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Breast cancer (BC) poses one of the major threats to female's health worldwide. Immune infiltration in BC is a key representative of the tumor microenvironment and has been proven highly relevant for prognosis. The role of the FREM1 (FRAS1-Related Extracellular Matrix 1) gene in carcinoma has not studied, moreover, the underlying mechanism remains largely unknown. This study aims to investigate the expression profile and potential action of FREM1 on BC progression. We applied series of bioinformatic methods as well as immunohistochemistry (IHC) and immunofluorescence (IF) to analyze FREM1 expression profile, its relationship with clinicopathological characteristics, impact on clinical outcomes, relevant functions, correlation with immune infiltration in BC. The results demonstrated that FREM1 had a dramatically reduced expression in BC tissues, possessed an inverse correlation with stage, age, and metastasis, and exhibited a higher level in invasive lobular breast carcinoma than in ductal one. Furthermore, decreased FREM1 expression was often associated with estrogen receptor (ER)/progesterone receptor (PR) negative and triple negative breast carcinoma (TNBC) status while human epidermal growth factor 2 (Her-2) positive status, and considerably correlated with a worse overall survival (OS) and recurrence-free survival (RFS). Meanwhile, the univariate/multivariate Cox model revealed that low-FREM1 expression can be an independent prognostic factor for BC. Additionally, FREM1 was mainly involved in the cell metabolism and immune cells infiltration. Moreover, IHC and IF demonstrated a positive correlation of its expression with the immune infiltrating levels of CD4 + , CD8 + T cells, and CD86 + M1 macrophages while a negative correlation with CD68 + pan-macrophages and CD163 + M2 macrophages. These findings suggest that FREM1 can be a potential biomarker for evaluating the immune infiltrating status, and the BC prognosis., (© 2020 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

Solitary intestinal lymphoid tissues such as cryptopatches (CPs) and isolated lymphoid follicles (ILFs) constitute steady-state activation hubs containing group 3 innate lymphoid cells (ILC3) that continuously produce interleukin (IL)-22. The outer surface of CPs and ILFs is demarcated by a poorly characterized population of CD11c + cells. Using genome-wide single-cell transcriptional profiling of intestinal mononuclear phagocytes and multidimensional flow cytometry, we found that CP- and ILF-associated CD11c + cells were a transcriptionally distinct subset of intestinal cDCs, which we term CIA-DCs. CIA-DCs required programming by CP- and ILF-resident CCR6 + ILC3 via lymphotoxin-β receptor signaling in cDCs. CIA-DCs differentially expressed genes associated with immunoregulation and were the major cellular source of IL-22 binding protein (IL-22BP) at steady state. Mice lacking CIA-DC-derived IL-22BP exhibited diminished expression of epithelial lipid transporters, reduced lipid resorption, and changes in body fat homeostasis. Our findings provide insight into the design principles of an immunoregulatory checkpoint controlling nutrient absorption., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Glioma (GM) is one of the major global health problems across the world. Circular RNAs (circRNAs) have been increasingly identified and characterized in almost every aspect of biology, especially in cancer biology. This study desires to investigate the mechanism of circ-PITX1 on regulating GM development. Quantitative reverse-transcription polymerase chain reaction was carried out to measure the expression of circ-PITX1, which was upregulated in matched cancerous tissues and adjacent noncancerous tissues from 52 patients and four cell lines of GM. Fisher's exact indicated the upregulation of circ-PITX1 was associated with patients' tumor size and World Health Organization grade. Gain and loss-of-function assays demonstrated that circ-PITX1 could facilitate the growth, migration, and invasion and inhibit cell apoptosis in GM cell lines. What's more, circ-PITX1 sponges miR-518a-5p to release its repression on 3'-untranslated region (3'UTR) of interleukin 17 receptor D (IL17RD) messenger RNA to exert its oncogenic functions in GM cells proved by dual-luciferase reporter and rescue assays. Taken together, circ-PITX1 may play a critical role in GM and may be used as a therapeutic target for GM., (© 2019 Wiley Periodicals, Inc.)

