Reciprocal regulation of IL-33 receptor-mediated inflammatory response and pulmonary fibrosis by TRAF6 and USP38.

The warning cytokine interleukin-33 receptor (IL-33R) mediates local inflammatory responses and plays crucial roles in the pathogenesis of immune diseases such as pulmonary fibrosis and rheumatoid arthritis. Whether and how IL-33R is regulated remain enigmatic. Here, we identified ubiquitin-specific protease 38 (USP38) as a negative regulator of IL-33R­mediated signaling. USP38 deficiency promotes interleukin-33 (IL-33)­induced downstream proinflammatory responses in vitro and in vivo. Usp38−/− mice are more susceptible to inflammatory damage and death and developed more serious pulmonary fibrosis after bleomycin treatment. USP38 is constitutively associated with IL-33R and deconjugates its K27-linked polyubiquitination at K511, resulting in its autophagic degradation. We further show that the E3 ubiquitin ligase tumor necrosis factor receptor­associated factor 6 (TRAF6) catalyzes K27-linked polyubiquitination of IL-33R at K511, and that deficiency of TRAF6 inhibits IL-33­mediated signaling. Our findings suggest that K27-linked polyubiquitination and deubiquitination of IL-33R by TRAF6 and USP38 reciprocally regulate IL-33R level and signaling, which represents a critical mechanism in the regulation of IL-33­triggered lung inflammatory response and pulmonary fibrosis.

The Paradigm Change of IL-33 in Vascular Biology.

In this review, we focus on the actual understanding of the role of IL-33 in vascular biology in the context of the historical development since the description of IL-33 as a member of IL-1 superfamily and the ligand for ST2 receptor in 2005. We summarize recent data on the biology, structure and signaling of this dual-function factor with both nuclear and extracellular cytokine properties. We describe cellular sources of IL-33, particularly within vascular wall, changes in its expression in different cardio-vascular conditions and mechanisms of IL-33 release. Additionally, we summarize the regulators of IL-33 expression as well as the effects of IL-33 itself in cells of the vasculature and in monocytes/macrophages in vitro combined with the consequences of IL-33 modulation in models of vascular diseases in vivo. Described in murine atherosclerosis models as well as in macrophages as an atheroprotective cytokine, extracellular IL-33 induces proinflammatory, prothrombotic and proangiogenic activation of human endothelial cells, which are processes known to be involved in the development and progression of atherosclerosis. We, therefore, discuss that IL-33 can possess both protective and harmful effects in experimental models of vascular pathologies depending on experimental conditions, type and dose of administration or method of modulation.

Basal epithelial stem cells cross an alarmin checkpoint for postviral lung disease.

Epithelial cells are charged with protection at barrier sites, but whether this normally beneficial response might sometimes become dysfunctional still needs definition. Here, we recognized a pattern of imbalance marked by basal epithelial cell growth and differentiation that replaced normal airspaces in a mouse model of progressive postviral lung disease due to the Sendai virus. Single-cell and lineage-tracing technologies identified a distinct subset of basal epithelial stem cells (basal ESCs) that extended into gas-exchange tissue to form long-term bronchiolar-alveolar remodeling regions. Moreover, this cell subset was selectively expanded by crossing a cell-growth and survival checkpoint linked to the nuclear-localized alarmin IL-33 that was independent of IL-33 receptor signaling and instead connected to autocrine chromatin accessibility. This mechanism creates an activated stem-progenitor cell lineage with potential for physiological or pathological function. Thus, conditional loss of Il33 gene function in basal epithelial cells disrupted the homeostasis of the epithelial barrier at skin and gut sites but also markedly attenuated postviral disease in the lung based on the downregulation of remodeling and inflammation. Thus, we define a basal ESC strategy to deploy innate immune machinery that appears to overshoot the primordial goal of self-defense. Our findings reveal new targets to stratify and correct chronic and often deadly postviral disease.

