MyD88s. The coordinated positive and negative regulation of the TIR activation ensures the appropriate modulation of the innate and inflammatory responses and avoids the risk of pathological derangement. This chapter will consider in detail the characteristics and functional role of the Ig domain receptor subfamily in the regulation of host defense and their possible role in pathology.

The suitability of different cellular in vitro immunotoxicity and genotoxicity methods for the analysis of nanoparticle-induced events

Abstract Suitable assays and test strategies are needed to analyze potential genotoxic and immunotoxic health effects caused by nanoparticle exposure. The development and validation of such methods is challenging because nanoparticles may show unexpected behavior, like aggregation or interference with optical measurements, when routine in vitro assays are performed. In our interdisciplinary study, the effects of inorganic gold (4.5 nm) and iron oxide (7.3 nm) nanoparticles with a narrow size distribution were tested on human cells using different assay systems. The results show that cytotoxicity as well as immunotoxicity and genotoxicity induced by these two inorganic nanoparticles was low or absent when using a panel of cell-based tests in different laboratories. However, several technical issues had to be tackled that were specific for working with nanoparticles. The methods used, their suitability for nanotoxicity testing, and the technical problems encountered are carefully described and discussed in this paper.

Model of interaction of the IL-1 receptor accessory protein IL-1RAcP with the IL-1β/IL-1RI complex

A preliminary model has been calculated for the activating interaction of the interleukin 1 receptor (IL‐1R) accessory protein IL‐1RAcP with the ligand/receptor complex IL‐1β/IL‐1RI. First, IL‐1RAcP was modeled on the crystal structure of IL‐1RI bound to IL‐1β. Then, the IL‐1RAcP model was docked using specific programs to the crystal structure of the IL‐1β/IL‐1RI complex. Two types of models were predicted, with comparable probability. Experimental data obtained with the use of IL‐1β peptides and antibodies, and with mutated IL‐1β proteins, support the BACK model, in which IL‐1RAcP establishes contacts with the back of IL‐1RI wrapped around IL‐1β.