An IL-9-pulmonary macrophage axis defines the allergic lung inflammatory environment.

Despite IL-9 functioning as a pleiotropic cytokine in mucosal environments, the IL-9-responsive cell repertoire is still not well defined. Here, we found that IL-9 mediates proallergic activities in the lungs by targeting lung macrophages. IL-9 inhibits alveolar macrophage expansion and promotes recruitment of monocytes that develop into CD11c+ and CD11c- interstitial macrophage populations. Interstitial macrophages were required for IL-9-dependent allergic responses. Mechanistically, IL-9 affected the function of lung macrophages by inducing Arg1 activity. Compared with Arg1-deficient lung macrophages, Arg1-expressing macrophages expressed greater amounts of CCL5. Adoptive transfer of Arg1+ lung macrophages but not Arg1- lung macrophages promoted allergic inflammation that Il9r-/- mice were protected against. In parallel, the elevated expression of IL-9, IL-9R, Arg1, and CCL5 was correlated with disease in patients with asthma. Thus, our study uncovers an IL-9/macrophage/Arg1 axis as a potential therapeutic target for allergic airway inflammation.

Arginine and Arginases Modulate Metabolism, Tumor Microenvironment and Prostate Cancer Progression.

Arginine availability and activation of arginine-related pathways at cancer sites have profound effects on the tumor microenvironment, far beyond their well-known role in the hepatic urea cycle. Arginine metabolism impacts not only malignant cells but also the surrounding immune cells behavior, modulating growth, survival, and immunosurveillance mechanisms, either through an arginase-mediated effect on polyamines and proline synthesis, or by the arginine/nitric oxide pathway in tumor cells, antitumor T-cells, myeloid-derived suppressor cells, and macrophages. This review presents evidence concerning the impact of arginine metabolism and arginase activity in the prostate cancer microenvironment, highlighting the recent advances in immunotherapy, which might be relevant for prostate cancer. Even though further research is required, arginine deprivation may represent a novel antimetabolite strategy for the treatment of arginine-dependent prostate cancer.

A Monocyte-Orchestrated IFN-I-to-IL-4 Cytokine Axis Instigates Protumoral Macrophages and Thwarts Poly(I:C) Therapy.

Type I IFNs (IFN-I) are important for tumor immune surveillance and contribute to the therapeutic responses for numerous treatment regimens. Nevertheless, certain protumoral activities by IFN-I have been increasingly recognized. Indeed, our recent work showed that systemic poly(I:C)/IFN treatment can undesirably trigger high arginase (ARG1) expression within the tumor-associated monocyte/macrophage compartment. Using a line of CRISPR-generated Arg1-YFP reporter knock-in mice, we have determined that a subset of tumor-associated macrophages represent the major Arg1-expressing cell type following poly(I:C)/IFN stimulation. More detailed analyses from in vitro and in vivo models demonstrate a surprising IFN-to-IL-4 cytokine axis in transitional monocytes, which can subsequently stimulate IL-4 target genes, including Arg1, in macrophages. Intriguingly, IFN stimulation of transitional monocytes yielded concurrent M2 (YFP+)- and M1 (YFP-)-skewed macrophage subsets, correlated with an inhibitory crosstalk between IFN-I and IL-4. Genetic abrogation of IL-4 signaling in mice diminished poly(I:C)/IFN-induced ARG1 in tumors, leading to enhanced activation of CD8+ T cells and an improved therapeutic effect. The present work uncovered a monocyte-orchestrated macrophage phenotype conversion mechanism that may have broad implications.

TRAF6 Regulates the Immunosuppressive Effects of Myeloid-Derived Suppressor Cells in Tumor-Bearing Host.