Inflammasomes and Fibrosis.

Fibrosis is the final common pathway of inflammatory diseases in various organs. The inflammasomes play an important role in the progression of fibrosis as innate immune receptors. There are four main members of the inflammasomes, such as NOD-like receptor protein 1 (NLRP1), NOD-like receptor protein 3 (NLRP3), NOD-like receptor C4 (NLRC4), and absent in melanoma 2 (AIM2), among which NLRP3 inflammasome is the most studied. NLRP3 inflammasome is typically composed of NLRP3, ASC and pro-caspase-1. The activation of inflammasome involves both "classical" and "non-classical" pathways and the former pathway is better understood. The "classical" activation pathway of inflammasome is that the backbone protein is activated by endogenous/exogenous stimulation, leading to inflammasome assembly. After the formation of "classic" inflammasome, pro-caspase-1 could self-activate. Caspase-1 cleaves cytokine precursors into mature cytokines, which are secreted extracellularly. At present, the "non-classical" activation pathway of inflammasome has not formed a unified model for activation process. This article reviews the role of NLRP1, NLRP3, NLRC4, AIM2 inflammasome, Caspase-1, IL-1ß, IL-18 and IL-33 in the fibrogenesis.

Mechanical Ventilation with Moderate Tidal Volume Exacerbates Extrapulmonary Sepsis-Induced Lung Injury via IL33-WISP1 Signaling Pathway.

ABSTRACT: IL-33 and WNT1-inducible secreted protein (WISP1) play central roles in acute lung injury (ALI) induced by mechanical ventilation with moderate tidal volume (MTV) in the setting of sepsis. Here, we sought to determine the inter-relationship between IL-33 and WISP1 and the associated signaling pathways in this process.We used a two-hit model of cecal ligation puncture (CLP) followed by MTV ventilation (4 h 10 mL/kg) in wild-type, IL-33-/- or ST2-/- mice or wild-type mice treated with intratracheal antibodies to WISP1. Macrophages (Raw 264.7 and alveolar macrophages from wild-type or ST2-/- mice) were used to identify specific signaling components.CLP + MTV resulted in ALI that was partially sensitive to genetic ablation of IL-33 or ST2 or antibody neutralization of WISP1. Genetic ablation of IL-33 or ST2 significantly prevented ALI after CLP + MTV and reduced levels of WISP1 in the circulation and bronchoalveolar lung fluid. rIL-33 increased WISP1 in alveolar macrophages in an ST2, PI3K/AKT, and ERK dependent manner. This WISP1 upregulation and WNT ß-catenin activation were sensitive to inhibition of the ß-catenin/TCF/CBP/P300 nuclear pathway.We show that IL-33 drives WISP1 upregulation and ALI during MTV in CLP sepsis. The identification of this relationship and the associated signaling pathways reveals a number of possible therapeutic targets to prevent ALI in ventilated sepsis patients.

Interleukin 33 Triggers Early Eosinophil-Dependent Events Leading to Metaplasia in a Chronic Model of Gastritis-Prone Mice.

