Clinical antiviral efficacy of favipiravir in early COVID-19 (PLATCOV): an open-label, randomised, controlled, adaptive platform trial.

In early symptomatic COVID-19 treatment, high dose oral favipiravir did not accelerate viral clearance. BACKGROUND: Favipiravir, an anti-influenza drug, has in vitro antiviral activity against SARS-CoV-2. Clinical trial evidence to date is inconclusive. Favipiravir has been recommended for the treatment of COVID-19 in some countries. METHODS: In a multicentre open-label, randomised, controlled, adaptive platform trial, low-risk adult patients with early symptomatic COVID-19 were randomised to one of ten treatment arms including high dose oral favipiravir (3.6g on day 0 followed by 1.6g daily to complete 7 days treatment) or no study drug. The primary outcome was the rate of viral clearance (derived under a linear mixed-effects model from the daily log10 viral densities in standardised duplicate oropharyngeal swab eluates taken daily over 8 days [18 swabs per patient]), assessed in a modified intention-to-treat population (mITT). The safety population included all patients who received at least one dose of the allocated intervention. This ongoing adaptive platform trial was registered at ClinicalTrials.gov (NCT05041907) on 13/09/2021. RESULTS: In the final analysis, the mITT population contained data from 114 patients randomised to favipiravir and 126 patients randomised concurrently to no study drug. Under the linear mixed-effects model fitted to all oropharyngeal viral density estimates in the first 8 days from randomisation (4,318 swabs), there was no difference in the rate of viral clearance between patients given favipiravir and patients receiving no study drug; a -1% (95% credible interval: -14 to 14%) difference. High dose favipiravir was well-tolerated. INTERPRETATION: Favipiravir does not accelerate viral clearance in early symptomatic COVID-19. The viral clearance rate estimated from quantitative measurements of oropharyngeal eluate viral densities assesses the antiviral efficacy of drugs in vivo with comparatively few studied patients.

Clinical Antiviral Efficacy of Remdesivir in Coronavirus Disease 2019: An Open-Label, Randomized Controlled Adaptive Platform Trial (PLATCOV).

BACKGROUND: Uncertainty over the therapeutic benefit of parenteral remdesivir in coronavirus disease 2019 (COVID-19) has resulted in varying treatment guidelines. METHODS: In a multicenter open-label, controlled, adaptive, pharmacometric platform trial, low-risk adult patients with early symptomatic COVID-19 were randomized to 1 of 8 treatment arms including intravenous remdesivir (200 mg followed by 100 mg daily for 5 days) or no study drug. The primary outcome was the rate of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) clearance (estimated under a linear model fit to the daily log10 viral densities, days 0-7) in standardized duplicate oropharyngeal swab eluates, in a modified intention-to-treat population. This ongoing adaptive trial is registered at ClinicalTrials.gov (NCT05041907). RESULTS: The 2 study arms enrolled 131 patients (remdesivir n = 67, no study drug n = 64) and estimated viral clearance rates from a median of 18 swab samples per patient (a total of 2356 quantitative polymerase chain reactions). Under the linear model, compared with the contemporaneous control arm (no study drug), remdesivir accelerated mean estimated viral clearance by 42% (95% credible interval, 18%-73%). CONCLUSIONS: Parenteral remdesivir accelerates viral clearance in early symptomatic COVID-19. Pharmacometric assessment of therapeutics using the method described can determine in vivo clinical antiviral efficacy rapidly and efficiently.

PI3K Isoforms in Cell Signalling and Innate Immune Cell Responses.

Phosphoinositide-3-kinases (PI3Ks) are enzymes involved in signalling and modification of the function of all mammalian cells. These enzymes phosphorylate the 3-hydroxyl group of the inositol ring of phosphatidylinositol, resulting in lipid products that act as second messengers responsible for coordinating many cellular functions, including activation, chemotaxis, proliferation and survival. The identification of the functions that are mediated by a specific PI3K isoform is complex and depends on the specific cell type and inflammatory context. In this chapter we will focus on the role of PI3K isoforms in the context of innate immunity, focusing on the mechanisms by which PI3K signalling regulates phagocytosis, the activation of immunoglobulin, chemokine and cytokines receptors, production of ROS and cell migration, and how PI3K signalling plays a central role in host defence against infections and tissue injury.