Myeloid-derived suppressor cells (MDSCs) are immature heterogeneous cells derived from the bone marrow and they are the major component of the tumor-induced immunosuppressive environment. Tumor necrosis factor receptor-associated factor 6 (TRAF6), an E3 ubiquitin ligase, catalyzes the polyubiquitination of target proteins. TRAF6 plays a critical role in modulating the immune system. However, whether TRAF6 is involved in the regulation of MDSCs has not been thoroughly elucidated to date. In this study, we found that the expression of TRAF6 in MDSCs derived from tumor tissue was significantly upregulated compared with that of MDSCs from spleen of tumor-bearing mice. Knockdown of TRAF6 remarkably attenuated the immunosuppressive effects of MDSCs. Mechanistically, TRAF6 might improve the immunosuppression of MDSCs by mediating K63-linked polyubiquitination and phosphorylation of signal transducer and activator of transcription 3 (STAT3). Additionally, it was discovered that the accumulation of MDSCs was abnormal in peripheral blood of lung cancer patients. TRAF6 and arginase 1 were highly expressed in MDSCs of patients with lung cancer. Taken together, our study demonstrated that TRAF6 participates in promoting the immunosuppressive function of MDSCs and provided a potential target for antitumor immunotherapy.

More than just protein building blocks: how amino acids and related metabolic pathways fuel macrophage polarization.

Macrophages represent the first line of defence in innate immune responses and additionally serve important functions for the regulation of host inflammation and tissue homeostasis. The M1/M2 model describes the two extremes of macrophage polarization states, which can be induced by multiple stimuli, most notably by LPS/IFN-γ and IL-4/IL-13. Historically, the expression of two genes encoding for enzymes, which use the same amino acid as their substrate, iNOS and ARG1, has been used to define classically activated M1 (iNOS) and alternatively activated M2 (ARG1) macrophages. This 'arginine dichotomy' has recently become a matter of debate; however, in parallel with the emerging field of immunometabolism there is accumulating evidence that these two enzymes and their related metabolites are fundamentally involved in the intrinsic regulation of macrophage polarization and function. The aim of this review is to highlight recent advances in macrophage biology and immunometabolism with a specific focus on amino acid metabolism and their related metabolic pathways: iNOS/ARG1 (arginine), TCA cycle and OXPHOS (glutamine) as well as the one-carbon metabolism (serine, glycine).

Melanoma-associated fibroblasts impair CD8+ T cell function and modify expression of immune checkpoint regulators via increased arginase activity.

This study shows that melanoma-associated fibroblasts (MAFs) suppress cytotoxic T lymphocyte (CTL) activity and reveals a pivotal role played by arginase in this phenomenon. MAFs and normal dermal fibroblasts (DFs) were isolated from surgically resected melanomas and identified as Melan-A-/gp100-/FAP+ cells. CTLs of healthy blood donors were activated in the presence of MAF- and DF-conditioned media (CM). Markers of successful CTL activation, cytotoxic degranulation, killing activity and immune checkpoint regulation were evaluated by flow cytometry, ELISPOT, and redirected killing assays. Soluble mediators responsible for MAF-mediated effects were identified by ELISA, flow cytometry, inhibitor assays, and knock-in experiments. In the presence of MAF-CM, activated/non-naïve CTLs displayed dysregulated ERK1/2 and NF-κB signaling, impeded CD69 and granzyme B production, impaired killing activity, and upregulated expression of the negative immune checkpoint receptors TIGIT and BTLA. Compared to DFs, MAFs displayed increased amounts of VISTA and HVEM, a known ligand of BTLA on T cells, increased L-arginase activity and CXCL12 release. Transgenic arginase over-expression further increased, while selective arginase inhibition neutralized MAF-induced TIGIT and BTLA expression on CTLs. Our data indicate that MAF interfere with intracellular CTL signaling via soluble mediators leading to CTL anergy and modify immune checkpoint receptor availability via L-arginine depletion.

Atherosclerosis Linked to Aberrant Amino Acid Metabolism and Immunosuppressive Amino Acid Catabolizing Enzymes.