BACKGROUND & AIMS: Interleukin (IL)33/IL1F11 is an important mediator for the development of type 2 T-helper cell (Th2)-driven inflammatory disorders and has also been implicated in the pathogenesis of gastrointestinal (GI)-related cancers, including gastric carcinoma. We therefore sought to mechanistically determine IL33's potential role as a critical factor linking chronic inflammation and gastric carcinogenesis using gastritis-prone SAMP1/YitFc (SAMP) mice. METHODS: SAMP and (parental control) AKR mice were assessed for baseline gastritis and progression to metaplasia. Expression/localization of IL33 and its receptor, ST2/IL1R4, were characterized in corpus tissues, and activation and neutralization studies were both performed targeting the IL33/ST2 axis. Dissection of immune pathways leading to metaplasia was evaluated, including eosinophil depletion studies using anti-IL5/anti-CCR3 treatment. RESULTS: Progressive gastritis and, ultimately, intestinalized spasmolytic polypeptide-expressing metaplasia (SPEM) was detected in SAMP stomachs, which was absent in AKR but could be moderately induced with exogenous, recombinant IL33. Robust peripheral (bone marrow) expansion of eosinophils and local recruitment of both eosinophils and IL33-expressing M2 macrophages into corpus tissues were evident in SAMP. Interestingly, IL33 blockade did not affect bone marrow-derived expansion and local infiltration of eosinophils, but markedly decreased M2 macrophages and SPEM features, while eosinophil depletion caused a significant reduction in both local IL33-producing M2 macrophages and SPEM in SAMP. CONCLUSIONS: IL33 promotes metaplasia and the sequelae of eosinophil-dependent downstream infiltration of IL33-producing M2 macrophages leading to intestinalized SPEM in SAMP, suggesting that IL33 represents a critical link between chronic gastritis and intestinalizing metaplasia that may serve as a potential therapeutic target for preneoplastic conditions of the GI tract.

IL-33 Mediates Lung Inflammation by the IL-6-Type Cytokine Oncostatin M.

The interleukin-1 family member IL-33 participates in both innate and adaptive T helper-2 immune cell responses in models of lung disease. The IL-6-type cytokine Oncostatin M (OSM) elevates lung inflammation, Th2-skewed cytokines, alternatively activated (M2) macrophages, and eosinophils in C57Bl/6 mice in vivo. Since OSM induces IL-33 expression, we here test the IL-33 function in OSM-mediated lung inflammation using IL-33-/- mice. Adenoviral OSM (AdOSM) markedly induced IL-33 mRNA and protein levels in wild-type animals while IL-33 was undetectable in IL-33-/- animals. AdOSM treatment showed recruitment of neutrophils, eosinophils, and elevated inflammatory chemokines (KC, eotaxin-1, MIP1a, and MIP1b), Th2 cytokines (IL-4/IL-5), and arginase-1 (M2 macrophage marker) whereas these responses were markedly diminished in IL-33-/- mice. AdOSM-induced IL-33 was unaffected by IL-6-/- deficiency. AdOSM also induced IL-33R+ ILC2 cells in the lung, while IL-6 (AdIL-6) overexpression did not. Flow-sorted ILC2 responded in vitro to IL-33 (but not OSM or IL-6 stimulation). Matrix remodelling genes col3A1, MMP-13, and TIMP-1 were also decreased in IL-33-/- mice. In vitro, IL-33 upregulated expression of OSM in the RAW264.7 macrophage cell line and in bone marrow-derived macrophages. Taken together, IL-33 is a critical mediator of OSM-driven, Th2-skewed, and M2-like responses in mouse lung inflammation and contributes in part through activation of ILC2 cells.

The IL-33/ST2 pathway suppresses murine colon cancer growth and metastasis by upregulating CD40 L signaling.

Interleukin (IL)-33 is a member of the IL-1 family, participating in both helper T1 (Th1)- and Th2-type immune responses, but its ambiguous effects on tumor growth and related immune mechanisms remain unclear. Here, we report that recombinant mouse IL-33 (mIL-33) significantly inhibited colon cancer growth and metastasis to lung and liver in a murine CT26 or MC38 tumor-cell engraftment model. This effect could be associated with CD4+ T cells and CD40 L signaling, as depletion of CD4+ T cells or blocking CD40 L signaling in vivo partly abolished the antitumor function of IL-33. In addition, IL-33 treatment upregulated CD40 L expression on tumor-infiltrating lymphocytes, and promoted the activation of CD4+ T, CD8+ T and natural killer cells via CD40 L signaling. Furthermore, IL-33 was sufficient to induce the ST2 expression on CD4+ T cells, but not on CD8+ T and natural killer cells, indicating that IL-33 acted on CD4+ T cells via a positive-feedback loop. Our findings shed new light on the IL-33-mediated antitumor effects and mechanisms of Th1 action, and also suggest that IL-33 may serve as an activator to boost anticancer immune responses in singular or combinatory therapies.