Background: Influenza attacks the epithelium of the lung, causing cell death and disruption of the epithelial barrier leading to fluid buildup in the lung and impairment of gas exchange. Limited treatment options for severe influenza pneumonia prioritize the need for the discovery of effective therapies. IL-22 is a cytokine that promotes tissue integrity and has strong promise as a treatment option. While research has been focused on the cytokine itself, there is limited understanding of the regulation of the IL-22 receptor (IL-22Ra1) at the epithelial surface during infection., Methods: IL-22Ra1 levels were measured by qRT-PCR, western blot and immunofluorescence following H1N1 influenza infection (A/PR/8/34 H1N1) or synthetic TLR3 mimetic, Poly (I:C). Regulation of the receptor was determined using STAT inhibitors (STAT1, STAT3 and PanSTAT inhibitors), TLR3 inhibition, and neutralization of interferon alpha receptor 2 (IFNAR2). Significance was determined by a p-value of greater than 0.05. Significance between two groups was measured using unpaired t-test and significance between more than two groups was measured using one-way ANOVA with Tukey Multiple Comparison Test., Results: Here we show both in vivo and in vitro that IL-22Ra1 was induced as early as 24 h after influenza (H1N1 PR8) infection. This induction was triggered by toll-like receptor 3 (TLR3) as a TLR3 mimetic [Poly (I:C)] also induced IL-22Ra1 and inhibition of endosomal formation required for TLR3 function inhibited this process. This upregulation was dependent upon IFNβ signaling through STAT1. Importantly, induction of IL-22Ra1 significantly increased IL-22 signaling as evidenced by pSTAT3 levels following IL-22 treatment., Conclusion: Collectively, these data suggest epithelial cells may optimize the beneficial effects of IL-22 through the induction of the IL-22 receptor during viral infection in the lung.

Background: Thoracic ossification of the posterior longitudinal ligament (T-OPLL) can cause thoracic spinal stenosis, which results in intractable myelopathy and radiculopathy. The etiology of T-OPLL is unknown and the condition is difficult to treat surgically. Whole-genome sequencing identified a genetic variant at rs199772854 of the interleukin 17 receptor C (IL17RC) gene as a potentially pathogenic locus associated with T-OPLL. We aimed to determine whether the rs199772854A site mutation causes abnormal expression of the IL17RC in Han Chinese patients with T-OPLL and predict the possible pathogenic mechanisms of T-OPLL. Analyses were performed to determine whether IL17RC is involved in the pathogenicity of T-OPLL., Methods: Peripheral blood and OPLL tissue were collected from a total of 72 patients with T-OPLL disease (36 patients carrying the rs199772854A site mutation in IL17RC and 36 wild-type patients). The expression of IL17RC was analyzed by enzyme-linked immunosorbent assay, reverse transcription-quantitative polymerase chain reaction, immunohistochemistry, and Western blotting., Results: rs199772854A mutation resulted in markedly increased IL17RC gene expression levels in peripheral blood samples and the OPLL tissue obtained following clinical surgery (P < 0.05)., Conclusions: The results suggest that the rs199772854A site mutation of IL17RC can significantly increase the expression of IL17RC. The IL17RC gene rs199772854A site polymorphism is a potential pathogenic mutation in T-OPLL disease, which may be associated with the occurrence of T-OPLL.

The objective of this study was to conduct an analysis of peripheral blood Th17 cells with the ability to home to gut mucosa (CD4 + Th17 + β7 + ) during recent or chronic human immunodeficiency virus (HIV) infections. The relationship between HIV load and systemic inflammation markers was studied. Twenty-five patients with recent (n = 10) or chronic (n = 15) untreated HIV infections; 30 treated HIV-infected patients with undetectable HIV load at the time of inclusion and 30 healthy controls were included. Bacterial translocation markers (16S rDNA), soluble CD14 (sCD14) and interleukin (IL)-6 monocyte activation parameters, CD4/CD8 ratio and T helper type 17 (Th17) subpopulations [CD4 + Th17 + expressing the IL-23 receptor (IL-23R) or β7] were analysed at baseline and after 6 and 12 months of anti-retroviral therapy (ART). 16S rDNA was detected in all patients. Significantly increased serum levels of sCD14 and IL-6 and a decreased CD4/CD8 ratio were observed in patients. Similar percentages of CD4 + IL-23R + and CD4 + Th17 + β7 + cells were observed in healthy controls and patients at baseline. After 12 months of therapy, patients with a recent HIV infection showed significant increases of CD4 + IL-23R + and CD4 + Th17 + β7 + cell percentages and a decrease in IL-6 levels, although 16S rDNA continued to be detectable in all patients. No significant differences were observed in Th17 subpopulations in patients with chronic HIV infection after therapy. Early initiation of ART helps to increase the number of Th17 cells with the ability to home to the intestinal mucosa and to partially restore gut mucosal homeostasis. These results provide a rationale for initiating ART during the acute phase of HIV infection., (© 2018 British Society for Immunology.)