Cardiovascular disease is the leading global health concern and responsible for more deaths worldwide than any other type of disorder. Atherosclerosis is a chronic inflammatory disease in the arterial wall, which underpins several types of cardiovascular disease. It has emerged that a strong relationship exists between alterations in amino acid (AA) metabolism and the development of atherosclerosis. Recent studies have reported positive correlations between levels of branched-chain amino acids (BCAAs) such as leucine, valine, and isoleucine in plasma and the occurrence of metabolic disturbances. Elevated serum levels of BCAAs indicate a high cardiometabolic risk. Thus, BCAAs may also impact atherosclerosis prevention and offer a novel therapeutic strategy for specific individuals at risk of coronary events. The metabolism of AAs, such as L-arginine, homoarginine, and L-tryptophan, is recognized as a critical regulator of vascular homeostasis. Dietary intake of homoarginine, taurine, and glycine can improve atherosclerosis by endothelium remodeling. Available data also suggest that the regulation of AA metabolism by indoleamine 2,3-dioxygenase (IDO) and arginases 1 and 2 are mediated through various immunological signals and that immunosuppressive AA metabolizing enzymes are promising therapeutic targets against atherosclerosis. Further clinical studies and basic studies that make use of animal models are required. Here we review recent data examining links between AA metabolism and the development of atherosclerosis.

Myeloid-derived suppressor cells shift Th17/Treg ratio and promote systemic lupus erythematosus progression through arginase-1/miR-322-5p/TGF-ß pathway.

Immune cells play important roles in systemic lupus erythematosus (SLE). We previously found that myeloid-derived suppressor cell (MDSC)-derived arginase-1 (Arg-1) promoted Th17 cell differentiation in SLE. In the present study, we performed RNA-chip to identify the microRNA regulation network between MDSCs and Th17 cells. miR-542-5p in humans, as the homologous gene of miR-322-5p in mice was significantly up-regulated in the Th17+MDSC group compared with Th17 cells cultured alone and down-regulated in the Th17+MDSC+Arg-1 inhibitor group compared with the Th17+MDSC group. We further evaluated the miR-322-5p and Th17/Treg balance in mice and found that the proportions of both Th17 cells and Tregs were elevated and that miR-322-5p overexpression activated the transforming growth factor-ß pathway. Moreover, although miR-322-5p expression was higher in SLE mice, it decreased after treatment with an Arg-1 inhibitor. The proportion of Th17 cells and Th17/Treg ratio correlated with miR-322-5p levels. In conclusion, MDSC-derived Arg-1 and mmu-miR-322-5p not only promote Th17 cell and Treg differentiation, but also shift the Th17/Treg ratio in SLE. The Arg-1/miR-322-5p axis may serve as a novel treatment target for SLE.

Extensive sequence and structural evolution of Arginase 2 inhibitory antibodies enabled by an unbiased approach to affinity maturation.

Affinity maturation is a powerful technique in antibody engineering for the in vitro evolution of antigen binding interactions. Key to the success of this process is the expansion of sequence and combinatorial diversity to increase the structural repertoire from which superior binding variants may be selected. However, conventional strategies are often restrictive and only focus on small regions of the antibody at a time. In this study, we used a method that combined antibody chain shuffling and a staggered-extension process to produce unbiased libraries, which recombined beneficial mutations from all six complementarity-determining regions (CDRs) in the affinity maturation of an inhibitory antibody to Arginase 2 (ARG2). We made use of the vast display capacity of ribosome display to accommodate the sequence space required for the diverse library builds. Further diversity was introduced through pool maturation to optimize seven leads of interest simultaneously. This resulted in antibodies with substantial improvements in binding properties and inhibition potency. The extensive sequence changes resulting from this approach were translated into striking structural changes for parent and affinity-matured antibodies bound to ARG2, with a large reorientation of the binding paratope facilitating increases in contact surface and shape complementarity to the antigen. The considerable gains in therapeutic properties seen from extensive sequence and structural evolution of the parent ARG2 inhibitory antibody clearly illustrate the advantages of the unbiased approach developed, which was key to the identification of high-affinity antibodies with the desired inhibitory potency and specificity.

Myeloid Cell-Derived Arginase in Cancer Immune Response.

Amino acid metabolism is a critical regulator of the immune response, and its modulating becomes a promising approach in various forms of immunotherapy. Insufficient concentrations of essential amino acids restrict T-cells activation and proliferation. However, only arginases, that degrade L-arginine, as well as enzymes that hydrolyze L-tryptophan are substantially increased in cancer. Two arginase isoforms, ARG1 and ARG2, have been found to be present in tumors and their increased activity usually correlates with more advanced disease and worse clinical prognosis. Nearly all types of myeloid cells were reported to produce arginases and the increased numbers of various populations of myeloid-derived suppressor cells and macrophages correlate with inferior clinical outcomes of cancer patients. Here, we describe the role of arginases produced by myeloid cells in regulating various populations of immune cells, discuss molecular mechanisms of immunoregulatory processes involving L-arginine metabolism and outline therapeutic approaches to mitigate the negative effects of arginases on antitumor immune response. Development of potent arginase inhibitors, with improved pharmacokinetic properties, may lead to the elaboration of novel therapeutic strategies based on targeting immunoregulatory pathways controlled by L-arginine degradation.

