LILRA5 is expressed by synovial tissue macrophages in rheumatoid arthritis,selectively induces pro‐inflammatory cytokines and IL‐10 and is regulated by TNF‐α, IL‐10 and IFN‐γ

Leukocyte immunoglobulin‐like receptor A5 (LILRA5) belongs to a family of receptors known to regulate leukocyte activation. There are two membrane‐bound and two soluble forms of LILRA5. The transmembrane LILRA5 contain a short cytoplasmic domain and a charged arginine residue within the transmembrane region. Cross‐linking of LILRA5 on monocytes induced production of pro‐inflammatory cytokines, suggesting that LILRA5 plays a role in inflammation. However, expression of LILRA5 in diseases with extensive inflammatory component is unknown. Rheumatoid arthritis (RA) is a chronic inflammatory synovitis characterized by unregulated activation of leukocytes leading to joint destruction. Here we demonstrate extensive LILRA5 expression on synovial tissue macrophages and in synovial fluid of patients with active RA but not in patients with osteoarthritis. We also show that LILRA5 associated with the common γ chain of the FcR and LILRA5 cross‐linking induced phosphorylation of Src tyrosine kinases and Spleen tyrosine kinase (Syk). Furthermore, LILRA5 induced selective production of pro‐inflammatory cytokines as well as IL‐10. LILRA5 mRNA and protein expression was tightly regulated by TNF‐α, IL‐10 and IFN‐γ. Increased expression of LILRA5 in rheumatoid tissue, together with its ability to induce key cytokines involved in RA, suggests that this novel receptor may contribute to disease pathogenesis.     

IL‐4 blocks M‐CSF‐dependent macrophage proliferation by inducing p21Waf1 in a STAT6‐dependent way

Macrophages are recruited from the blood stream to the inflammatory loci to carry out their functional activities. In an early phase of the cell cycle, macrophages become activated by Th1‐type cytokines (i.e. IFN‐γ), thereby producing several factors (cytokines, NO, etc.) and developing pro‐inflammatory activities. When bacteria and apoptotic bodies are removed, through the interaction with Th2‐type cytokines (i.e. IL‐4), macrophages become anti‐inflammatory and repair damaged tissues. Incubation of bone‐marrow‐derived macrophages with IFN‐γ or IL‐4 blocked their proliferation. While M‐CSF withdrawal caused cell cycle arrest at the early G₁ phase, treatment of macrophages with IFN‐γ or IL‐4 caused this arrest later, at the G₁/S boundary. Proliferation arrest was not due to an induction of apoptosis. IFN‐γ and IL‐4 induced the expression of the cyclin‐dependent kinase (Cdk) inhibitor p21^Waf1. Using KO mice and iRNA experiments, we found that p21^Waf1is required for IL‐4‐ but not for IFN‐γ‐dependent inhibition of macrophage proliferation. IL‐4 inhibited M‐CSF‐dependent Cdk‐2 and Cdk‐4 activities, which are necessary for entry and passage through the S phase of the cell cycle. The signal transduction used to induce the expression of p21^Waf1after interaction of IL‐4 with the corresponding receptor was mediated by STAT6. Thus, IL‐4 and IFN‐γ blocked M‐CSF‐induced macrophage proliferation through distinct mechanisms.     

Both IFN‐γ and IL‐17 are required for the development of severe autoimmune gastritis

IL‐17, produced by a distinct lineage of CD4⁺ helper T (Th) cells termed Th17 cells, induces the production of pro‐inflammatory cytokines from resident cells and it has been demonstrated that over‐expression of IL‐17 plays a crucial role in the onset of several auto‐immune diseases. Here we examined the role of IL‐17 in the pathogenesis of autoimmune gastritis, a disease that was previously believed to be mediated by IFN‐γ. Significantly higher levels of IL‐17 and IFN‐γ were found in the stomachs and stomach‐draining lymph nodes of mice with severe autoimmune gastritis. Unlike IL‐17, which was produced solely by CD4⁺ T cells in gastritic mice, the majority of IFN‐γ‐producing cells were CD8⁺ T cells. However, CD8⁺ T cells alone were not able to induce autoimmune gastritis. T cells that were deficient in IL‐17 or IFN‐γ production were able to induce autoimmune gastritis but to a much lower extent compared with the disease induced by wild‐type T cells. These data demonstrate that production of neither IL‐17 nor IFN‐γ by effector T cells is essential for the initiation of autoimmune gastritis, but suggest that both are required for the disease to progress to the late pathogenic stage that includes significant tissue disruption.     