Hepatitis B virus X protein (HBx) promotes ST2 expression by GATA2 in liver cells.

At present, most studies on the relationship between hepatitis B virus (HBV) and IL-33/ST2 axis focus on clinical detection, but the underlying molecular mechanisms of HBx and IL-33/ST2 axis regulation and Th cell function regulation have not been explored. In this study, serum samples of patients with chronic hepatitis B (CHB) and HBV-related liver cancer (HBV-HCC), and healthy controls, as well as the supernatant solutions of HL7702-WT, HL7702-NC, and HL7702-HBx cells were collected to detect the content of soluble ST2 (sST2). The contents of Th1 cytokines (TNF-α and TNF-γ) and Th2 cytokines (IL-6 and IL-10) in the supernatant of different co-culture groups were detected. The effects of GATA2 on ST2 promoter transcription were investigated by upregulation or interference with GATA2 expression, dual-luciferase reporting, and ChIP experiments. The combined detection of sST2 and FIB-4 was beneficial to the non-invasive diagnosis of liver fibrosis. HBx promotes sST2 expression in liver cells, upregulates Th2 cell function, and inhibits Th1 cell function through IL-33/ST2 axis. HBx interacts with GATA2 to influence the activity of ST2 promoter. Serum sST2 detection is an invaluable indicator for the assessment of the progress of HBV infectious diseases, and the IL-33/ST2 axis plays an important role in changing the cellular immune function caused by HBV infection.

IL-33 is essential to prevent high-fat diet-induced obesity in mice infected with an intestinal helminth.

Intestinal helminthes induce immunosuppressive responses as well as type 2 immunity. Their suppressive properties are intended to regulate inflammatory diseases such as allergies and autoimmune diseases. This study evaluated whether helminthic infections suppress obesity, a chronic inflammatory state, using an intestinal nematode, Heligmosomoides polygyrus (Hp). Infection with Hp at the same time as feeding a high-fat diet (HFD) prevented weight gain, dyslipidaemia and glucose intolerance observed in uninfected obese mice. Immunologically, Hp infection skewed M1 macrophages to M2 macrophages and induced type 2 innate lymphoid cells in adipose tissues. The expression of interleukin (IL)-33, a potent initiator of type 2 responses, was also increased in association with uncoupled protein 1 (UCP1). To further investigate the anti-obesity effects of IL-33 in mice infected with Hp, IL-33-deficient mice were fed the HFD and infected with Hp. These mutant mice rapidly gained weight compared with wild-type mice, indicating the anti-obesity effect of IL-33. In the absence of IL-33, the rapid increase in weight was not prevented, and type 2 responses and UCP1 expression were not observed even during Hp infection. These results suggested that the suppression of obesity by Hp is dependent on IL-33.

Could interleukin-33 and its suppressor of tumorigenicity 2 (ST2) receptor have a role in cervical human papillomavirus (HPV) infections?

Why most women can clear human papillomavirus (HPV) infections while others can develop permanent infections. The stimulation of immunotolerance of the immune system of the host by the persistent HPV infection may be the answer to this question. Interleukin-33 (IL-33) may play a role in the pathogenesis of HPV infection, this hypothesis was thought to be due to the rapid release of IL-33 from damaged cells following tissue damage, necrosis, and activation of the inflammasome. Thus, in this study, the role of IL-33/suppressor of tumorigenicity 2 (ST2) was emphasized in HPV positive and HPV negative cervical tissues. A total of 80 were assessed. The reduced levels of IL-33 and ST2 are associated with cervical HPV infections. There was a statistically significant 42% positive correlation between IL-33 and ST2 in the HPV-positive group. Surprisingly, our data showed no significant difference between the expression levels of IL-33 or ST2 and working status, type of delivery, pre- and post-operative pathology, cigarette, educational status, locality, birth control method, gynecological, and colposcopic findings. We found that as a result of our study; low IL-33 and ST2 levels were associated with HPV infections.

