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OBJECTIVE - Alternatively activated macrophages (M2) are associated with the progression of spasmolytic polypeptide-expressing metaplasia (SPEM) in the stomach. However, the precise mechanism(s) and critical mediators that induce SPEM are unknown.
DESIGN - To determine candidate genes important in these processes, macrophages from the stomach corpus of mice with SPEM (DMP-777-treated) or advanced SPEM (L635-treated) were isolated and RNA sequenced. Effects on metaplasia development after acute parietal cell loss induced by L635 were evaluated in interleukin (IL)-33, IL-33 receptor (ST2) and IL-13 knockout (KO) mice.
RESULTS - Profiling of metaplasia-associated macrophages in the stomach identified an M2a-polarised macrophage population. Expression of IL-33 was significantly upregulated in macrophages associated with advanced SPEM. L635 induced metaplasia in the stomachs of wild-type mice, but not in the stomachs of IL-33 and ST2 KO mice. While IL-5 and IL-9 were not required for metaplasia induction, IL-13 KO mice did not develop metaplasia in response to L635. Administration of IL-13 to ST2 KO mice re-established the induction of metaplasia following acute parietal cell loss.
CONCLUSIONS - Metaplasia induction and macrophage polarisation after parietal cell loss is coordinated through a cytokine signalling network of IL-33 and IL-13, linking a combined response to injury by both intrinsic mucosal mechanisms and infiltrating M2 macrophages.
Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.
BACKGROUND & AIMS - There is controversy regarding the role of the type 2 immune response in the pathogenesis of ulcerative colitis (UC)-few data are available from treatment-naive patients. We investigated whether genes associated with a type 2 immune response in the intestinal mucosa are up-regulated in treatment-naive pediatric patients with UC compared with patients with Crohn's disease (CD)-associated colitis or without inflammatory bowel disease (IBD), and whether expression levels are associated with clinical outcomes.
METHODS - We used a real-time reverse-transcription quantitative polymerase chain reaction array to analyze messenger RNA (mRNA) expression patterns in rectal mucosal samples from 138 treatment-naive pediatric patients with IBD and macroscopic rectal disease, as well as those from 49 children without IBD (controls), enrolled in a multicenter prospective observational study from 2008 to 2012. Results were validated in real-time reverse-transcription quantitative polymerase chain reaction analyses of rectal RNA from an independent cohort of 34 pediatric patients with IBD and macroscopic rectal disease and 17 controls from Cincinnati Children's Hospital Medical Center.
RESULTS - We measured significant increases in mRNAs associated with a type 2 immune response (interleukin [IL]5 gene, IL13, and IL13RA2) and a type 17 immune response (IL17A and IL23) in mucosal samples from patients with UC compared with patients with colon-only CD. In a regression model, increased expression of IL5 and IL17A mRNAs distinguished patients with UC from patients with colon-only CD (P = .001; area under the receiver operating characteristic curve, 0.72). We identified a gene expression pattern in rectal tissues of patients with UC, characterized by detection of IL13 mRNA, that predicted clinical response to therapy after 6 months (odds ratio [OR], 6.469; 95% confidence interval [CI], 1.553-26.94), clinical response after 12 months (OR, 6.125; 95% CI, 1.330-28.22), and remission after 12 months (OR, 5.333; 95% CI, 1.132-25.12).
CONCLUSIONS - In an analysis of rectal tissues from treatment-naive pediatric patients with IBD, we observed activation of a type 2 immune response during the early course of UC. We were able to distinguish patients with UC from those with colon-only CD based on increased mucosal expression of genes that mediate type 2 and type 17 immune responses. Increased expression at diagnosis of genes that mediate a type 2 immune response is associated with response to therapy and remission in pediatric patients with UC.
Copyright © 2017 AGA Institute. Published by Elsevier Inc. All rights reserved.