The immunomodulatory and self-renewable features of human adipose-derived mesenchymal stem cells (hAD-MSCs) mark their importance in regenerative medicine. Interleukin (IL)-23 as a proinflammatory cytokine suppresses T regulatory cells and promotes the response of T helper 17 and T helper 1 cells. This pathway initiates inflammation and immunosuppression in several autoimmune diseases. The current study aimed at producing recombinant IL-23 decoy receptor (RIL-23R) using hAD-MSCs as a good candidate for ex vivo cell-based gene therapy purposes to reduce inflammation in autoimmune diseases. hAD-MSCs was isolated from lipoaspirate and then characterized by differentiation. RIL-23R was designed and cloned into a pCDH813A-1 lentiviral vector. The transduction of hAD-MSCs was performed at multiplicity of infection = 50 with pCDH-EFI α-RIL-23R-PGK copGFP. Expressions of RIL-23R and octamer-binding transcription factor 4 (OCT-4) were determined by real-time polymerase chain reaction. Self-renewing properties were assayed with OCT-4. Bioactivity of the designed RIL-23R was evaluated by IL-17 and IL-10 expression of mouse splenocytes. The results showed that the transducted hAD-MSCs/RIL-23R, expressing IL-23 decoy receptor, can provide a useful approach for a basic research on cell-based gene therapy for autoimmune disorders.

Background: Interleukin (IL)-23 can facilitate the differentiation of IL-17-producing helper T cells (Th17). The IL-23/IL-17 axis is known to play a key role in the immunopathogenesis of ankylosing spondylitis (AS). We hypothesized that the expression of microRNAs (miRNAs, miRs) would be regulated by IL-23 and that these miRNAs could participate in the immunopathogenesis of AS., Methods: Expression profiles of human miRNAs in K562 cells, cultured in the presence or absence of IL-23 for 3 days, were analyzed by microarray. Potentially aberrantly expressed miRNAs were validated using T-cell samples from 24 patients with AS and 16 control subjects. Next-generation sequencing (NGS) was conducted to search for gene expression and biological functions regulated by specific miRNAs in the IL-23-mediated signaling pathway., Results: Initial analysis revealed that the expression levels of 12 miRNAs were significantly higher, whereas those of 4 miRNAs were significantly lower, in K562 cells after coculture with IL-23 for 3 days. Among these IL-23-regulated miRNAs, the expression levels of miR-29b-1-5p, miR-4449, miR-211-3p, miR-1914-3p, and miR-7114-5p were found to be higher in AS T cells. The transfection of miR-29b-1-5p mimic suppressed IL-23-mediated signal transducer and activator of transcription 3 (STAT3) phosphorylation in K562 cells. After NGS analysis and validation, we found that miR-29b-1-5p upregulated the expression of angiogenin, which was also upregulated in K562 cells after coculture with IL-23. Increased expression of miR-29b-1-5p or miR-211-3p could enhance interferon-γ expression., Conclusions: Among the miRNAs regulated by IL-23, expression levels of five miRNAs were increased in T cells from patients with AS. The transfection of miR-29b-1-5p mimic could inhibit the IL-23-mediated STAT3 phosphorylation and might play a role in negative feedback control in the immunopathogenesis of AS.

