Interleukin-13 Propagates Prothrombin Kringle-2-Induced Neurotoxicity in Hippocampi In Vivo via Oxidative Stress.

The present study investigated expression of endogenous interleukin-13 (IL-13) and its possible function in the hippocampus of prothrombin kringle-2 (pKr-2)-lesioned rats. Here we report that intrahippocampal injection of pKr-2 revealed a significant loss of NeuN-immunopositive (NeuN+) and Nissl+ cells in the hippocampus at 7 days after pKr-2. In parallel, pKr-2 increased IL-13 levels, which reached a peak at 3 days post pKr-2 and sustained up to 7 days post pKr-2. IL-13 immunoreactivity was seen exclusively in activated microglia/macrophages and neutrophils, but not in neurons or astrocytes. In experiments designed to explore the involvement of IL-13 in neurodegeneration, IL-13 neutralizing antibody (IL-13Nab) significantly increased survival of NeuN+ and Nissl+ cells. Accompanying neuroprotection, immunohistochemical analysis indicated that IL-13Nab inhibited pKr-2-induced expression of inducible nitric oxide synthase and myeloperoxidase within activated microglia/macrophages and neutrophils, possibly resulting in attenuation of reactive oxygen species (ROS) generation and oxidative damage of DNA and protein. The current findings suggest that the endogenous IL-13 expressed in pKr-2 activated microglia/macrophages and neutrophils might be harmful to hippocampal neurons via oxidative stress.

Critical role of IL-33, but not IL-25 or TSLP, in silica crystal-mediated exacerbation of allergic airway eosinophilia.

Silica crystals (silica), which are a major mineral component of volcanic ash and desert dust, contribute to the pathogenesis of pulmonary disorders such as asthma and fibrosis. Although administration of silica or sand dust to rodents exacerbates development of ovalbumin-induced or house dust mite-induced asthma-like airway inflammation, the detailed mechanisms remain unclear. Here, using murine models, we found that silica can induce IL-33 expression in pulmonary epithelial cells. IL-33, but not IL-25 or TSLP, and type 2 cytokines such as IL-5 and IL-13 were critically involved in silica's exacerbation of OVA-induced airway eosinophilia in mice. Innate lymphoid cells (ILCs), but not T, B or NKT cells, were also involved in the setting. Moreover, a scavenger receptor that recognized silica was important for silica's exacerbating effect. These observations suggest that IL-33 induced in epithelial cells by silica activates ILCs to produce IL-5 and/or IL-13, contributing to silica's exacerbation of OVA-induced airway eosinophilia in mice. Our findings provide new insight into the underlying mechanisms of exacerbation of pulmonary disorders such as asthma following inhalation of silica-containing materials such as volcanic ash and desert dust.

Efficient RNP-directed Human Gene Targeting Reveals SPDEF Is Required for IL-13-induced Mucostasis.

Primary human bronchial epithelial cell (HBEC) cultures are a useful model for studies of lung health and major airway diseases. However, mechanistic studies have been limited by our ability to selectively disrupt specific genes in these cells. Here we optimize methods for gene targeting in HBECs by direct delivery of single guide RNA (sgRNA) and rCas9 (recombinant Cas9) complexes by electroporation, without a requirement for plasmids, viruses, or antibiotic selection. Variations in the method of delivery, sgRNA and rCas9 concentrations, and sgRNA sequences all had effects on targeting efficiency, allowing for predictable control of the extent of gene targeting and for near-complete disruption of gene expression. To demonstrate the value of this system, we targeted SPDEF, which encodes a transcription factor previously shown to be essential for the differentiation of MUC5AC-producing goblet cells in mouse models of asthma. Targeting SPDEF led to proportional decreases in MUC5AC expression in HBECs stimulated with IL-13, a central mediator of allergic asthma. Near-complete targeting of SPDEF abolished IL-13-induced MUC5AC expression and goblet cell differentiation. In addition, targeting of SPDEF prevented IL-13-induced impairment of mucociliary clearance, which is likely to be an important contributor to airway obstruction, morbidity, and mortality in asthma. We conclude that direct delivery of sgRNA and rCas9 complexes allows for predictable and efficient gene targeting and enables mechanistic studies of disease-relevant pathways in primary HBECs.