Cytokines IL-4 and IL-13 play important roles in polarization of macrophages/dendritic cells to an M2 phenotype, which is important for recovery from acute kidney injury. Both IL-4 and IL-13 activate JAK3/STAT6 signaling. In mice with diphtheria toxin receptor expression in proximal tubules (selective injury model), a relatively selective JAK3 inhibitor, tofacitinib, led to more severe kidney injury, delayed recovery from acute kidney injury, increased inflammatory M1 phenotype markers and decreased reparative M2 phenotype markers of macrophages/dendritic cells, and development of more severe renal fibrosis after diphtheria toxin administration. Similarly, there was delayed recovery and increased tubulointerstitial fibrosis in these diphtheria toxin-treated mice following tamoxifen-induced deletion of both IL-4 and IL-13, with increased levels of M1 and decreased levels of M2 markers in the macrophages/dendritic cells. Furthermore, deletion of IL-4 and IL-13 led to a decrease of tissue reparative M2a phenotype markers but had no effect on anti-inflammatory M2c phenotype markers. Deletion of IL-4 and IL-13 also inhibited recovery from ischemia-reperfusion injury in association with increased M1 and decreased M2 markers and promoted subsequent tubulointerstitial fibrosis. Thus, IL-4 and IL-13 are required to effectively polarize macrophages/dendritic cells to an M2a phenotype and to promote recovery from acute kidney injury.
Copyright © 2016 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.
Allergic airway diseases are immune disorders associated with heightened type 2 immune responses and IL-5 and IL-13 production at the site of inflammation. We have previously reported that cyclooxygenase (COX) inhibition by indomethacin augmented allergic airway inflammation in a STAT6-independent manner. However, the key COX product(s) responsible for restraining indomethacin-mediated STAT6-independent allergic inflammation is unknown. In this study, using the mouse model of OVA-induced allergic airway inflammation, we identified that PGI2 receptor (IP) signaling was critical for indomethacin-induced, STAT6-independent proallergic effects. We demonstrated that IP deficiency increased inflammatory cell infiltration, eosinophilia, and IL-5 and IL-13 expression in the lung in a STAT6-independent manner. The augmented STAT6-independent allergic inflammation correlated with enhanced primary immune responses to allergic sensitization and elevated production of multiple inflammatory chemokines (CCL11, CCL17, CCL22, and CXCL12) in the lung after allergen challenge. We also showed that the PGI2 analogue cicaprost inhibited CD4 T cell proliferation and IL-5 and IL-13 expression in vitro, and IP deficiency diminished the stimulatory effect of indomethacin on STAT6-independent IL-5 and IL-13 responses in vivo. The inhibitory effects of PGI2 and the IP signaling pathway on CD4 T cell activation, inflammatory chemokine production, and allergic sensitization and airway inflammation suggest that PGI2 and its analogue iloprost, both Food and Drug Administration-approved drugs, may be useful in treating allergic diseases and asthma. In addition, inhibiting PGI2 signaling by drugs that either block PGI2 production or restrain IP signaling may augment STAT6-independent pathways of allergic inflammation.
Copyright © 2016 by The American Association of Immunologists, Inc.
Adipose tissue (AT) inflammation during obesity is mediated by immune cells and closely correlates with systemic insulin resistance. In lean AT, eosinophils are present in low but significant numbers and capable of promoting alternative macrophage activation in an IL-4/IL-13-dependent manner. In WT mice, obesity causes the proportion of AT eosinophils to decline, concomitant with inflammation and classical activation of AT macrophages. In this study, we show that CCR2 deficiency leads to increased eosinophil accumulation in AT. Furthermore, in contrast to WT mice, the increase in eosinophils in CCR2(-/-) AT is sustained and even amplified during obesity. Interestingly, a significant portion of eosinophils is found in CLSs in AT of obese CCR2(-/-) mice, which is the first time eosinophils have been shown to localize to these inflammatory hot spots. CCR2(-/-) bone marrow precursors displayed increased expression of various key eosinophil genes during in vitro differentiation to eosinophils, suggesting a potentially altered eosinophil phenotype in the absence of CCR2. In addition, the proportion of eosinophils in AT positively correlated with local expression of Il5, a potent eosinophil stimulator. The increase in eosinophils in CCR2(-/-) mice was detected in all white fat pads analyzed and in the peritoneal cavity but not in bone marrow, blood, spleen, or liver. In AT of CCR2(-/-) mice, an increased eosinophil number positively correlated with M2-like macrophages, expression of the Treg marker Foxp3, and type 2 cytokines, Il4, Il5, and Il13. This is the first study to link CCR2 function with regulation of AT eosinophil accumulation.