The role of IL‐28, IFN‐γ, and TNF‐α in predicting response to pegylated interferon/ribavirin in chronic HCV patients

The primary goal of HCV therapy is to achieve a sustained virological response (SVR). Many host and viral factors influence the treatment response. Cytokines play an important role in the defense against viral infections, where successful treatment of hepatitis C depends on a complex balance between pro‐ and anti‐inflammatory responses. In the present study, we investigated the relationship between the presence and percentage of some cytokines (IL‐28, IFN‐γ, and TNF‐α) regarding different clinicopathological parameters including response to therapy in chronic HCV patients using immunohistochemical technique. This study was carried out on 64 chronic HCV patients (34 responders and 30 non‐responders). Of cases, 54% showed IL‐28 expression, which was associated with low AST (p = 0.002) and low HAI score (p = 0.006). Of cases, 67 and 45% showed IFN‐γ and TNF‐α expression, respectively, where the median percentage of TNF‐α expression was higher in grade II spotty necrosis compared to grade I. Some inflammatory cytokines expressed by intrahepatic inflammatory cells in chronic HCV patients promote inflammation and injury (pro‐inflammatory) such as TNF‐α. Other cytokines aid in resolving inflammation and injury (anti‐inflammatory) such as IL‐28. The balance between these cytokines will determine the degree of inflammatory state. None of the investigated cytokines proved its clear cut role in affecting response to therapy, however, their levels varied between responders and non‐responders for further investigations to clarify.     

Adipose invariant natural killer T cells

Adipose tissue is a dynamic organ that makes up a substantial proportion of the body; in severe obesity it can account for 50% of body mass. Details of the unique immune system resident in human and murine adipose tissue are only recently emerging, and so it has remained a largely unexplored and unappreciated immune site until now. Adipose tissue harbours a unique collection of immune cells, which often display unusual functions compared with their counterparts elsewhere in the body. These resident immune cells are key to maintaining tissue and immune homeostasis, yet in obesity their chronic aberrant stimulation can contribute to the inflammation and pathogenesis associated with obesity. Anti‐inflammatory adipose‐resident lymphocytes are often depleted in obesity, whereas pro‐inflammatory immune cells accumulate, leading to an overall inflammatory state, which is a key step in the development of obesity‐induced metabolic disease. A good example is invariant natural killer T (iNKT) cells, which make up a large proportion of lymphocytes in human and murine adipose tissue. Here, they are unusually poised to produce anti‐inflammatory or regulatory cytokines, however in obesity, iNKT cells are greatly reduced. As iNKT cells are potent transactivaors of other immune cells, and can act as a bridge between innate and adaptive immunity, their loss in obesity represents the loss of a major regulatory population. Restoring iNKT cells, or activating them in obese mice leads to improved glucose handling, insulin sensitivity, and even weight loss, and hence represents an exciting therapeutic avenue to be explored for restoring homeostasis in obese adipose tissue.     