The pro-apoptotic ARTS protein induces neutrophil apoptosis, efferocytosis, and macrophage reprogramming to promote resolution of inflammation.

ARTS (Sept4_i2) is a pro-apoptotic protein and a product of the Sept4 gene. ARTS acts upstream of mitochondria to initiate caspase activation. ARTS induces apoptosis by specifically binding XIAP and allowing de-repression of active caspases required for Mitochondrial Outer Membrane Permeabilzation (MOMP). Moreover, ARTS promotes apoptosis by inducing ubiquitin-mediated degradation of both major anti-apoptotic proteins XIAP and Bcl-2. In the resolution phase of inflammation, the infiltrating leukocytes, which execute the acute innate response, undergo apoptosis and are subsequently cleared by phagocytic macrophages (i.e. efferocytosis). In this course, macrophages undergo reprogramming from inflammatory, to anti-inflammatory, and eventually to resolving macrophages that leave the injury sites. Since engulfment of apoptotic leukocytes is a key signaling step in macrophage reprogramming and resolution of inflammation, we hypothesized that a failed apoptosis in leukocytes in vivo would result in an impaired resolution process. To test this hypothesis, we utilized the Sept4/ARTS-/- mice, which exhibit resistance to apoptosis in many cell types. During zymosan A-induced peritonitis, Sept4/ARTS-/- mice exhibited impaired resolution of inflammation, characterized by reduced neutrophil apoptosis, macrophage efferocytosis and expression of pro-resolving mediators. This was associated with increased pro-inflammatory cytokines and reduced anti-inflammatory cytokines, secreted by resolution-phase macrophages. Moreover, ARTS overexpression in leukocytes in vitro promoted an anti-inflammatory behavior. Overall, our results suggest that ARTS is a key master-regulator necessary for neutrophil apoptosis, macrophage efferocytosis and reprogramming to the pro-resolving phenotype during the resolution of inflammation.

Autoradiography and inmmunolabeling suggests that lizard blastema contains arginase-positive M2-like macrophages that may support tail regeneration.

BACKGROUND: The regenerating blastema of the tail in the lizard Podarcis muralis contains numerous macrophages among the prevalent mesenchymal cells. Some macrophages are phagocytic but others are devoid of phagosomes suggesting that they have other roles aside phagocytosis. METHODS: The presence of healing macrophages (M2-like) has been tested using autoradiographic, immunohistochemical and ultrastructural studies. RESULTS: Autoradiography shows an uptake of tritiated arginine in sparse cells of the blastema and in the regenerating epidermis. Bioinformatics analysis suggests that epitopes for arginase-1 and -2, recognized by the employed antibody, are present in lizards. Immunofluorescence shows sparse arginase immunopositive macrophages in the blastema and few macrophages also in the apical wound epidermis. The ultrastructural study shows that macrophages contain dense secretory granules, most likely inactive lysosomes, and small cytoplasmic pale vesicles. Some of the small vesicles are arginase-positive while immunolabeling is very diffuse in the macrophage cytoplasm. CONCLUSIONS: The presence of cells incorporating arginine and of arginase 1-positive cells suggests that M2-like macrophages are present among mesenchymal and epidermal cells of the regenerative tail blastema. M2-like macrophages may promote tail regeneration differently from the numerous pro-inflammatory macrophages previously detected in the scarring limb. The presence of M2-like macrophages in addition to hyaluronate, support the hypothesis that the regenerative blastema of the tail in lizards is an immuno-privileged organ where cell proliferation and growth occur without degenerating in a tumorigenic outgrowth.

High-Throughput Fluorescence-Based Activity Assay for Arginase-1.