© Society for Leukocyte Biology.
INTRODUCTION - Parenteral nutrition (PN) increases the risk of infection in critically ill patients and is associated with defects in gastrointestinal innate immunity. Goblet cells produce mucosal defense compounds, including mucin (principally MUC2), trefoil factor 3 (TFF3), and resistin-like molecule β (RELMβ). Bombesin (BBS), a gastrin-releasing peptide analogue, experimentally reverses PN-induced defects in Paneth cell innate immunity. We hypothesized that PN reduces goblet cell product expression and PN+BBS would reverse these PN-induced defects.
METHODS - Two days after intravenous cannulation, male Institute of Cancer Research mice were randomized to chow (n = 15), PN (n = 13), or PN+BBS (15 µg tid) (n = 12) diets for 5 days. Defined segments of ileum and luminal fluid were analyzed for MUC2, TFF3, and RELMβ by quantitative reverse transcriptase polymerase chain reaction and Western blot. Th2 cytokines interleukin (IL)-4 and IL-13 were measured by enzyme-linked immunosorbent assay.
RESULTS - Compared with chow, PN significantly reduced MUC2 in ileum (P < .01) and luminal fluid (P = .01). BBS supplementation did not improve ileal or luminal MUC2 compared with PN (P > .3). Compared with chow, PN significantly reduced TFF3 in ileum (P < .02) and luminal fluid (P < .01). BBS addition did not improve ileal or luminal TFF3 compared with PN (P > .3). Compared with chow, PN significantly reduced ileal RELMβ (P < .01). BBS supplementation significantly increased ileal RELMβ to levels similar to chow (P < .03 vs PN; P > .6 vs chow). Th2 cytokines were decreased with PN and returned to chow levels with BBS.
CONCLUSION - PN significantly impairs the goblet cell component of innate mucosal immunity. BBS only preserves goblet cell RELMβ during PN but not other goblet cell products measured.
© 2015 American Society for Parenteral and Enteral Nutrition.
Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, a multienzyme complex, is the major source for production of reactive oxygen species (ROS). ROS are increased in allergic diseases, such as asthma, but the role of ROS in disease pathogenesis remains uncertain. We hypothesized that mice unable to generate ROS via the NADPH oxidase pathway would have decreased allergic airway inflammation. To test this hypothesis, we studied gp91phox(-/-) mice in a model of allergic airway inflammation after sensitization and challenge with ovalbumin. Serum, bronchoalveolar lavage fluid, and lungs were then examined for evidence of allergic inflammation. We found that mice lacking a functional NADPH oxidase complex had significantly decreased ROS production and allergic airway inflammation, compared with wild-type (WT) control animals. To determine the mechanism by which allergic inflammation was inhibited by gp91phox deficiency, we cultured bone marrow-derived dendritic cells from WT and gp91phox(-/-) mice and activated them with LPS. IL-12 expression was significantly increased in the gp91phox(-/-) bone marrow-derived dendritic cells, suggesting that the cytokine profile produced in the absence of gp91phox enhanced the conditions leading to T helper (Th) type 1 differentiation, while inhibiting Th2 polarization. Splenocytes from sensitized gp91phox(-/-) animals produced significantly less IL-13 in response to ovalbumin challenge in vitro compared with splenocytes from sensitized WT mice, suggesting that NADPH oxidase promotes allergic sensitization. In contrast, inflammatory cytokines produced by T cells cultured from WT and gp91phox(-/-) mice under Th0, Th1, Th2, and Th17 conditions were not significantly different. This study demonstrates the importance of NADPH oxidase activity and ROS production in a murine model of asthma.