Gal‐3 regulates the capacity of dendritic cells to promote NKT‐cell‐induced liver injury

Galectin‐3 (Gal‐3), an endogenous lectin, exhibits pro‐ and anti‐inflammatory effects in various disease conditions. In order to explore the role of Gal‐3 in NKT‐cell‐dependent pathology, we induced hepatitis in C57BL/6 WT and Gal‐3‐deficient mice by using specific ligand for NKT cells: α‐galactosylceramide, glycolipid Ag presented by CD1d. The injection of α‐galactosylceramide significantly enhanced expression of Gal‐3 in liver NKT and dendritic cells (DCs). Genetic deletion or selective inhibition of Gal‐3 (induced by Gal‐3‐inhibitor TD139) abrogated the susceptibility to NKT‐cell‐dependent hepatitis. Blood levels of pro‐inflammatory cytokines (TNF‐α, IFN‐γ, IL‐12) and their production by liver DCs and NKT cells were also downregulated. Genetic deletion or selective inhibition of Gal‐3 alleviated influx of inflammatory CD11c⁺CD11b⁺ DCs in the liver and favored tolerogenic phenotype and IL‐10 production of liver NKT and DCs. Deletion of Gal‐3 attenuated the capacity of DCs to support liver damage in the passive transfer experiments and to produce pro‐inflammatory cytokines in vitro. Gal‐3‐deficient DCs failed to optimally stimulate production of pro‐inflammatory cytokines in NKT cells, in vitro and in vivo. In conclusion, Gal‐3 regulates the capacity of DCs to support NKT‐cell‐mediated liver injury, playing an important pro‐inflammatory role in acute liver injury.     

Adipose tissue macrophages

It is now broadly accepted that low-grade chronic inflammation associated with obesity leads to the onset of insulin resistance and type 2 diabetes mellitus. Obesity-associated inflammation is characterized by an increased abundance of macrophages in adipose tissue along with production of inflammatory cytokines. Adipose tissue macrophages (ATMs) are suspected to be the major source of inflammatory mediators such as TNF-alpha and IL-6 that interfere with adipocyte function by inhibiting insulin action. However, ATMs phenotypically resemble alternatively activated (M2) macrophages and are capable of anti-inflammatory mediator production challenging the concept that ATMs are simply the "bad guys" in obese adipose tissue. Triggers promoting ATM recruitment, ATM functions and dysfunctions, and stimuli and molecular mechanisms that drive them into becoming detrimental to their environment are subject to current research. Strategies to interfere with ATM recruitment and adverse activation could give rise to novel options for treatment and prevention of insulin resistance and type 2 diabetes mellitus.     

Type I interferon potentiates T‐cell receptor mediated induction of IL‐10‐producing CD4+ T cells

Type I interferons (IFNs) have the dual ability to promote the development of the immune response and exert an anti‐inflammatory activity. We analyzed the integrated effect of IFN‐α, TCR signal strength, and CD28 costimulation on human CD4⁺ T‐cell differentiation into cell subsets producing the anti‐ and proinflammatory cytokines IL‐10 and IFN‐γ. We show that IFN‐α boosted TCR‐induced IL‐10 expression in activated peripheral CD45RA⁺CD4⁺ T cells and in whole blood cultures. The functional cooperation between TCR and IFN‐α efficiently occurred at low engagement of receptors. Moreover, IFN‐α rapidly cooperated with anti‐CD3 stimulation alone. IFN‐α, but not IL‐10, drove the early development of type I regulatory T cells that were mostly IL‐10⁺ Foxp3^? IFN‐γ^? and favored IL‐10 expression in a fraction of Foxp3⁺ T cells. Our data support a model in which IFN‐α costimulates TCR toward the production of IL‐10 whose level can be amplified via an autocrine feedback loop.     

IL‐18‐induced expression of high‐affinity IL‐2R on murine NK cells is essential for NK‐cell IFN‐γ production during murine Plasmodium yoelii infection