Ouratea spectabilis and its Biflavanone Ouratein D Exert Potent Anti-inflammatory Activity in MSU Crystal-induced Gout in Mice.

Gouty arthritis (GA) is an inflammatory arthritis triggered by the deposition of monosodium urate monohydrate (MSU) crystals, causing pain, inflammation, and joint damage. Several drugs are currently employed to manage acute flares of GA, but they either have limited effectiveness or induce severe adverse reactions. Ouratea spectabilis is traditionally used in Brazil to treat gastric ulcers and rheumatism. The ethanolic extract of O. spectabilis stems (OSpC) and four biflavanones (ouratein A - D) isolated thereof were evaluated in a murine model of GA induced by the injection of MSU crystals. The underlying mechanism of action of ouratein D was investigated in vitro in cell cultures by measurement of IL-1ß levels by ELISA and Western blot analysis. The administration of OSpC (10, 30 or 100 mg/Kg, p. o.) reduced the migration of total inflammatory cells, monocytes, and neutrophils and diminished the levels of IL-1ß and CXCL1 in the synovial tissue. Among the tested compounds, only ouratein D (1 mg/Kg) reduced the migration of the inflammatory cells and it was shown to be active up to 0.01 mg/Kg (equivalent to 0.34 nM/Kg, p. o.). Treatment of pre-stimulated THP-1 cells (differentiated into macrophages) or BMDMs with ouratein D reduced the release of IL-1ß in both macrophage lines. This biflavanone reduced the activation of caspase-1 (showed by the increase in the cleaved form) in supernatants of cultured BMDMs, evidencing its action in modulating the inflammasome pathway. The obtained results demonstrate the anti-gout properties of O. spectabilis and point out ouratein D as the bioactive component of the assayed extract.

Eosinophil plays a crucial role in intestinal mucositis induced by antineoplastic chemotherapy.

Mucositis is a major clinical complication associated with cancer treatment and may limit the benefit of chemotherapy. Leukocytes and inflammatory mediators have been extensively associated with mucositis severity. However, the role of eosinophils in the pathophysiology of chemotherapy-induced mucositis remains to be elucidated. Here, using GATA-1-deficient mice, we investigated the role of eosinophils in intestinal mucositis. There was marked accumulation of eosinophils in mice given irinotecan and eosinophil ablation inhibited intestinal mucositis. Treatment with Evasin-4, a chemokine receptor antagonist, reduced the recruitment of eosinophils and decreased irinotecan-induced mucositis. Importantly, Evasin-4 did not interfere negatively with the antitumour effects of irinotecan. Evasin-4 was of benefit for mice given high doses of irinotecan once Evasin-4-treated mice presented delayed mortality. Altogether, our findings suggest that Evasin-4 may have significant mucosal-protective effects in the context of antineoplastic chemotherapy and may, therefore, be useful in combination with anticancer treatment in cancer patients.

PAF signaling plays a role in obesity-induced adipose tissue remodeling.

BACKGROUND/OBJECTIVES: Platelet-activating factor receptor (PAFR) activation controls adipose tissue (AT) expansion in animal models. Our objective was twofold: (i) to check whether PAFR signaling is involved in human obesity and (ii) investigate the PAF pathway role in hematopoietic or non-hematopoietic cells to control adipocyte size. MATERIALS/SUBJECTS AND METHODS: Clinical parameters and adipose tissue gene expression were evaluated in subjects with obesity. Bone marrow (BM) transplantation from wild-type (WT) or PAFR-/- mice was performed to obtain chimeric PAFR-deficient mice predominantly in hematopoietic or non-hematopoietic-derived cells. A high carbohydrate diet (HC) was used to induce AT remodeling and evaluate in which cell compartment PAFR signaling modulates it. Also, 3T3-L1 cells were treated with PAF to evaluate fat accumulation and the expression of genes related to it. RESULTS: PAFR expression in omental AT from humans with obesity was negatively correlated to different corpulence parameters and more expressed in the stromal vascular fraction than adipocytes. Total PAFR-/- increased adiposity compared with WT independent of diet-induced obesity. Differently, WT mice receiving PAFR-/--BM exhibited similar adiposity gain as WT chimeras. PAFR-/- mice receiving WT-BM showed comparable augmentation in adiposity as total PAFR-/- mice, demonstrating that PAFR signaling modulates adipose tissue expansion through non-hematopoietic cells. Indeed, the PAF treatment in 3T3-L1 adipocytes reduced fat accumulation and expression of adipogenic genes. CONCLUSIONS: Therefore, decreased PAFR signaling may favor an AT accumulation in humans and animal models. Importantly, PAFR signaling, mainly in non-hematopoietic cells, especially in adipocytes, appears to play a significant role in regulating diet-induced AT expansion.