The role of interleukin-13 in chronic inflammatory intestinal disorders.

Interleukin (IL)-13 is a cytokine playing a pivotal role in T helper (Th)2 immune response supposed to be implicated in some intestinal disorders. IL-13 is produced by Th2 cells, natural killer T cell, innate lymphoid cells and innate immune cells, which contribute to trigger and maintain a chronic idiopathic intestinal inflammation. In murine models IL-13 exerts pleiotropic functions, playing either pathogenic or protective roles according to the different experimental conditions. As regards celiac disease, IL-13 is considered to be involved mostly in the refractory phase rather than at uncomplicated stage. Discrepancies have been observed in the role of IL-13 upon the inflammation and fibrosis in ulcerative colitis (UC) and in Crohn's disease, respectively. Failure of the anti-IL-13 monoclonal antibodies tralokinumab and anrukinzumab in UC patients in clinical trials support the absence of a role for IL-13 in UC. This review deals with IL-13 in several experimental colitis models -such as oxazolone-, trinitrobenzene sulfonic acid- or dextran sodium sulphate-induced colitis- and chronic intestinal inflammatory disorders -including celiac disease, UC and Crohn's disease-, and it also highlights the attempts to modulate IL-13 as therapeutic tool.

IL-4 receptor engagement in human neutrophils impairs their migration and extracellular trap formation.

BACKGROUND: Type 2 immunity serves to resist parasitic helminths, venoms, and toxins, but the role and regulation of neutrophils during type 2 immune responses are controversial. Helminth models suggested a contribution of neutrophils to type 2 immunity, whereas neutrophils are associated with increased disease severity during type 2 inflammatory disorders, such as asthma. OBJECTIVE: We sought to evaluate the effect of the prototypic type 2 cytokines IL-4 and IL-13 on human neutrophils. METHODS: Human neutrophils from peripheral blood were assessed without or with IL-4 or IL-13 for (1) expression of IL-4 receptor subunits, (2) neutrophil extracellular trap (NET) formation, (3) migration toward CXCL8 in vitro and in humanized mice, and (4) CXCR1, CXCR2, and CXCR4 expression, as well as (5) in nonallergic versus allergic subjects. RESULTS: Human neutrophils expressed both types of IL-4 receptors, and their stimulation through IL-4 or IL-13 diminished their ability to form NETs and migrate toward CXCL8 in vitro. Likewise, in vivo chemotaxis in NOD-scid-Il2rg-/- mice was reduced in IL-4-stimulated human neutrophils compared with control values. These effects were accompanied by downregulation of the CXCL8-binding chemokine receptors CXCR1 and CXCR2 on human neutrophils on IL-4 or IL-13 stimulation in vitro. Ex vivo analysis of neutrophils from allergic patients or exposure of neutrophils from nonallergic subjects to allergic donor serum in vitro impaired their NET formation and migration toward CXCL8, thereby mirroring IL-4/IL-13-stimulated neutrophils. CONCLUSION: IL-4 receptor signaling in human neutrophils affects several neutrophil effector functions, which bears important implications for immunity in type 2 inflammatory disorders.

Association of IL-13, S100B, and TLR-7 Gene Polymorphisms with Enterovirus 71 Infection in Hand, Foot, and Mouth Disease in China.