Early production of pro‐inflammatory cytokines, including IFN‐γ, is essential for control of blood‐stage malaria infections. We have shown that IFN‐γ production can be induced among human natural killer (NK) cells by coculture with Plasmodium falciparum infected erythrocytes, but the importance of this response is unclear. To further explore the role of NK cells during malaria infection, we have characterized the NK‐cell response of C57BL/6 mice during lethal (PyYM) or nonlethal (Py17XNL) P. yoelii infection. Ex vivo flow cytometry revealed that NK cells are activated within 24 h of Py17XNL blood‐stage infection, expressing CD25 and producing IFN‐γ; this response was blunted and delayed during PyYM infection. CD25 expression and IFN‐γ production were highly correlated, suggesting a causal relationship between the two responses. Subsequent in vitro experiments revealed that IL‐18 signaling is essential for induction of CD25 and synergizes with IL‐12 to enhance CD25 expression on splenic NK cells. In accordance with this, Py17XNL‐infected erythrocytes induced NK‐cell CD25 expression and IFN‐γ production in a manner that is completely IL‐18‐ and partially IL‐12‐dependent, and IFN‐γ production is enhanced by IL‐2. These data suggest that IL‐2 signaling via CD25 amplifies IL‐18‐ and IL‐12‐mediated NK‐cell activation during malaria infection.     

T-cell immunoglobulin- and mucin-domain-containing molecule-4 maintains adipose tissue homeostasis by orchestrating M2 macrophage polarization via nuclear factor kappa B pathway

Obesity is generally associated with low-grade inflammation. Adipose tissue macrophages (ATMs) orchestrate metabolic inflammation. The classical (M1-like) or alternative (M2-like) activation of ATMs is functionally coupled with the metabolic status of fat tissues. It has been found that T-cell immunoglobulin- and mucin-domain-containing molecule-4 (Tim-4) inhibits inflammation by regulating macrophages. However, the exact role of Tim-4 in macrophage polarization and obesity remains unknown. Here, we identified Tim-4 as a critical switch governing macrophage M1/M2 polarization and energy homeostasis. Tim-4 deletion led to spontaneous obesity in elder mice and promoted obesity severity of db/db mice. Obesity microenvironment enhanced the expression of Tim-4 in white adipose tissue and ATMs. In vitro, we detected an increase in M1-like cells and decrease in M2-like cells in both peritoneal macrophages and bone marrow-derived macrophages from Tim-4 knockout mice. Mechanistically, we demonstrated that Tim-4 promoted M2-like macrophages polarization via suppressing nuclear factor kappa B (NF-κB) signaling pathway. In addition, we found that Tim-4 promoted TLR4 internalization, which might contribute to regulation of NF-κB signaling. Collectively, these results indicated that Tim-4 maintained adipose tissue homeostasis by regulating macrophage polarization via NF-κB pathway, which would provide a new target for obesity intervention.     

Pulmonary inflammation in mice with collagen‐induced arthritis is conditioned by complete Freund's adjuvant and regulated by endogenous IFN‐γ

Following immunization with collagen II (CII) in complete Freund's adjuvant (CFA), DBA/1 mice develop arthritis of major joints. This collagen‐induced arthritis (CIA) is used as a model for rheumatoid arthritis (RA) in man. Inflammatory changes in lung tissue commonly occur in RA. However, evidence for pulmonary inflammation in CIA is scarce and ambiguous. Here, we demonstrate pulmonary inflammation accompanying CIA in wild‐type DBA/1 mice. In IFN‐γ receptor‐deficient (IFN‐γR KO) mice, inflammation was more frequent and more severe. Injection of CFA only (without CII) proved to be as efficient in eliciting pulmonary inflammation as immunization with CFA + CII, though being less effective in causing arthritis. Significant correlation in severity between joint and pulmonary involvement could not be demonstrated. Macroscopic, microscopic, and functional characteristics of pulmonary inflammation in the mice resembled those seen in human RA. Increased inflammation in IFN‐γR KO mice was accompanied by augmented expression of various cytokines and chemokines, as measured by RT‐PCR on affected tissue. Treatment with a TNF‐α inhibitor ameliorated lung pathology. We conclude that CIA in DBA/1 mice is accompanied by pulmonary inflammation. Although both disease processes are kept in check by endogenous IFN‐γ, lack of strict parallelism indicates that overlap in their pathogeneses is partial.     