Mediators of the Resolution of the Inflammatory Response.

The termination of inflammation is governed by endogenous molecules collectively referred to as 'mediators of resolution' of inflammation. There is now strong evidence to suggest that failed resolution may underpin autoimmune and inflammatory diseases and could thus be targeted to decrease inflammation. There are many molecules that have been described as mediators of resolution, and new players are still being continuously discovered. To support the emerging field of 'resolution pharmacology', here we discuss the scientific strategies required to qualify a molecule as a resolution mediator. Systematic definition of the players of resolution, their receptors, and downstream mechanisms remains a necessary knowledge to move the field forward and suggest new targets for the development of novel therapies to treat inflammatory diseases.

Future directions for the discovery of natural product-derived immunomodulating drugs: an IUPHAR positional review.

Drug discovery from natural sources is going through a renaissance, having spent many decades in the shadow of synthetic molecule drug discovery, despite the fact that natural product-derived compounds occupy a much greater chemical space than those created through synthetic chemistry methods. With this new era comes new possibilities, not least the novel targets that have emerged in recent times and the development of state-of-the-art technologies that can be applied to drug discovery from natural sources. Although progress has been made with some immunomodulating drugs, there remains a pressing need for new agents that can be used to treat the wide variety of conditions that arise from disruption, or over-activation, of the immune system; natural products may therefore be key in filling this gap. Recognising that, at present, there is no authoritative article that details the current state-of-the-art of the immunomodulatory activity of natural products, this in-depth review has arisen from a joint effort between the International Union of Basic and Clinical Pharmacology (IUPHAR) Natural Products and Immunopharmacology Sections, with contributions from a number of world-leading researchers in the field of natural product drug discovery, to provide a "position statement" on what natural products has to offer in the search for new immunomodulatory argents. To this end, we provide a historical look at previous discoveries of naturally occurring immunomodulators, present a picture of the current status of the field and provide insight into the future opportunities and challenges for the discovery of new drugs to treat immune-related diseases.

Influenza Virus Infection Impairs the Gut's Barrier Properties and Favors Secondary Enteric Bacterial Infection through Reduced Production of Short-Chain Fatty Acids.

Along with respiratory tract disease per se, viral respiratory infections can also cause extrapulmonary complications with a potentially critical impact on health. In the present study, we used an experimental model of influenza A virus (IAV) infection to investigate the nature and outcome of the associated gut disorders. In IAV-infected mice, the signs of intestinal injury and inflammation, altered gene expression, and compromised intestinal barrier functions peaked on day 7 postinfection. As a likely result of bacterial component translocation, gene expression of inflammatory markers was upregulated in the liver. These changes occurred concomitantly with an alteration of the composition of the gut microbiota and with a decreased production of the fermentative, gut microbiota-derived products short-chain fatty acids (SCFAs). Gut inflammation and barrier dysfunction during influenza were not attributed to reduced food consumption, which caused in part gut dysbiosis. Treatment of IAV-infected mice with SCFAs was associated with an enhancement of intestinal barrier properties, as assessed by a reduction in the translocation of dextran and a decrease in inflammatory gene expression in the liver. Lastly, SCFA supplementation during influenza tended to reduce the translocation of the enteric pathogen Salmonella enterica serovar Typhimurium and to enhance the survival of doubly infected animals. Collectively, influenza virus infection can remotely impair the gut's barrier properties and trigger secondary enteric infections. The latter phenomenon can be partially countered by SCFA supplementation.

CD300a contributes to the resolution of articular inflammation triggered by MSU crystals by controlling neutrophil apoptosis.