Objective: Vogt-Koyanagi-Harada (VKH) disease is an autoimmune disease mediated by T cells that target melanocytes. It has been shown that IL-23 receptor (IL-23R) signaling promotes the generation of pathogenic T helper 17 cells. The aim of this study was designed to detect the possible role of IL-23R in VKH disease., Methods: Subjects were divided into an active and inactive VKH patient group and a normal control group. The IL-23R level in peripheral blood mononuclear cells (PBMCs) was measured by flow cytometry and real-time polymerase chain reaction. PBMCs were stimulated with serum from patients or controls to detect the influence of serum from VKH patients on IL-23R expression., Results: The IL-23R mRNA level was markedly increased in PBMCs from the active VKH patient group as compared to normal controls. Flow cytometry analysis showed that there was also an elevated IL-23R protein level in PBMCs in active VKH patients. The IL-23R protein level was higher in PBMCs obtained from healthy controls when they were cultured with serum from active VKH patient as compared to cell cultured with serum from normal controls. After the intraocular inflammation in VKH patients was controlled, the IL-23R gene expression returned back to normal levels., Conclusion: Our study suggests that an elevated IL-23R level may participate in the development of VKH disease.

Background: Interleukin-23 (IL-23) has been implicated in inflammatory and autoimmune diseases by skewing CD4 + T helper cells towards a pathogenic Th17 phenotype. In this study we investigated the presence of IL-23 receptor (IL-23R)-expressing cells in the atherosclerotic aorta and evaluated the effect of IL-23R deficiency on atherosclerosis development in mice., Methods and Results: We used heterozygous Ldlr -/- Il23r e GFP / WT knock-in mice to identify IL-23R-expressing cells by flow cytometry and homozygous Ldlr -/- Il23r e GFP / eGFP ( Ldlr -/- Il23r -/- ) mice to investigate the effect of lack of IL-23R in atherosclerosis. We demonstrate the presence of relatively rare IL-23R-expressing cells in lymphoid tissue and aorta (≈0.1-1% IL23R + cells of all CD45 + leukocytes). After 10 weeks on a high-fat diet, production of IL-17, but not interferon-γ, by CD4 + T cells and other lymphocytes was reduced in Ldlr -/- Il23r -/- compared with Ldlr -/- controls. However, Ldlr -/- and Ldlr -/- Il23r -/- mice had equivalent amounts of aortic sinus and descending aorta lesions. Adoptive transfer of IL-23R-deficient CD4 + T cells to lymphopenic Ldlr -/- Rag1 -/- resulted in dramatically reduced IL-17-producing T cells but did not reduce atherosclerosis, compared with transfer of IL-23R-sufficient CD4 + T cells., Conclusions: These data demonstrate that loss of IL-23R does not affect development of experimental atherosclerosis in LDLr-deficient mice, despite a role for IL-23 in differentiation of IL-17-producing T cells., (© 2018 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley.)

IL-17RB, a member of the IL-17 receptor family that can be activated by IL-17B, has been proved to be involved in inflammatory diseases and cancers. However, the function of IL-17RB in thyroid cancer is still unknown. In this study, IL-17RB expression in thyroid cancer cell lines and tissues was examined by real-time PCR and western blot. The effects of IL-17RB on cell invasion and migration were determined by in vitro invasion and migration assays, while the effects of IL-17RB on cell metastasis were analyzed by in vivo experiments. The results showed that IL-17RB expression was upregulated in both thyroid cancer cells and tissues. IL-17B dose-dependently promoted the invasion, growth and migration of thyroid cancer cells, whereas knockdown of IL-17RB attenuated the effects of IL-17B in vitro. Moreover, IL-17RB was involved in the metastasis and growth of thyroid cancer cells in vivo. In addition, IL-17RB induced ERK1/2 activation and increased MMP-9 expression in vitro and in vivo. Inhibition of ERK1/2 pathway blocked the IL-17RB-mediated thyroid cancer cell invasion and MMP-9 expression. Together, our findings demonstrate that IL-17RB can enhance thyroid cancer cell invasion and metastasis via ERK1/2 pathway-mediated MMP-9 expression, suggesting that IL-17RB may act as a potential therapeutic target for thyroid cancer therapy., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Purpose: To survey the changes of promoter CpG methylation status and mRNA expression of IL17RC (interleukin 17 receptor C) gene in retinal pigment epithelium (RPE) cells under chemical hypoxia condition for choroidal neovascularization (CNV) modeling in vitro., Materials and Methods: RPE cells were cultured in both untreated as a control group and treated by cobalt chloride media as a hypoxia group for various concentrations (100-150μM) and times (24-36 hrs.) To confirm chemical hypoxia condition, mRNA expression of HIF (Hypoxia Inducible Factor) -1α, -2α, and Vascular Endothelial Growth Factor (VEGF) was compared between two groups by Real-time PCR. Also, in normoxia and hypoxia conditions, IL17RC expression changes and promoter CpG methylation status were evaluated by Real-time PCR and methylation-specific PCR (MSP) techniques, respectively., Results: Overexpression of HIF-1α, HIF-2α, and VEGF was significant in hypoxia versus normoxia conditions. Our data showed overexpression of IL17RC (2.1- to 6.3-fold) and decreasing of its promoter methylation in comparison with hypoxia and normoxia conditions. It was found that there are significant association between promoter methylation status and expression of IL17RC in chemical hypoxia condition., Conclusion: Therefore, methylation of IL17RC could play as a marker in CNV and degeneration of RPE cells in vitro. Additionally, HIF-α and methylation phenomena may be considered as critical targets for blocking in angiogenesis of age-related degeneration in future studies.