AIM: This study was conducted to determine if single nucleotide polymorphisms within the interleukin (IL)-13 (rs20541 locus), the S100B (rs9722 locus), and the toll-like receptor (TLR)-7 (rs179019 and rs3853839 loci) genes are associated with the clinical severity of disease caused by enterovirus 71 (EV71) in children suffering from hand, foot, and mouth disease (HFMD). MATERIALS AND METHODS: A total of 355 children, diagnosed with HFMD, were divided into two groups: severe (totaling 162 cases) and mild (totaling 193 cases). Three hundred healthy children were recruited as a control group. The gene polymorphisms of the rs20541 locus in the IL-13 gene; the rs9722 locus in the S100B gene; and the rs179019 and the rs3853839 loci in the TLR-7 gene were analyzed with Sanger sequencing. The expression levels of IL-13, S100B, interferon (IFN)-α, IL-6 and the relative expression level of TLR-7 were calculated for each genotype. RESULTS: This study demonstrated that the T allele at the rs9722 locus of the S100B gene was a significant risk factor for severe HFMD. The rs3853839 C allele of the TLR-7 gene was also a risk factor for severe HFMD in both male and female patients. The G allele at the rs20541 locus of IL-13 gene and the A allele at the rs179019 locus of the TLR-7 gene were not risk factors for severe HFMD in either male or female patients. CONCLUSION: The T allele at the rs9722 locus of S100B gene is a risk factor for the severe HFMD caused by EV71 infection, of which the mechanism may be due to the promotion of S100B protein secretion. The allele C at TLR-7 rs3853839 locus is a risk factor for the severe HFMD caused by EV71 infection, which may be related to a reduction of the relative expression of TLR-7, IFN-α, and IL-6.

Th2 cytokines orchestrate the secretion of MUC5AC and MUC5B in IL-5-positive chronic rhinosinusitis with nasal polyps.

BACKGROUND: Mucin over-secretion is a significant characteristic of chronic rhinosinusitis with nasal polyps (CRSwNP). This study aimed to investigate the relationship between Th2 cytokines and MUC5AC or MUC5B, and the mechanism of mucin over-secretion in the type-2 inflammatory endotype of CRSwNP. METHODS: Main Th-cell cytokines, associated mediators, and mucins were determined in the homogenates of nasal polyp samples from 21 CRSwNP patients and inferior turbinate samples from 8 controls, by ELISA or UniCAP system. Secretion of MUC5AC and MUC5B was measured in the supernatants of IL-5, IL-4, or IL-13 primed nasal polyp fragments. Co-localization of MUC5AC, MUC5B, and IL-4 receptor α (IL-4Rα) in CRSwNP and controls was evaluated by immunohistochemistry. Gene expression of IL-4Rα in the samples was measured by real-time reverse transcription-polymerase chain reaction. RESULTS: Baseline protein levels of the Th2-cytokines IL-4, IL-5, and IL-13, and mucins MUC5AC and MUC5B were significantly higher in the IL-5(+) CRSwNP group, compared to control and IL-5(-) CRSwNP groups. MUC5AC and MUC5B secretions were significantly increased in IL-4- or IL-13-primed, but not IL-5-primed fragments of nasal polyps. Immuno-stained serial sections demonstrated that IL-4Rα was widely expressed in the epithelium and submucosal glands in control and nasal polyp tissues. Gene expression of IL-4Rα was elevated in nasal polyp tissues, specifically in the IL-5(+) CRSwNP group. CONCLUSIONS: In type-2 inflammatory nasal polyps, characterized by the tissue expression of IL-5, MUC5AC and MUC5B are overexpressed. Both IL-4 and IL-13 may upregulate mucin expression via IL-4Rα, which is also overexpressed in IL-5(+) CRSwNP.

Targeting IL-13 as a Host-Directed Therapy Against Ulcerative Colitis.

The role of interleukin-13 in mediating ulcerative colitis remains under scrutiny. Compelling evidence from both man and mouse suggests that IL-13 not only contributes to the pathology associated with disease but is also involved in mediating the inflammatory response. These studies have led to the approach of targeting IL-13 as a promising treatment strategy in alleviating ulcerative colitis disease. Despite this evidence, recent clinical trial data suggests that specifically blocking the receptor through which IL-13 signals, IL-4 receptor-alpha (IL-4Rα) in ulcerative colitis patients, is insufficient in protecting them from disease outcome. This challenges the importance of IL-13 as a therapeutic target. This review describes the role of IL-13 in ulcerative colitis and current treatment strategies that target IL-13. The potential role of IL-13 signaling independently of IL-4Rα in mediating ulcerative colitis is highlighted as an important consideration when targeting the signaling mechanisms of IL-13 for therapeutic approaches.