Gout is an inflammatory disease triggered by deposition of monosodium urate (MSU) crystals in the joints, resulting in high neutrophil influx and pain. Here, we studied the role of the inhibitory receptor CD300a in the resolution process in a murine model of gout. We found increased CD300a expression on neutrophils emigrated to the joint. When compared to WT mice, CD300a-/- mice had persistent neutrophil influx till 24 hr after MSU injection. This was associated with increased concentration of IL-1ß and greater tissue damage in the joints of CD300a-/- mice. There was an increase in the percentage of apoptotic neutrophils in the synovial lavage of WT mice, as compared to CD300a-/- mice. This difference was reflected in the decline of efferocytic events in the synovial cavity of CD300a-/- mice 24 hr after MSU injection. A CD300a agonistic antibody was shown, for the first time, to increase apoptosis of human neutrophils, and this was associated with cleavage of caspase-8. In conclusion, our results reveal an important role of CD300a in the control of leucocyte infiltration, IL-1ß production and caspase-8 cleavage in neutrophils, contributing to the resolution of inflammation triggered by MSU injection.

Therapeutic potential of the FPR2/ALX agonist AT-01-KG in the resolution of articular inflammation.

The resolution of inflammation is a dynamic process, characterized by the biosynthesis of pro-resolving mediators, including the lipid Lipoxin A4 (LXA4). LXA4 acts on the N-formyl peptide receptor 2 (FPR2/ALX) to mediate anti-inflammatory and pro-resolving effects. In order to exploit the therapeutic potential of endogenous LXA4 in the context of inflammation we have recently developed synthetic LXA4 mimetics (sLXms) including a dimethyl-imidazole-containing FPR2/ALX agonist designated AT-01-KG. Here, we have investigated the effect of treatment with AT-01-KG in established models of articular inflammation. In a model of gout, mice were injected with MSU crystals and treated with AT-01-KG at the peak of inflammatory response. The treatment decreased the number of neutrophils in the knee exudate, an effect which was accompanied by low levels of myeloperoxidase, CXCL1 and IL-1ß in periarticular tissue. AT-01-KG treatment led to reduced tissue damage and hypernociception. The effects of AT-01-KG on neutrophil accumulation were not observed in MSU treated FPR2/3-/-mice. Importantly, AT-01-KG induced resolution of articular inflammation by increasing neutrophil apoptosis and subsequent efficient efferocytosis. In a model of antigen-induced arthritis, AT-01-KG treatment also attenuated inflammatory responses. These data suggest that AT-01-KG may be a potential new therapy for neutrophilic inflammation of the joints.

Relevance of angiotensin-(1-7) and its receptor Mas in pneumonia caused by influenza virus and post-influenza pneumococcal infection.

Resolution failure of exacerbated inflammation triggered by Influenza A virus (IAV) prevents return of pulmonary homeostasis and survival, especially when associated with secondary pneumococcal infection. Therapeutic strategies based on pro-resolving molecules have great potential against acute inflammatory diseases. Angiotensin-(1-7) [Ang-(1-7)] is a pro-resolving mediator that acts on its Mas receptor (MasR) to promote resolution of inflammation. We investigated the effects of Ang-(1-7) and the role of MasR in the context of primary IAV infection and secondary pneumococcal infection and evaluated pulmonary inflammation, virus titers and bacteria counts, and pulmonary damage. Therapeutic treatment with Ang-(1-7) decreased neutrophil recruitment, lung injury, viral load and morbidity after a primary IAV infection. Ang-(1-7) induced apoptosis of neutrophils and efferocytosis of these cells by alveolar macrophages, but had no direct effect on IAV replication in vitro. MasR-deficient (MasR-/-) mice were highly susceptible to IAV infection, displaying uncontrolled inflammation, increased viral load and greater lethality rate, as compared to WT animals. Ang-(1-7) was not protective in MasR-/- mice. Interestingly, Ang-(1-7) given during a sublethal dose of IAV infection greatly reduced morbidity associated with a subsequent S. pneumoniae infection, as seen by decrease in the magnitude of neutrophil influx, number of bacteria in the blood leading to a lower lethality. Altogether, these results show that Ang-(1-7) is highly protective against severe primary IAV infection and protects against secondary bacterial infection of the lung. These effects are MasR-dependent. Mediators of resolution of inflammation, such as Ang-(1-7), should be considered for the treatment of pulmonary viral infections.