IL (Interleukin)-33 Suppresses Abdominal Aortic Aneurysm by Enhancing Regulatory T-Cell Expansion and Activity.

Objective- Inflammation occurs during the progression of abdominal aortic aneurysm (AAA). IL (interleukin)-33 is a pleiotropic cytokine with multiple immunomodulatory effects, yet its role in AAA remains unknown. Approach and Results- Immunoblot, immunohistochemistry, and immunofluorescent staining revealed increased IL-33 expression in adventitia fibroblasts from mouse AAA lesions. Daily intraperitoneal administration of recombinant IL-33 or transgenic IL-33 expression ameliorated periaorta CaPO4 injury- and aortic elastase exposure-induced AAA in mice, as demonstrated by blunted aortic expansion, reduced aortic wall elastica fragmentation, enhanced AAA lesion collagen deposition, attenuated T-cell and macrophage infiltration, reduced inflammatory cytokine production, skewed M2 macrophage polarization, and reduced lesion MMP (matrix metalloproteinase) expression and cell apoptosis. Flow cytometry analysis, immunostaining, and immunoblot analysis showed that exogenous IL-33 increased CD4+Foxp3+ regulatory T cells in spleens, blood, and aortas in periaorta CaPO4-treated mice. Yet, ST2 deficiency muted these IL-33 activities. Regulatory T cells from IL-33-treated mice also showed significantly stronger activities in suppressing smooth muscle cell inflammatory cytokine and chemokine expression, macrophage MMP expression, and in increasing M2 macrophage polarization than those from vehicle-treated mice. In contrast, IL-33 failed to prevent AAA and lost its beneficial activities in CaPO4-treated mice after selective depletion of regulatory T cells. Conclusions- Together, this study established a role of IL-33 in protecting mice from AAA formation by enhancing ST2-dependent aortic and systemic regulatory T-cell expansion and their immunosuppressive activities.

IL-33 induced inflammation exacerbated the development of chronic obstructive pulmonary disease through oxidative stress.

OBJECTIVE: We aimed at exploring the role of IL-33 in mouse chronic obstructive pulmonary disease and its potential molecular mechanism. MATERIALS AND METHODS: The chronic obstructive pulmonary disease (COPD) mice model was established by cigarette smoking (CS). COPD mice were randomly assigned into PBS group and IL-33 antibody group. The peripheral blood and lung tissues of mice from two groups were collected for the following experiments. Pathological changes of the lung tissues in both groups were analyzed by hematoxylin and eosin (HE) staining. IL-33 positive cells in lung tissues were detected by immunohistochemistry. Then, the mRNA and protein levels of IL-33, sST2, ERK and TNF-α in the mice peripheral blood of the two groups were accessed by Real-time polymerase chain reaction (RT-PCR) and Western blot. Finally, the indicators related to oxidative stress, including superoxide dismutase (SOD), malondialdehyde (MDA) and reactive oxygen species (ROS) in the mice serum of two groups were measured. RESULTS: After successful construction of COPD mouse model by CS, HE staining illustrated that the structure of airway wall of lung tissue in mice from PBS group was irregular. The ciliated columnar epithelium presented significant degeneration, necrosis and shedding. A large amount of inflammation cell infiltration was observed in vascular tissues. The alveolar epithelial structure was severely damaged and alveolar septum was narrowed and ruptured. Adjacent alveoli were found to be fused into larger cysts. The above pathological changes were relatively better in mice from IL-33 antibody group. Immunohistochemical results demonstrated that IL-33 was remarkably deposited in the lung tissue of PBS group. The mRNA and protein levels of IL-33, sST2, ERK and TNF-α in peripheral blood of PBS group were much higher than those of IL-33 antibody group. At the same time, SOD level in PBS group decreased, while MDA level and ROS production increased. CONCLUSIONS: IL-33 aggravates lung injury in COPD mice by increasing inflammation response and oxidative stress, which may serve as a target for predicting and treating COPD.