Transgenic Mouse Reporter to Study Muc5b In Vivo.

Dysregulation of gel-forming mucins is associated with many airway diseases. Better knowledge of the pathophysiological mechanisms linking mucins and respiratory diseases will advance the understanding of their pathogenesis and should provide opportunities to develop new therapeutic compounds for treatment. MUC5B and MUC5AC are the two main gel-forming mucins in the respiratory tract. The organization in domains and the expression profile of mouse Muc5b are very similar to those in humans, which makes the mouse a relevant model for studies of the translational activities of human mucins. To assess the in vivo biological functions of Muc5b, a mouse reporter tagged in frame with the green fluorescent protein marker has been engineered by homologous recombination. The proof of concept that this reporter model may be informative for translational studies was confirmed by the finding that interleukin-13 administration in living mice upregulated Muc5b production.

Regulation of monoamine oxidase A (MAO-A) expression, activity, and function in IL-13-stimulated monocytes and A549 lung carcinoma cells.

Monoamine oxidase A (MAO-A) is a mitochondrial flavoenzyme implicated in the pathogenesis of atherosclerosis and inflammation and also in many neurological disorders. MAO-A also has been reported as a potential therapeutic target in prostate cancer. However, the regulatory mechanisms controlling cytokine-induced MAO-A expression in immune or cancer cells remain to be identified. Here, we show that MAO-A expression is co-induced with 15-lipoxygenase (15-LO) in interleukin 13 (IL-13)-activated primary human monocytes and A549 non-small cell lung carcinoma cells. We present evidence that MAO-A gene expression and activity are regulated by signal transducer and activator of transcription 1, 3, and 6 (STAT1, STAT3, and STAT6), early growth response 1 (EGR1), and cAMP-responsive element-binding protein (CREB), the same transcription factors that control IL-13-dependent 15-LO expression. We further established that in both primary monocytes and in A549 cells, IL-13-stimulated MAO-A expression, activity, and function are directly governed by 15-LO. In contrast, IL-13-driven expression and activity of MAO-A was 15-LO-independent in U937 promonocytic cells. Furthermore, we demonstrate that the 15-LO-dependent transcriptional regulation of MAO-A in response to IL-13 stimulation in monocytes and in A549 cells is mediated by peroxisome proliferator-activated receptor γ (PPARγ) and that signal transducer and activator of transcription 6 (STAT6) plays a crucial role in facilitating the transcriptional activity of PPARγ. We further report that the IL-13-STAT6-15-LO-PPARγ axis is critical for MAO-A expression, activity, and function, including migration and reactive oxygen species generation. Altogether, these results have major implications for the resolution of inflammation and indicate that MAO-A may promote metastatic potential in lung cancer cells.

IL-13 -1112 polymorphism and periodontitis susceptibility: a meta-analysis.

BACKGROUND: Several studies have examined the association between the IL-13 -1112C/T polymorphism and the risk of periodontitis. However, these studies have reached different conclusions. The aim of the current study was to investigate the link between this IL-13 -1112 polymorphism and susceptibility to periodontitis. METHODS: We utilized electronic databases, including the CNKI (China National Knowledge Infrastructure), Wanfang, PubMed, Embase, and Cochrane Library databases, to manually search for relevant research published through November 30, 2016. The Chinese and English terms used to search the literature included "periodontitis", "periodontal disease", "IL 13", "IL-13", and "interleukin-13". In accordance with our inclusion criteria, we selected studies that involved case-control trials. All of these case-control trials described their objectives, design and specific statistical methods. For all included studies, odds ratios (ORs) and 95% confidence intervals (95% CIs) were provided or could be calculated from the study data. The quality of the included literature was evaluated using the Newcastle-Ottawa scale (NOS). STATA 12.0 was used to calculate the sizes of the combined effects and conduct a sensitivity analysis of the results. RESULTS: Our meta-analysis included 4 articles representing 5 case-control studies with a total of 710 cases and 671 control subjects. The meta-analysis results indicated that the CC vs TT model, CT vs TT model and TT vs CT + CC model (CC VS TT: OR = 0.615, 95% CI = 0.395-0.957; CT vs TT: OR = 0.518, 95% CI = 0.323-0.830; and TT vs CT + CC: OR = 1.739, 95% CI = 1.130-2.676) were significant in five IL-13 -1112 gene polymorphism and periodontitis susceptibility models. Subgroup analysis indicated that the CC vs TT, CT vs TT and TT vs CT + CC models were significant in the chronic periodontitis (CP) group, whereas no significant differences were found in the five aggressive periodontitis (AgP) group models. The sensitivity analysis showed that dropping any single study did not affect the pooled analysis results. CONCLUSION: The IL-13 -1112 polymorphism may be associated with susceptibility to periodontitis. The IL-13 -1112 gene polymorphism may be associated with susceptibility to CP but not to AgP. Thus, large-scale, multi-ethnic case-control trials are still warranted.