The Long Pentraxin 3 Contributes to Joint Inflammation in Gout by Facilitating the Phagocytosis of Monosodium Urate Crystals.

The purpose of this study was to investigate the role of pentraxin 3 (PTX3), a pivotal component of the innate immune system, in gout. Levels of PTX3 and IL-1ß in human samples were evaluated by ELISA. Development of murine gout was evaluated through the levels of cytokines (PTX3, CXCL1, and IL-1ß) and neutrophil recruitment into the joint cavity. Phagocytosis of monosodium urate (MSU) crystals and caspase-1 activation were determined by flow cytometer. Acute gout patients showed elevated concentration of PTX3 in plasma and synovial fluid as compared with healthy and osteoarthritic subjects. Moreover, there was a positive correlation between intra-articular PTX3 and IL-1ß levels. PTX3 was induced in the periarticular tissue of mice postinjection of MSU crystals. Importantly, Ptx3-deficient mice showed reduced inflammation in response to MSU crystal injection compared with wild-type mice, including reduction of neutrophil recruitment into the joint cavity and IL-1ß and CXCL1 production. Interestingly, addition of PTX3 in vitro enhanced MSU crystal phagocytosis by monocytes and resulted in higher production of IL-1ß by macrophages. This contribution of PTX3 to the phagocytosis of MSU crystals and consequent production of IL-1ß occurred through a mechanism mainly dependent on FcγRIII. Thus, our results suggest that PTX3 acts as a humoral pattern recognition molecule in gout facilitating MSU crystal phagocytosis and contributing to the pathogenesis of gouty arthritis.

The role of annexin A1 in the modulation of the NLRP3 inflammasome.

Annexins are well-known Ca2+ phospholipid-binding proteins, which have a wide variety of cellular functions. The role of annexin A1 (AnxA1) in the innate immune system has focused mainly on the anti-inflammatory and proresolving properties through its binding to the formyl-peptide receptor 2 (FPR2)/ALX receptor. However, studies suggesting an intracellular role of AnxA1 are emerging. In this study, we aimed to understand the role of AnxA1 for interleukin (IL)-1ß release in response to activators of the nucleotide-binding domain leucine-rich repeat (NLR) and pyrin domain containing receptor 3 (NLRP3) inflammasome. Using AnxA1 knockout mice, we observed that AnxA1 is required for IL-1ß release in vivo and in vitro. These effects were due to reduction of transcriptional levels of IL-1ß, NLRP3 and caspase-1, a step called NLRP3 priming. Moreover, we demonstrate that AnxA1 co-localize and directly bind to NLRP3, suggesting the role of AnxA1 in inflammasome activation is independent of its anti-inflammatory role via FPR2. Therefore, AnxA1 regulates NLRP3 inflammasome priming and activation in a FPR2-independent manner.

ACKR2 contributes to pulmonary dysfunction by shaping CCL5:CCR5-dependent recruitment of lymphocytes during influenza A infection in mice.

Inflammation triggered by influenza A virus (IAV) infection is important for viral clearance, induction of adaptive responses, and return to lung homeostasis. However, an exaggerated immune response, characterized by the overproduction of chemokines, can lead to intense lung injury, contributing to mortality. Chemokine scavenger receptors, such as ACKR2, control the levels of CC chemokines influencing the immune responses. Among the chemokine targets of ACKR2, CCL5 is important to recruit and activate lymphocytes. We investigated the role of ACKR2 during IAV infection in mice. Pulmonary ACKR2 expression was increased acutely after IAV infection preceding the virus-induced lung dysfunction. ACKR2-knockout (ACKR2-/-) mice were protected from IAV, presenting decreased viral burden and lung dysfunction. Mechanistically, the absence of ACKR2 resulted in augmented airway CCL5 levels, secreted by mononuclear and plasma cells in the lung parenchyma. The higher chemokine gradient led to an augmented recruitment of T and B lymphocytes, formation of inducible bronchus-associated lymphoid tissue and production of IgA in the airways of ACKR2-/- mice post-IAV. CCL5 neutralization in ACKR2-/- mice prevented lymphocyte recruitment and increased bronchoalveolar lavage fluid protein levels and pulmonary dysfunction. Finally, CCR5-/- mice presented increased disease severity during IAV infection, displaying increased neutrophils, pulmonary injury and dysfunction, and accentuated lethality. Collectively, our data showed that ACKR2 dampens CCL5 levels and the consequent recruitment of CCR5+ T helper 1 (Th1), T regulatory cells (Tregs), and B lymphocytes during IAV infection, decreasing pathogen control and promoting lung dysfunction in wild type mice. Therefore, ACKR2 is detrimental and CCR5 is protective during IAV infection coordinating innate and adaptive immune responses in mice.