Arginase-1, which converts the amino acid L-arginine into L-ornithine and urea, is a promising new drug target for cancer immunotherapy, as it has a role in the regulation of T-cell immunity in the tumor microenvironment. To enable the discovery of small-molecule Arginase-1 inhibitors by high-throughput screening, we developed a novel homogeneous (mix-and-measure) fluorescence-based activity assay. The assay measures the conversion of L-arginine into L-ornithine by a decrease in fluorescent signal due to quenching of a fluorescent probe, Arginase Gold. This way, inhibition of Arginase-1 results in a gain of signal when compared with the uninhibited enzyme. Side-by-side profiling of reference inhibitors in the fluorescence-based assay and a colorimetric urea formation assay revealed similar potencies and the same potency rank order among the two assay formats. The fluorescence-based assay was successfully automated for high-throughput screening of a small-molecule library in 384-well format with a good Z'-factor and hit confirmation rate. Finally, we show that the assay can be used to study the binding kinetics of inhibitors.

Treatment of mice with a ligand binding blocking anti-CD28 monoclonal antibody improves healing after myocardial infarction.

Both conventional and regulatory CD4+ T-cells rely on costimulatory signals mediated by cell surface receptors including CD28 for full activation. We showed previously that stimulation of CD4+ Foxp3+ regulatory T-cells by superagonistic anti-CD28 monoclonal antibodies (mAb) improves myocardial healing after experimental myocardial infarction (MI). However, the effect of ligand binding blocking anti-CD28 monoclonal antibodies has not yet been tested in this context. We hypothesize that ligand blocking anti-CD28 mAb treatment might favorably impact on healing after MI by limiting the activation of conventional CD4+ T-cells. Therefore, we studied the therapeutic effect of the recently characterized mAb E18 which blocks ligand binding to CD28 in a mouse permanent coronary ligation model. E18 or an irrelevant control mAb was applied once on day two after myocardial infarction to wildtype mice. Echocardiography was performed on day 7 after MI. E18 treatment improved the survival and reduced the incidence of left ventricular ruptures after experimental myocardial infarction. Accordingly, although we found no difference in infarct size, there was significantly less left ventricular dilation after E18 treatment in surviving animals as determined by echocardiography at day 7 after MI. In sham operated control mice neither antibody had an impact on body weight, survival, and echocardiographic parameters. Mechanistically, compared to control immunoglobulin, E18 treatment reduced the number of CD4+ T-cells and monocytes/macrophages within the infarct and periinfarct zone on day 5. This was accompanied by an upregulation of arginase which is a marker for alternatively differentiated macrophages. The data indicate that CD28-dependent costimulation of CD4+ T-cells impairs myocardial healing and anti-CD28 antibody treatment constitutes a potentially clinically translatable approach to improve the outcome early after MI.

Interactions between macrophages and helminths.

Macrophages, the major population of tissue-resident mononuclear phagocytes, contribute significantly to the immune response during helminth infection. Alternatively activated macrophages (AAM) are induced early in the anti-helminth response following tissue insult and parasite recognition, amplifying the early type 2 immune cascade initiated by epithelial cells and ILC2s, and subsequently driving parasite expulsion. AAM also contribute to functional alterations in tissues infiltrated with helminth larvae, mediating both tissue repair and inflammation. Their activation is amplified and occurs more rapidly following reinfection, where they can play a dual role in trapping tissue migratory larvae and preventing or resolving the associated inflammation and damage. In this review, we will address both the known and emerging roles of tissue macrophages during helminth infection, in addition to considering both outstanding research questions and new therapeutic strategies.

Arginase promotes immune evasion of Echinococcus granulosus in mice.