Interleukin-33 induces interleukin-8 expression via JNK/c-Jun/AP-1 pathway in human umbilical vein endothelial cells.

Interleukin (IL)-33 is a member of the IL-1 cytokine family with dual functions as a traditional cytokine and as a transcriptional regulator. We recently reported that IL-33 up-regulated growth regulated oncogene (GRO)-α/CXCL1 expression in human vascular endothelial cells. The aim of this study was to investigate the effect of IL-33 on the expression of IL-8/CXCL8, another member of the CXC-chemokine family, and to elucidate its signaling pathways in human umbilical vein endothelial cells (HUVECs). Immunocytochemical staining and Western immunoblot analysis revealed that IL-33 augmented IL-8 protein expression in HUVECs. Real time reverse transcription-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) showed that IL-33 significantly increased IL-8 mRNA and secretion in a dose- and time-dependent manner. IL-33 preferentially stimulated proliferating subconfluent cells, and increased IL-8 secretion to a higher level compared with confluent cells. IL-33 also stimulated phosphorylations of c-Jun N-terminal kinase (JNK) and c-Jun, and enhanced activator protein (AP)-1 DNA-binding activity, all of which were suppressed by SP600125, a JNK inhibitor. Moreover, IL-33-induced IL-8 mRNA and secretion were also suppressed by SP600125. Transfection of c-Jun small interfering RNA into cultured HUVECs significantly reduced the IL-33-induced increase in IL-8 secretion from HUVECs. The present study demonstrates that IL-33 induces IL-8 expression via JNK/c-Jun/AP-1 pathway in human vascular endothelial cells, and provides a new insight into the role of IL-33-induced IL-8 in the pathophysiology of atherosclerosis and vascular inflammation.

Disruption of the Epidermal Barrier Induces Regulatory T Cells via IL-33 in Mice.

Disturbance of the epidermal barrier by UVR is associated with the release of antimicrobial peptides and inflammatory cytokines for the purpose of a danger response. On the other hand, UVR causes immunosuppression via regulatory T cells (Treg) that limit the inflammatory reaction. The concurrent induction of antimicrobial peptides and Treg by UVR may represent a counter-regulatory mechanism in response to barrier disruption, preventing microbial superinfection and sensitization to contact allergens, respectively, both of which cross impaired epidermis more easily. Thus, using a model of murine contact hypersensitivity we examined if disruption of the epidermal barrier only initiates similar counter-regulatory mechanisms via the generation of Treg. Sensitization through tape-stripped skin induced a weaker contact hypersensitivity response than in control mice. This was due to the induction of antigen-specific Treg, as demonstrated in adoptive transfer and depletion experiments utilizing DEREG mice. Treg induction by tape stripping was linked to the expression of the alarmin IL-33, as blockade of IL-33 exacerbated contact hypersensitivity, whereas injection of IL-33 inhibited contact hypersensitivity and induced Treg. These results demonstrate that epidermal barrier disruption, in addition to danger signals, induces regulatory events that prevent exaggerated skin inflammation and that IL-33 appears to be critically involved in this process.

MYB-GATA1 fusion promotes basophilic leukaemia: involvement of interleukin-33 and nerve growth factor receptors.