Inhibition of the sphingosine-1-phosphate pathway promotes the resolution of neutrophilic inflammation.

Sphingosine-1-phosphate (S1P) is an important sphingolipid derived from plasma membrane and has a known role in productive phase of inflammation, but its role in neutrophil survival and resolution phase of inflammation is unknown. Here, we investigated the effects of inhibition of S1P receptors and the blockade of S1P synthesis in BALB/c mice and human neutrophils. S1P and S1PR1-3 receptors expression were increased in cells from the pleural cavity stimulated with LPS. Using different antagonists of S1PRs and inhibitors of different steps of the metabolic pathway of S1P production, we show that S1P and its receptors are involved in regulating neutrophil survival and resolution of inflammation in the pleural cavity. Given the role of the S1P-S1PR axis in resolution of inflammation, we sought to identify whether blockade at different levels of the sphingosine-1-phosphate synthesis pathway could affect neutrophil survival in vitro. Inhibitors of the S1P pathway were also able to induce human neutrophil apoptosis. In addition, blockade of S1P synthesis or its receptor facilitated the efferocytosis of apoptotic neutrophil. Taken together, our data demonstrate a fundamental role for S1P in regulating the outcome of inflammatory responses, and position S1P-S1PR axis as a potential target for treatment of neutrophilic inflammation.

Decreased expression of neuronal nitric oxide synthase contributes to the endothelial dysfunction associated with cigarette smoking in human.

Endothelial nitric oxide synthase (eNOS) malfunctioning has been proposed to contribute to the endothelial damage produced by cigarette. Besides eNOS, neuronal NOS (nNOS) is also expressed in most vascular tissues and plays an important role in the endothelium-dependent vascular relaxation. We hypothesize that nNOS may contribute to the endothelium dysfunction produced by cigarette in smokers. Vascular function was assessed in human resistance mesenteric arteries using a wire myograph, the level of protein expression by Western blot, eNOS and nNOS localization by immunofluorescence. Measurement of NO was assessed by fluorescence microscopy. Arteries of smokers showed impaired endothelium-dependent vascular relaxation in response to acetylcholine. Pharmacological nonselective blockade of NOS with l-NAME and selective nNOS blockade with inhibitor 1 reduced the relaxation of the mesenteric artery of both smokers and nonsmokers. Interestingly, the inhibitory effect of NOS inhibitors was greater in nonsmokers than in smokers. The expression of total nNOS and eNOS and the level of phosphorylation at eNOS-pSer1177 were reduced in arteries of smokers as compared with nonsmokers. No differences between groups were observed in the expression of total COX-1, COX-2, catalase and SOD-1. Immunofluorescence analysis showed the presence of nNOS in the vascular endothelium in both groups. Acetylcholine-induced NO production was impaired in arteries from smokers as compared to nonsmokers. Selective inhibition of nNOS caused a decreased in NO production, which was greater in nonsmokers than in smokers. Our data show that a decrease in nNOS expression contributes to the endothelial dysfunction caused by cigarette smoking in human.

Blame the signaling: Role of cAMP for the resolution of inflammation.