Enhanced protection from fibrosis and inflammation in the combined absence of IL‐13 and IFN‐γ

Persistent or dysregulated IL‐13 responses are key drivers of fibrosis in multiple organ systems, and this identifies this cytokine as an important therapeutic target. Nevertheless, the mechanisms by which IL‐13 blockade leads to the amelioration of fibrosis remain unclear. Because IFN‐γ exhibits potent anti‐fibrotic activity, and IL‐4Rα signalling antagonizes IFN‐γ effector function, compensatory increases in IFN‐γ activity following IL‐13/IL‐4Rα blockade might contribute to the reduction in fibrosis. To investigate the role of IFN‐γ, we developed novel IL‐13^?/?/IFN‐γ^?/? double cytokine‐deficient mice and examined disease progression in models of type 2‐driven fibrosis. As predicted, we showed that fibrosis in the lung and liver are both highly dependent on IL‐13. We also observed increased IFN‐γ production and inflammatory activity in the tissues of IL‐13‐deficient mice. Surprisingly, however, an even greater reduction in fibrosis was observed in IL‐13/IFN‐γ double deficient mice, most notably in the livers of mice chronically infected with Schistosoma mansoni. The increased protection was associated with marked decreases in Tgfb1, Mmp12, and Timp1 mRNA expression in the tissues; reduced inflammation; and decreased expression of important pro‐inflammatory mediators such as TNF‐α. Experiments conducted with neutralizing monoclonal antibodies to IL‐13 and IFN‐γ validated the findings with the genetically deficient mice. Together, these studies demonstrate that the reduction in fibrosis observed when IL‐13 signalling is suppressed is not dependent on increased IFN‐γ activity. Instead, by reducing compensatory increases in type 1‐associated inflammation, therapeutic strategies that block IFN‐γ and IL‐13 activity simultaneously can confer greater protection from progressive fibrosis than IL‐13 blockade alone. Published 2016. This article is a U.S. Government work and is in the public domain in the USA.     

Morphological and Inflammatory Changes in Visceral Adipose Tissue During Obesity

Obesity is a major health burden worldwide and is a major factor in the development of insulin resistance and metabolic complications such as type II diabetes. Chronic nutrient excess leads to visceral adipose tissue (VAT) expansion and dysfunction in an active process that involves the adipocytes, their supporting matrix, and immune cell infiltrates. These changes contribute to adipose tissue hypoxia, adipocyte cell stress, and ultimately cell death. Accumulation of lymphocytes, macrophages, and other immune cells around dying adipocytes forms the so-called “crown-like structure”, a histological hallmark of VAT in obesity. Cross talk between immune cells in adipose tissue dictates the overall inflammatory response, ultimately leading to the production of pro-inflammatory mediators which directly induce insulin resistance in VAT. In this review, we summarize recent studies demonstrating the dramatic changes that occur in visceral adipose tissue during obesity leading to low-grade chronic inflammation and metabolic disease.     

Expression and Identification of Immunological Activities of the HIV‐gp120N‐Human Interferon Gamma Fusion Protein

The human immunodeficiency virus type 1 (HIV‐1) envelope glycoprotein gp120 is a vaccine immunogen that has been studied extensively. To enhance the immune response of cells against HIV‐1 gp120, we tested the coexpression of gp120N with interferon‐γ (IFN‐γ) as an immune adjuvant. Two recombinant prokaryotic plasmids were constructed: the pET44b‐HIV‐1‐gp120N plasmid construct carried the HIV‐1 gp120N gene (pET44‐gp120N), whereas the pET44b‐HIV‐1‐gp120N‐IFN‐γ plasmid construct carried a fusion gp120N‐IFN‐γ gene (pET44b‐gp120N‐IFN‐γ). Target protein expression was achieved in E. coli BL21 (DE3) cells by chemical induction. To test the immunological activity of the proteins, mice were injected with a control, gp120N, or the fusion gp120N‐IFN‐γ protein. The serum and spleen cells of the mice were collected for immunological detection. Results showed that specific T lymphocyte proliferation and the expression of the Th1‐type cytokines (IL‐2 and IFN‐γ) were higher in the gp120N‐IFN‐γ group than the other two groups (P < 0.05). No difference was observed in the expression levels of the Th2‐type cytokines (IL‐4 and IL‐10; P > 0.05). These results suggest that IFN‐γ plays a prominent role as an immune adjuvant when coexpressed with HIV‐1 gp120N. IFN‐γ enhances the specific cell immune response of mice against HIV‐1 gp120. Anat Rec, 292:381–386, 2009. © 2009 Wiley‐Liss, Inc.     