BACKGROUND: Cystic echinococcosis is a chronic disease caused by infection with the larvae of Echinococcus granulosus. The parasite's ability to establish persistent infection is partly due to its evolving immune evasion strategies. One strategy may involve the protective effect of arginase, which impedes the control of pathogens or tumors, whereas it remains largely unknown during E. granulosus infection. Here, we analyzed whether arginase was produced in peritoneal cells and assessed its role in immunosuppression in mice infected with protoscoleces of E. granulosus. METHODS: BALB/c mice injected with protoscoleces of E. granulosus were used to evaluate the expression of arginase (ARG) in mRNA and protein levels. The profiles of ARG-1 expression in peritoneal cells and CD3ζ expression in T cells from spleens were assessed at different time points (3, 6, 9 and 12 months post-infection) by flow cytometry. In vitro, peritoneal cells were co-cultured with purified T cells in a transwell system, and the levels of CD3ζ re-expression were compared by flow cytometry. Meanwhile, the changes of L-arginine and its related metabolites in serum were tested. RESULTS: Compared to the control group, the peritoneal cells from infected mice showed higher levels of ARG-1 mRNA and protein, unchanged ARG-2 and iNOS. Enhanced ARG-1 expression was present in SSClowCD11b+F4/80+, CD11b+CD11c+, CD11b+Gr-1+Ly-6C+Ly-6G-, CD11b+Gr-1+Ly-6C-Ly-6G+, CD11b+Gr-1+ and CD11b+Ly-6G+ cells. The proportion of cells and the proportion of ARG-1 expression in corresponding cells exhibited a rising trend along with the extension of infection time, except for fluctuations in SSClowCD11b+F4/80+ and CD11b+CD11c+ cells at 12 months post-infection, whereas the expression of CD3ζ chain in CD4+ and CD8+ T cells showed a descending trend. Purified T cells showed declined re-expression of CD3ζ when co-cultured with peritoneal cells from infected mice, and CD3ζ was regenerated by supplement of L-arginine or arginase inhibitor BEC, rather than NOS inhibitor L-NMMA or catalase. Meanwhile, the concentrations of L-arginine, L-citrulline and NO decreased, and those of L-ornithine and urea increased in serum post-infection. CONCLUSIONS: Our findings demonstrated that ARG-1 expression is enhanced in multiple myeloid cells from peritoneum and promotes immune evasion of E. granulosus in mice by inhibiting the expression of T cell receptor CD3ζ chain and antagonism against iNOS.

LncRNA AK036396 Inhibits Maturation and Accelerates Immunosuppression of Polymorphonuclear Myeloid-Derived Suppressor Cells by Enhancing the Stability of Ficolin B.

Long noncoding RNAs (lncRNA) are emerging as crucial regulators of cell biology. However, the role of lncRNAs in the development and function of polymorphonuclear myeloid-derived suppressor cells (PMN-MDSC) remains unclear. Here, we identified that the lncRNA F730016J06Rik (AK036396) was highly expressed in PMN-MDSCs and that lncRNA AK036396 knockdown promoted the maturation and decreased the suppressive function of PMN-MDSCs. Ficolin B (Fcnb), the expression of which could be assessed as a surrogate for PMN-MDSC development, was the predicted target gene of lncRNA AK036396 based on microarray results. LncRNA AK036396 knockdown attenuated Fcnb protein stability in a manner dependent on the ubiquitin-proteasome system. Moreover, Fcnb inhibition downregulated the suppressive function of PMN-MDSCs. In addition, the expression of human M-ficolin, which is an ortholog of mouse Fcnb, was increased and positively correlated with arginase1 (ARG1) expression. This suppressive molecule is released by MDSCs, and its production is commonly used to represent the suppressive activity of MDSCs in patients with lung cancer, suggesting clinical relevance for these findings. These results indicate that lncRNA AK036396 can inhibit maturation and accelerate immunosuppression of PMN-MDSCs by enhancing Fcnb protein stability.

Detection and functional evaluation of arginase-1 isolated from human PMNs and murine MDSC.

Immunotherapy has emerged as a potent alternative for cancer treatment, unfortunately, the clinical benefit remains limited to few patients and immunotherapy resistance due to immunosuppressive tumor microenvironment represents the major reason of such a failure. Arginase-1 is one of the enzymes contributing to the establishment of such immunosuppression. Among the human immune cells, polymorphonuclear cells (PMNs) represent the major source of arginase-1, while myeloid-derived suppressor cells (MDSCs) are the main arginase-1 producing cells in mice. Due to arginase-1 potential impact in dampening the immune response, there is a growing interest in assaying arginase-1 levels and functions. Thus, in this chapter we propose how to evaluate the expression and activity of arginase in human peripheral blood-derived PMNs and in MDSCs isolated from tumor-bearing mice.