Acute basophilic leukaemia (ABL) is a rare subtype of acute myeloblastic leukaemia. We previously described a recurrent t(X;6)(p11;q23) translocation generating an MYB-GATA1 fusion gene in male infants with ABL. To better understand its role, the chimeric MYB-GATA1 transcription factor was expressed in CD34-positive haematopoietic progenitors, which were transplanted into immunodeficient mice. Cells expressing MYB-GATA1 showed increased expression of markers of immaturity (CD34), of granulocytic lineage (CD33 and CD117), and of basophilic differentiation (CD203c and FcϵRI). UT-7 cells also showed basophilic differentiation after MYB-GATA1 transfection. A transcriptomic study identified nine genes deregulated by both MYB-GATA1 and basophilic differentiation. Induction of three of these genes (CCL23, IL1RL1, and NTRK1) was confirmed in MYB-GATA1-expressing CD34-positive cells by reverse transcription quantitative polymerase chain reaction. Interleukin (IL)-33 and nerve growth factor (NGF), the ligands of IL-1 receptor-like 1 (IL1RL1) and neurotrophic receptor tyrosine kinase 1 (NTRK1), respectively, enhanced the basophilic differentiation of MYB-GATA1-expressing UT-7 cells, thus demonstrating the importance of this pathway in the basophilic differentiation of leukaemic cells and CD34-positive primary cells. Finally, gene reporter assays confirmed that MYB and MYB-GATA1 directly activated NTRK1 and IL1RL1 transcription, leading to basophilic skewing of the blasts. MYB-GATA1 is more efficient than MYB, because of better stability. Our results highlight the role of IL-33 and NGF receptors in the basophilic differentiation of normal and leukaemic cells. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

IL-33 Contributes to Schistosoma japonicum-induced Hepatic Pathology through Induction of M2 Macrophages.

Interleukin (IL)-33 is involved in T helper (Th)2-biased immune responses in mice infected with Schistosoma, but the precise mechanism remains to be elucidated. Herein, we investigated the role of IL-33 and its receptor ST2L in hepatic granuloma pathology induced by Schistosoma japonicum infection. We found that IL-33 induced the increased production of IL-5 and IL-13 from splenocytes and liver mononuclear cells (MNCs) of infected mice. The infected mice developed significantly higher number of ST2L-expressing cells in spleen and liver. Most of the ST2L-expressing cells in liver were F4/80(+) macrophages, indicating the key role of macrophages in the response to IL-33. However, the liver MNCs in male-only worm infection had a poor response to IL-33, though elevated serum IL-33 was observed. ST2L(+)F4/80(+) cells were lower in male-only worm infection than that of mixed infection. IL-33 and soluble egg antigen (SEA) upregulated ST2L expression on macrophages in vitro and ST2L-expressing macrophage displayed MHCII(-)CD11b(+)M2 phenotype. Macrophage deletion significantly attenuated IL-33-induced type 2 immunity and egg granuloma formation during S. japonicum infection. These data demonstrate that IL-33 contributes to hepatic granuloma pathology through induction of M2 macrophages during S. japonicum infection.

Interferon γ-Induced Nuclear Interleukin-33 Potentiates the Release of Esophageal Epithelial Derived Cytokines.

BACKGROUND: Esophageal epithelial cells are an initiating cell type in esophageal inflammation, playing an essential role in the pathogenesis of gastroesophageal reflux disease (GERD). A new tissue-derived cytokine, interleukin-33 (IL-33), has been shown to be upregulated in esophageal epithelial cell nuclei in GERD, taking part in mucosal inflammation. Here, inflammatory cytokines secreted by esophageal epithelial cells, and their regulation by IL-33, were investigated. METHODS: In an in vitro stratified squamous epithelial model, IL-33 expression was examined using quantitative RT-PCR, western blot, ELISA, and immunofluorescence. Epithelial cell secreted inflammatory cytokines were examined using multiplex flow immunoassay. IL-33 was knocked down with small interfering RNA (siRNA) in normal human esophageal epithelial cells (HEECs). Pharmacological inhibitors and signal transducers and activators of transcription 1 (STAT1) siRNA were used to explore the signaling pathways. RESULTS: Interferon (IFN)γ treatment upregulated nuclear IL-33 in HEECs. Furthermore, HEECs can produce various inflammatory cytokines, such as IL-6, IL-8, monocyte chemoattractant protein 1 (MCP-1), regulated on activation normal T-cell expressed and presumably secreted (RANTES), and granulocyte-macrophage colony-stimulating factor (GM-CSF) in response to IFNγ. Nuclear, but not exogenous IL-33, amplified IFN induction of these cytokines. P38 mitogen-activated protein kinase (MAPK) and janus protein tyrosine kinases (JAK)/STAT1 were the common signaling pathways of IFNγ-mediated induction of IL-33 and other cytokines. CONCLUSIONS: Esophageal epithelial cells can actively participate in GERD pathogenesis through the production of various cytokines, and epithelial-derived IL-33 might play a central role in the production of these cytokines.