[Pathogenesis of atopic dermatitis]. / Pathogenese des atopischen Ekzems.

Atopic dermatitis (AD) is one of the most common chronic inflammatory diseases and is also one of the most frequent reasons to consult a dermatologist. Over the past few years there has been a rapidly growing understanding of the cellular, molecular and immunological relationships as well as genetic variations, which leads to a better comprehension of the disease. Consequently, there are innovative targeted therapies in clinical studies or already approved for therapy. To make reasonable use of the new targeted therapies a good understanding of the pathogenesis is very important. In the future, stratification of patients with AD and the resulting personalized therapies will gain in importance. This review depicts the up to date state of knowledge on the complex pathogenesis of AD.

Cell Biology of Tight Junction Barrier Regulation and Mucosal Disease.

Mucosal surfaces are lined by epithelial cells. In the intestine, the epithelium establishes a selectively permeable barrier that supports nutrient absorption and waste secretion while preventing intrusion by luminal materials. Intestinal epithelia therefore play a central role in regulating interactions between the mucosal immune system and luminal contents, which include dietary antigens, a diverse intestinal microbiome, and pathogens. The paracellular space is sealed by the tight junction, which is maintained by a complex network of protein interactions. Tight junction dysfunction has been linked to a variety of local and systemic diseases. Two molecularly and biophysically distinct pathways across the intestinal tight junction are selectively and differentially regulated by inflammatory stimuli. This review discusses the mechanisms underlying these events, their impact on disease, and the potential of using these as paradigms for development of tight junction-targeted therapeutic interventions.

Anti-inflammatory (M2) macrophage media reduce transmission of oligomeric amyloid beta in differentiated SH-SY5Y cells.

Neuroinflammation plays an influential role in Alzheimer's disease (AD), although the mechanisms underlying this phenomenon remain largely unknown. Microglia are thought to be responsible for the majority of these effects and can be characterized into resting (M0), proinflammatory (M1), or anti-inflammatory (M2) functional phenotypes. We investigated the effects of conditioned macrophage media, as an analogue to microglia, on the transfer of oligomeric amyloid beta (oAß) between differentiated SH-SY5Y cells. We also investigated how the different inflammatory environments related to intercellular and intracellular changes. We demonstrate that M2 products decrease interneuronal transfer of oAß, while recombinant interleukin (IL)-4, IL-10, and IL-13 increase transfer. There were no alterations to the mRNA of a number of AD-related genes in response to the combination of oAß and M0, M1, or M2, but several intracellular proteins, some relating to protein trafficking and the endosomal/lysosomal system, were altered. Stimulating microglia to an M2 phenotype may thus slow down the progression of AD and could be a target for future therapies.

Growth Differentiation Factor 15 Mediates Systemic Glucose Regulatory Action of T-Helper Type 2 Cytokines.