Mucous cell metaplasia is a hallmark of asthma, and may be mediated by signal transducers and activators of transcription (STAT)-6 signaling. IL-17A is increased in the bronchoalveolar lavage fluid of patients with severe asthma, and IL-17A also increases mucus production in airway epithelial cells. Asthma therapeutics are being developed that inhibit STAT6 signaling, but the role of IL-17A in inducing mucus production in the absence of STAT6 remains unknown. We hypothesized that IL-17A induces mucous cell metaplasia independent of STAT6, and we tested this hypothesis in two murine models in which increased IL-17A protein expression is evident. In the first model, ovalbumin (OVA)-specific D011.10 Th17 cells were adoptively transferred into wild-type (WT) or STAT6 knockout (KO) mice, and the mice were challenged with OVA or PBS. WT-OVA and STAT6 KO-OVA mice demonstrated increased airway IL-17A and IL-13 protein expression and mucous cell metaplasia, compared with WT-PBS or STAT6 KO-PBS mice. In the second model, WT, STAT1 KO, STAT1/STAT6 double KO (DKO), or STAT1/STAT6/IL-17 receptor A (RA) triple KO (TKO) mice were challenged with respiratory syncytial virus (RSV) or mock viral preparation, and the mucous cells were assessed. STAT1 KO-RSV mice demonstrated increased airway mucous cell metaplasia compared with WT-RSV mice. STAT1 KO-RSV and STAT1/STAT6 DKO-RSV mice also demonstrated increased mucous cell metaplasia, compared with STAT1/STAT6/IL17RA TKO-RSV mice. We also treated primary murine tracheal epithelial cells (mTECs) from WT and STAT6 KO mice. STAT6 KO mTECs showed increased periodic acid-Schiff staining with IL-17A but not with IL-13. Thus, asthma therapies targeting STAT6 may increase IL-17A protein expression, without preventing IL-17A-induced mucus production.
The Th2 cytokines interleukin (IL)-4 and -13 are acknowledged regulators of lymphocyte proliferation and activation. They have also been well studied in the regulation of various myeloid-derived populations in tumor biology. It has become clear, however, that both cytokines can have direct effects on epithelial tumor cells expressing appropriate receptors. Changes in tumor proliferation, survival, and metastatic capability have all been ascribed to IL-4 and/or IL-13 action. Here, we evaluate the evidence to support direct tumor-promoting roles of these cytokines. We also identify the questions that should be addressed before proceeding with therapeutic approaches based on neutralization of IL-4 or IL-13 pathways.
BACKGROUND - Interleukin 13 (IL-13) is upregulated in ulcerative colitis (UC) and increases colon epithelial permeability by inducing apoptosis and expression of the pore-forming tight junction protein claudin-2. IL-13 induces activation of signal transducer and activator of transcription 6 (STAT6). However, the STAT6 phosphorylation status in patients with UC is unknown, as is the effect of STAT6 inhibition on colonic epithelium exposed to IL-13. The study aims were to determine if mucosal STAT6 phosphorylation is increased in patients with UC, and if STAT6 inhibition attenuates IL-13-induced colon epithelial cell dysfunction.
METHODS - Immunohistochemical staining for phosphorylated (p) STAT6 was performed on colonic tissue from newly diagnosed pediatric subjects with UC (early UC) or Crohn's disease (CD), colectomy tissue from adults with UC (advanced UC), and controls. Colon HT-29 and T84 cells were transfected with STAT6 small interfering RNA (siRNA), or treated with suberoylanilide hydroxamic acid (SAHA), a histone deacetylase inhibitor that inhibits STAT6, prior to IL-13 treatment.
RESULTS - The median score for epithelial pSTAT6 was 0 in control subjects, 2 in early UC (versus control P = 0.019), 4 in advanced UC (P = 0.003), and 0 in CD (P = 0.4). Cell transfection with STAT6 siRNA prevented IL-13-induced apoptosis and claudin-2 expression. SAHA inhibited IL-13-induced STAT6 phosphorylation, apoptosis, and claudin-2 expression, and mitigated IL-13-induced reductions in transepithelial resistance.
CONCLUSIONS - UC is associated with increased colonic epithelial STAT6 phosphorylation, and STAT6 inhibition prevents IL-13-induced apoptosis and barrier disruption. These data identify STAT6 as a novel target for UC treatment and support further study of SAHA as a therapeutic agent.
Copyright © 2011 Crohn's & Colitis Foundation of America, Inc.