An antitumorigenic role for the IL-33 receptor, ST2L, in colon cancer.

BACKGROUND: Despite the importance of inflammation in cancer, the role of the cytokine IL-33, and its receptor ST2, in colon cancer is unclear. The aim of this study was to investigate the role of IL-33, and its receptor isoforms (ST2 and ST2L), in colon cancer. METHODS: Serum levels of IL-33 and sST2 were determined with ELISA. ST2 and IL-33 expression was detected with quantitative real-time PCR (qRT-PCR), western blotting and immunohistochemistry. ST2 expression in CT26 cells was stably suppressed using ST2-specific shRNA. Cytokine and chemokine gene expression was detected with qRT-PCR. RESULTS: Human colon tumours showed lower expression of ST2L as compared with adjacent non-tumour tissue (P<0.01). Moreover, the higher the tumour grade, the lower the expression of ST2L (P=0.026). Colon cancer cells expressed ST2 and IL-33 in vitro. Functional analyses showed that stimulation of tumour cells with IL-33 induced the expression of chemokine (C-C motif) ligand 2 (CCL2). Knockdown of ST2 in murine colon cancer cells resulted in enhanced tumour growth (P<0.05) in BALB/c mice in vivo. This was associated with a decrease in macrophage infiltration, with IL-33-induced macrophage recruitment reduced by antagonising CCL2 in vitro. CONCLUSION: The IL-33/ST2 signalling axis may have a protective role in colon carcinogenesis.

Osteoprotective Effects of IL-33/ST2 Link to Osteoclast Apoptosis.

The relevance of IL-33 and its receptor ST2 for bone remodeling is not well-defined. Our aim was to assess the role and underlying mechanisms of IL-33/ST2 in mechanically induced bone remodeling. BALB/c (wild type) and ST2 deficient (St2(-/-)) mice were subjected to mechanical loading in alveolar bone. Microtomography, histology, and real-time quantitative PCR were performed to analyze bone parameters, apoptosis and bone cell counts, and expression of bone remodeling markers, respectively. MC3T3-E1 osteoblastic cells and bone marrow cells were used to verify if mechanical force triggered IL-33 and ST2 expression as well as the effects of IL-33 on osteoclast differentiation and activity. Mechanical loading increased the expression of IL-33 and ST2 in alveolar bone in vivo and in osteoblastic cells in vitro. St2(-/-) mice had increased mechanical loading-induced bone resorption, number of osteoclasts, and expression of proresorptive markers. In contrast, St2(-/-) mice exhibited reduced numbers of osteoblasts and apoptotic cells in periodontium and diminished expression of osteoblast signaling molecules. In vitro, IL-33 treatment inhibited osteoclast differentiation and activity even in the presence of receptor activator of NF-κB ligand. IL-33 also increased the expression of pro-apoptotic molecules, including Bcl-2-associated X protein (BAX), cell-surface Fas receptor (FAS), FASL, FAS-associated death domain, tumor necrosis factor-related apoptosis-inducing ligand, and BH3 interacting-domain death (BID). Overall, these findings suggest that IL-33/ST2 have anti-osteoclastogenic effects and reduce osteoclast formation and activity by inducing their apoptosis.