T-helper type 2 (Th2) cytokines, including interleukin (IL)-13 and IL-4, produced in adipose tissue, are critical regulators of intra-adipose and systemic lipid and glucose metabolism. Furthermore, IL-13 is a potential therapy for insulin resistance in obese mouse models. Here, we examined mediators produced by adipocytes that are responsible for regulating systemic glucose homeostasis in response to Th2 cytokines. We used RNA sequencing data analysis of cultured adipocytes to screen factors secreted in response to recombinant IL-13. Recombinant IL-13 induced expression of growth differentiation factor 15 (GDF15) via the Janus kinase-activated STAT6 pathway. In vivo administration of α-galactosylceramide or IL-33 increased IL-4 and IL-13 production, thereby increasing GDF15 levels in adipose tissue and in plasma of mice; however, these responses were abrogated in STAT6 knockout mice. Moreover, administration of recombinant IL-13 to wild-type mice fed a high-fat diet (HFD) improved glucose intolerance; this was not the case for GDF15 knockout mice fed the HFD. Taken together, these data suggest that GDF15 is required for IL-13-induced improvement of glucose intolerance in mice fed an HFD. Thus, beneficial effects of Th2 cytokines on systemic glucose metabolism and insulin sensitivity are mediated by GDF15. These findings open up a potential pharmacological route for reversing insulin resistance associated with obesity.

ILC2s regulate adaptive Th2 cell functions via PD-L1 checkpoint control.

Group 2 innate lymphoid cells (ILC2s) are important effector cells driving the initiation of type 2 immune responses leading to adaptive T helper 2 (Th2) immunity. Here we show that ILC2s dynamically express the checkpoint inhibitor molecule PD-L1 during type 2 pulmonary responses. Surprisingly, PD-L1:PD-1 interaction between ILC2s and CD4+ T cells did not inhibit the T cell response, but PD-L1-expressing ILC2s stimulated increased expression of GATA3 and production of IL-13 by Th2 cells both in vitro and in vivo. Conditional deletion of PD-L1 on ILC2s impaired early Th2 polarization and cytokine production, leading to delayed worm expulsion during infection with the gastrointestinal helminth Nippostrongylus brasiliensis Our results identify a novel PD-L1-controlled mechanism for type 2 polarization, with ILC2s mediating an innate checkpoint to control adaptive T helper responses, which has important implications for the treatment of type 2 inflammation.

A type 2 cytokine axis for thymus emigration.

In the thymus, stromal microenvironments support a developmental program that generates mature T cells ready for thymic exit. The cellular and molecular specialization within thymic stromal cells that enables their regulation of specific stages of thymocyte development is poorly understood. Here, we show the thymic microenvironment expresses the type 2 IL-4R complex and is functionally responsive to its known ligands, IL-4 and IL-13. Absence of IL-4Rα limits thymocyte emigration, leading to an intrathymic accumulation of mature thymocytes within medullary perivascular spaces and reduced numbers of recent thymic emigrants. Thymus transplantation shows this requirement maps to IL-4Rα expression by stromal cells, and we provide evidence that it regulates thymic exit via a process distinct from S1P-mediated migration. Finally, we reveal a cellular mechanism by which IL-4+IL-13+ invariant NKT cells are necessary for IL-4Rα signaling that regulates thymic exit. Collectively, we define a new axis for thymic emigration involving stimulation of the thymic microenvironment via type 2 cytokines from innate T cells.

Eosinophilic esophagitis: unclear roles of IgE and eosinophils.

Eosinophilic esophagitis (EoE) is a chronic inflammatory disease of the oesophagus. Recognized as a distinct entity only two decades ago, the emergence of the disease along with the availability of new technologies has rapidly opened new research avenues and outlined the main features of the pathogenesis of EoE. Yet, each advance in our understanding of the disease has raised new questions about the previous consensus. Currently, new subsets of the disease challenge our diagnostic criteria. For instance, it was believed that EoE did not respond to proton pump inhibitor (PPI) therapy; however, it has now been shown that a substantial proportion of EoE patients indeed respond to PPIs. In addition, a new subset of patients not even presenting eosinophil infiltrates in the oesophagus has also been described. Moreover, approaches for better understanding the heritability of the disease bring into question the dogma of predominant genetic involvement. Furthermore, the specificity and sensitivity of allergy testing for targeted food avoidance is highly controversial, and the production of specific antibodies in EoE now includes IgG4 in addition to IgE. In conclusion, EoE is perceived as 'a moving target' and the aim of this review was to summarize the current understanding of EoE pathogenesis.