BACKGROUND IL-23/IL-23R signaling plays a pivotal role during the course of inflammatory bowel diseases (IBD). However, the underlying mechanisms are poorly characterized. Foxp3+ regulatory T cells are critical in the maintenance of gut immune homeostasis and therefore are important in preventing the development of IBD. This study was performed to clarify the association between IL-23/IL-23R signaling and Foxp3+ regulatory T cells in colitis. MATERIAL AND METHODS Acute and chronic mouse colitis models were established by administering mice DSS in drinking water. IL-23R, IL-23, IL-I7, and IFN-γ expression level, as well as regulatory T cell, Th17-, and Th1-related transcription factors Foxp3, RORgt, and T-bet were assayed by real-time PCR. The frequency of Foxp3+ RORγt+ cells in a Foxp3+ cell population in colon mucosa during acute and chronic colitis was evaluated through flow cytometry. The signaling pathway mediated by IL-23R in the colon mucosa from acute colitis mice and chronic colitis mice was monitored by Western blot analysis. RESULTS We detected elevated IL-23R, IL-23, and IFN-γ expression in colon mucosa during acute and chronic colitis and found increased IL-17 in acute colitis mice. Transcription factors Foxp3 and T-bet were elevated in colon mucosa during acute and chronic colitis. Phosphorylation of Stat3 was greatly enhanced, indicating the activation of IL-23R function in colitis mice. The percentage of Foxp3+ T cells in acute and chronic colitis mice was comparable to control mice, but there was a 2-fold increase of Foxp3+ RORγt+ cells among the Foxp3+ cell population in acute and chronic colitis mice compared to control mice. CONCLUSIONS These findings indicate that the induction of Foxp3+ RORgt+ T cells could be enhanced during inflammation in the intestine where IL-23R expression is greatly induced. Our study highlights the importance of IL-23R expression level and the instability of Foxp3+ regulatory T cells in the development of inflammatory bowel diseases.