Immunosuppressive Amino-Acid Catabolizing Enzymes in Multiple Sclerosis.

Multiple sclerosis (MS) is a chronic inflammatory demyelinating disease that affects the central nervous system. Although the pathogenesis of MS is not yet fully elucidated, several evidences suggest that autoimmune processes mediated by Th1, Th17, and B cells play an important role in the development of the disease. Similar to other cells, immune cells need continuous access to amino acids (AA) in order to maintain basal metabolism and maintain vitality. When immune cells are activated by inflammation or antigenic signals, their demand for AA increases rapidly. Although AA deprivation itself may weaken the immune response under certain conditions, cells also have AA sensitive pathways that can activate intense alterations in cell metabolism based on changes in AA levels. Several data indicate that cells expressing enzymes that can degrade AA can regulate the functions of antigen-presenting cells and lymphocytes, revealing that the AA pathways are essential for controlling the function, and survival of immune cells, as well as immune cell gene expression. Basal AA catabolism may contribute to immune homeostasis and prevent autoimmunity, while increased AA catalytic activity may enhance immune suppression. In addition, there is increasing evidence that some downstream AA metabolites are important biological mediators of autoimmune response regulation. Two of the most important AA that modulate the immune response are L-Tryptophan (Trp) and L-Arginine (Arg). Tryptophan is catabolized through 2,3-dioxygenase (TDO) and indoleamine 2,3-dioxygenase (IDO) 1 and IDO2 enzymes, while three other enzymes catabolize Arg: inducible nitric oxide synthetase (iNOS), and two arginase isoforms (ARG1, ARG2). Genes encoding IDO, iNOS and ARG are induced by inflammatory cues such as cytokines, a key feature that distinguishes them from enzymes that catabolize other AA. Evidence suggests that AA catabolism is decreased in MS patients and that this decrease has functional consequences, increasing pro-inflammatory cytokines and decreasing Treg cell numbers. These effects are mediated by at least two distinct pathways involving serine/threonine kinases: the general control nonderepressible 2 kinase (GCN2K) pathway; and the mammalian target of rapamycin (mTOR) pathway. Similarly, IDO1-deficient mice showed exacerbation of experimental autoimmune encephalomyelitis (EAE), increased Th1 and Th17 cells, and decreased Treg cells. On the contrary, the administration of downstream Trp metabolite 3-HAA, inhibits Th1/Th17 effector cells and promotes Treg response by up-regulating TGF-ß production by dendritic cells, thereby improving EAE. Collectively, these observations stand out the significance of AA catabolism in the regulation of the immune responses in MS patients. The molecules related to these pathways deserve further exploration as potential new therapeutic targets in MS.

Transcriptome Meta-Analysis Deciphers a Dysregulation in Immune Response-Associated Gene Signatures during Sepsis.

Sepsis is a life-threatening disease induced by a systemic inflammatory response, which leads to organ dysfunction and mortality. In sepsis, the host immune response is depressed and unable to cope with infection; no drug is currently available to treat this. The lungs are frequently the starting point for sepsis. This study aimed to identify potential genes for diagnostics and therapeutic purposes in sepsis by a comprehensive bioinformatics analysis. Our criteria are to unravel sepsis-associated signature genes from gene expression datasets. Differentially expressed genes (DEGs) were identified from samples of sepsis patients using a meta-analysis and then further subjected to functional enrichment and protein‒protein interaction (PPI) network analysis for examining their potential functions. Finally, the expression of the topmost upregulated genes (ARG1, IL1R2, ELANE, MMP9) was quantified by reverse transcriptase-PCR (RT-PCR), and myeloperoxidase (MPO) expression was confirmed by immunohistochemistry (IHC) staining in the lungs of a well-established sepsis mouse model. We found that all the four genes were upregulated in semiquantitative RT-PCR studies; however, MMP9 showed a nonsignificant increase in expression. MPO staining showed strong immunoreactivity in sepsis as compared to the control. This study demonstrates the role of significant and widespread immune activation (IL1R2, MMP9), along with oxidative stress (ARG1) and the recruitment of neutrophils, in sepsis (ELANE, MPO).