A complex intracellular signaling governs different cellular responses in inflammation. Extracellular stimuli are sensed, amplified, and transduced through a dynamic cellular network of messengers converting the first signal into a proper response: production of specific mediators, cell activation, survival, or death. Several overlapping pathways are coordinated to ensure specific and timely induction of inflammation to neutralize potential harms to the tissue. Ideally, the inflammatory response must be controlled and self-limited. Resolution of inflammation is an active process that culminates with termination of inflammation and restoration of tissue homeostasis. Comparably to the onset of inflammation, resolution responses are triggered by coordinated intracellular signaling pathways that transduce the message to the nucleus. However, the key messengers and pathways involved in signaling transduction for resolution are still poorly understood in comparison to the inflammatory network. cAMP has long been recognized as an inducer of anti-inflammatory responses and cAMP-dependent pathways have been extensively exploited pharmacologically to treat inflammatory diseases. Recently, cAMP has been pointed out as coordinator of key steps of resolution of inflammation. Here, we summarize the evidence for the role of cAMP at inducing important features of resolution of inflammation.

Glucocorticoid-induced leucine zipper modulates macrophage polarization and apoptotic cell clearance.

Macrophages are professional phagocytes that display remarkable plasticity, with a range of phenotypes that can be broadly characterized by the M1/M2 dichotomy. Glucocorticoid (GC)-induced leucine zipper (GILZ) is a protein known to mediate anti-inflammatory and some pro-resolving actions, including as neutrophil apoptosis. However, the role of GILZ in key macrophage function is not well understood. Here, we investigated the role of GILZ on macrophage reprogramming and efferocytosis. Using murine bone-marrow-derived macrophages (BMDMs), we found that GILZ was expressed in naive BMDMs and exhibited increased expression in M2-like macrophages (IL4-differentiated). M1-like macrophages (IFN/LPS-differentiated) from GILZ-/- mice showed higher expression of the M1 markers CD86, MHC class II, iNOS, IL-6 and TNF-α, associated with increased levels of phosphorylated STAT1 and lower IL-10 levels, compared to M1-differentiated cells from WT mice. There were no changes in the M2 markers CD206 and arginase-1 in macrophages from GILZ-/- mice differentiated with IL-4, compared to cells from WT animals. Treatment of M1-like macrophages with TAT-GILZ, a cell-permeable GILZ fusion protein, decreased the levels of CD86 and MHC class II in M1-like macrophages without modifying CD206 levels in M2-like macrophages. In line with the in vitro data, increased numbers of M1-like macrophages were found into the pleural cavity of GILZ-/- mice after LPS-injection, compared to WT mice. Moreover, efferocytosis was defective in the context of GILZ deficiency, both in vitro and in vivo. Conversely, treatment of LPS-injected mice with TAT-GILZ promoted inflammation resolution, associated with lower numbers of M1-like macrophages and increased efferocytosis. Collectively, these data indicate that GILZ is a regulator of important macrophage functions, contributing to macrophage reprogramming and efferocytosis, both key steps for the resolution of inflammation.

IL-33 enhances macrophage release of IL-1ß and promotes pain and inflammation in gouty arthritis.

OBJECTIVE: To investigate the role of IL-33 in gouty arthritis. MATERIAL: 174 Balb/c (wild-type) and 54 ST2-/- mice were used in this study. In vitro experiments were conducted in bone marrow-derived macrophages (BMDMs). Synovial fluid samples from gouty arthritis (n = 7) and osteoarthritis (n = 8) hospital patients were used to measure IL-33 and sST2 levels. METHODS: Gout was induced by injection of monosodium urate (MSU) crystals in the knee joint of mice. Pain was determined using the electronic von Frey and static weight bearing. Neutrophil recruitment was determined by H&E staining, Rosenfeld staining slides, and MPO activity. ELISA was used for cytokine and sST2 measurement. The priming effect of IL-33 was determined in BMDM. RESULTS: Synovial fluid of gout patients showed higher IL-33 levels and neutrophil counts than osteoarthritis patients. In mice, the absence of ST2 prevented mechanical pain, knee joint edema, neutrophil recruitment to the knee joint, and lowered IL-1ß and superoxide anion levels. In macrophages, IL-33 enhanced the release of IL-1ß and TNF-α, and BMDMs from ST2-/- showed reduced levels of these cytokines after stimulus with MSU crystals. CONCLUSION: IL-33 mediates gout pain and inflammation by boosting macrophages production of cytokines upon MSU crystals stimulus.