Reverse plasticity: TGF‐β and IL‐6 induce Th1‐to‐Th17‐cell transdifferentiation in the gut

Th17 cells are a heterogeneous population of pro‐inflammatory T cells that have been shown to mediate immune responses against intestinal bacteria. Th17 cells are highly plastic and can transdifferentiate to Th1/17 cells or unconventional Th1 cells, which are highly pathogenic in animal models of immune‐mediated diseases such as inflammatory bowel diseases. A recent European Journal of Immunology article by Liu et al. (Eur. J. Immunol. 2015. 45:1010–1018) showed, surprisingly, that Th1 cells have a similar plasticity, and could transdifferentiate to Th17 cells. Thus, IFN‐γ‐producing Th1 effector cells specific for an intestinal microbial antigen were shown to acquire IL‐17‐producing capacities in the gut in a mouse model of colitis, and in response to TGF‐β and IL‐6 in vitro. TGF‐β induced Runx1, and together with IL‐6 was shown to render the ROR‐γt and IL‐17 promoters in Th1 cells accessible for Runx1 binding. In this commentary, we discuss how this unexpected plasticity of Th1 cells challenges our view on the generation of Th1/17 cells with the capacity to co‐produce IL‐17 and IFN‐γ, and consider possible implications of this Th1‐to‐Th17‐cell conversion for therapies of inflammatory bowel diseases and protective immune responses against intracellular pathogens.     

Functional interleukin‐33 receptors are expressed in early progenitor stages of allergy‐related granulocytes

Interleukin‐33 (IL‐33) induces T helper type 2 (Th2) cytokine production and eosinophilia independently of acquired immunity, leading to innate immunity‐mediated allergic inflammation. Allergy‐related innate myeloid cells such as eosinophils, basophils and mast cells express the IL‐33 receptor (IL‐33R), but it is still unknown how IL‐33 regulates allergic inflammation involving these cells and their progenitors. Here, we revealed that the functional IL‐33R was expressed on eosinophil progenitors (EoPs), basophil progenitors (BaPs) and mast cell progenitors (MCPs). In the presence of IL‐33, these progenitors did not expand, but produced a high amount of Th2 and pro‐inflammatory cytokines such as IL‐9, IL‐13, IL‐1β and IL‐6. The amount of cytokines produced by these progenitors was greater than that by mature cells. In vivo, IL‐33 stimulated the expansion of EoPs, but it was dependent upon the elevated serum IL‐5 that is presumably derived from type 2 innate lymphoid cells that express functional IL‐33R. These data collectively suggest that EoPs, BaPs and MCPs are not only the sources of allergy‐related granulocytes, but can also be sources of allergy‐related cytokines in IL‐33‐induced inflammation. Because such progenitors can differentiate into mature granulocytes at the site of inflammation, they are potential therapeutic targets in IL‐33‐related allergic diseases.     

An update on immune dysregulation in obesity‐related insulin resistance

Obesity is associated with chronic low‐grade inflammation of the adipose tissue (AT) that might develop into systemic inflammation, insulin resistance (IR) and an increased risk of type 2 diabetes mellitus (T2DM) in severe obese rodents and humans. In the lean state, small normal adipocytes and AT macrophages interact with each other to maintain metabolic homeostasis but during obesity, enlarged adipocytes secrete inflammatory mediators and express immune receptors to recruit immune cells and aggravate the inflammation. The better understanding of the obesity‐related inflammatory milieu and the sequential events leading to IR could be helpful in designing new preventive and therapeutic strategies. The present review will discuss the cellular and molecular abnormalities participating in the pathogenesis of obesity in obese individuals as well as high‐fat diet (HFD)‐fed mice, a mouse model of obesity.