Background: Biomarkers that can be used to accurately assess the residual risk of disease recurrence in women with hormone receptor-positive breast cancer are clinically valuable. We evaluated the prognostic value of the Breast Cancer Index (BCI), a continuous risk index based on a combination of HOXB13:IL17BR and molecular grade index, in women with early breast cancer treated with either tamoxifen alone or tamoxifen plus octreotide in the NCIC MA.14 phase III clinical trial (ClinicalTrials.gov Identifier NCT00002864; registered 1 November 1999)., Methods: Gene expression analysis of BCI by real-time polymerase chain reaction was performed blinded to outcome on RNA extracted from archived formalin-fixed, paraffin-embedded tumor samples of 299 patients with both lymph node-negative (LN-) and lymph node-positive (LN+) disease enrolled in the MA.14 trial. Our primary objective was to determine the prognostic performance of BCI based on relapse-free survival (RFS). MA.14 patients experienced similar RFS on both treatment arms. Association of gene expression data with RFS was evaluated in univariate analysis with a stratified log-rank test statistic, depicted with a Kaplan-Meier plot and an adjusted Cox survivor plot. In the multivariate assessment, we used stratified Cox regression. The prognostic performance of an emerging, optimized linear BCI model was also assessed in a post hoc analysis., Results: Of 299 samples, 292 were assessed successfully for BCI for 146 patients accrued in each MA.14 treatment arm. BCI risk groups had a significant univariate association with RFS (stratified log-rank p = 0.005, unstratified log-rank p = 0.007). Adjusted 10-year RFS in BCI low-, intermediate-, and high-risk groups was 87.5 %, 83.9 %, and 74.7 %, respectively. BCI had a significant prognostic effect [hazard ratio (HR) 2.34, 95 % confidence interval (CI) 1.33-4.11; p = 0.004], although not a predictive effect, on RFS in stratified multivariate analysis, adjusted for pathological tumor stage (HR 2.22, 95 % CI 1.22-4.07; p = 0.01). In the post hoc multivariate analysis, higher linear BCI was associated with shorter RFS (p = 0.002)., Conclusions: BCI had a strong prognostic effect on RFS in patients with early-stage breast cancer treated with tamoxifen alone or with tamoxifen and octreotide. BCI was prognostic in both LN- and LN+ patients. This retrospective study is an independent validation of the prognostic performance of BCI in a prospective trial.

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., (Copyright © 2015 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

IL-17-producing CD4+ T cells (Th17 cells) have well-described pathogenic roles in tissue inflammation and autoimmune diseases, such as experimental autoimmune encephalomyelitis (EAE); however, the involvement of IL-21 in these processes has remained controversial. While IL-21 is an essential autocrine amplification factor for differentiation of Th17 cells, the loss of IL-21 or IL-21 receptor (IL-21R) does not protect mice from actively induced EAE. Here, we utilized a transgenic EAE mouse model, in which T and B cells overexpress receptors for myelin oligodendrocyte glycoprotein (MOG) (referred to as 2D2xTH mice), and demonstrated that IL-21 is critical for the development of a variant form of spontaneous EAE in these animals. Il21r deletion in 2D2xTH mice reduced the incidence and severity of spontaneous EAE, which was associated with a defect in Th17 cell generation. Moreover, IL-21R deficiency limited IL-23R expression on Th17 cells and inhibited expression of key molecules involved in the generation of pathogenic Th17 cells. Conversely, loss of IL-23R in 2D2xTH mice resulted in complete resistance to the development of spontaneous EAE. Our data identify a previously unappreciated role for IL-21 in EAE and reveal that IL-21-mediated signaling supports generation and stabilization of pathogenic Th17 cells and development of spontaneous autoimmunity.

The mucosal immune response in the setting of intestinal inflammation contributes to colorectal cancer. IL10 signaling has a central role in gut homeostasis and is impaired in inflammatory bowel disease (IBD). Out of two IL10 receptor subunits, IL10R1 and IL10R2, the latter is shared among the IL10 family of cytokines and activates STAT signaling. STAT3 is oncogenic in colorectal cancer; however, knowledge about IL10 signaling upstream of STAT3 in colorectal cancer is lacking. Here, expression of IL10 signaling genes was examined in matched pairs from normal and tumor tissue from colorectal cancer patients showing overexpression (mRNA, protein) of IL10R2 and STAT3 but not IL10R1. IL10R2 overexpression was related to microsatellite stability. Transient overexpression of IL10R2 in HT29 cells increased proliferation upon ligand activation (IL10 and IL22). IL22, and not IL10, phosphorylated STAT3 along with increased phosphorylation of AKT and ERK. A significantly higher expression of IL22R1 and IL10R2 was also confirmed in a separate cohort of colorectal cancer samples. IL22 expression was elevated in gut mucosa from patients with IBD and colitis-associated cancer, which also exhibited increased expression of IL22R1 but not its coreceptor IL10R2. Overall, these data indicate that overexpression of IL10R2 and STAT3 contributes to colorectal carcinogenesis in microsatellite-stable tumors through IL22/STAT3 signaling., (©2015 American Association for Cancer Research.)

Cytokines play important roles in cardiac repair and regeneration. Recently, we demonstrated that interleukin (IL)-6 family cytokines induce the endothelial differentiation of Sca-1+ cardiac resident stem cells through STAT3/Pim-1 signaling pathway. In contrast, the biological functions of IL-12 family cytokines in heart remain to be elucidated, though they show structural homology with IL-6. In the present study, we examined the effects of IL-12 family cytokines on the transdifferentiation of cardiac Sca-1+ cells into cardiac cells. RT-PCR analyses revealed that IL-27 receptor α (IL-27Rα), but not IL-12R or IL-23R, was expressed in cardiac Sca-1+ cells. The transcript expression of IL-27 was elevated in murine hearts in cardiac injury models. Intriguingly, IL-27 stimulation for 14 days induced the endothelial cell (EC) marker genes, such as CD-31 and VE-cadherin. Immunoblot analyses clarified that IL-27 treatment rapidly phosphorylated STAT3. IL-27 upregulated the expression of Pim-1, but the overexpression of dominant negative STAT3 abrogated the induction of Pim-1 by IL-27. Finally, adenoviral transfection of dominant negative Pim-1 inhibited IL-27-induced EC differentiation of cardiac Sca-1+ cells. These findings demonstrated that IL-27 promoted the commitment of cardiac stem cells into the EC lineage, possibly leading to neovascularization as a novel biological function. IL-27 could not only regulate the inflammation but also contribute to the maintenance of the tissue homeostasis through stem cell differentiation at inflammatory sites., (Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Differences in the activity of monocytes/macrophages, important target cells of Mycobacterium tuberculosis, might influence tuberculosis progression. With the purpose of identifying candidate genes for tuberculosis susceptibility we infected monocytes from both healthy elderly individuals (a tuberculosis susceptibility group) and elderly tuberculosis patients with M. tuberculosis, and performed a microarray experiment. We detected 78 differentially expressed transcripts and confirmed these results by quantitative PCR of selected genes. We found that monocytes from tuberculosis patients showed similar expression patterns for these genes, regardless of whether they were obtained from younger or older patients. Only one of the detected genes corresponded to a cytokine: IL26, a member of the interleukin-10 (IL-10) cytokine family which we found to be down-regulated in infected monocytes from tuberculosis patients. Non-infected monocytes secreted IL-26 constitutively but they reacted strongly to M. tuberculosis infection by decreasing IL-26 production. Furthermore, IL-26 serum concentrations appeared to be lower in the tuberculosis patients. When whole blood was infected, IL-26 inhibited the observed pathogen-killing capability. Although lymphocytes expressed IL26R, the receptor mRNA was not detected in either monocytes or neutrophils, suggesting that the inhibition of anti-mycobacterial activity may be mediated by lymphocytes. Additionally, IL-2 concentrations in infected blood were lower in the presence of IL-26. The negative influence of IL-26 on the anti-mycobacterial activity and its constitutive presence in both serum and monocyte supernatants prompt us to propose IL26 as a candidate gene for tuberculosis susceptibility., (© 2014 John Wiley & Sons Ltd.)

The present results demonstrated that high glucose (G), salt (S), and cholesterol C (either alone or in combination), as mimicking extracellular changes in metabolic syndrome, damage cardiomyocyte-like H9c2 cells and reduce their viability in a time-dependent manner. However, the effects were greatest when cells were exposed to all three agents (GSC). The mRNA of glycoprotein (gp) 130 and WSX-1, both components of the interleukin (IL)-27 receptor, were present in H9c2 cells. Although mRNA expression was not affected by exogenous treatment with IL-27, the expression of gp130 mRNA (but not that of WSX-1 mRNA) was attenuated by GSC. Treatment of IL-27 to H9c2 cells increased activation of signal transducer and activator of transcription 3 (STAT3) and protected cells from GSC-induced cytochrome c release and cell damage. The protective effects of IL-27 were abrogated by the STAT3 inhibitor, stattic. The results of the present study clearly demonstrate that the STAT3 pathway triggered by anti-inflammatory IL-27 plays a role in protecting cardiomyocytes against GSC-mediated damage.

Interleukin-33 (IL-33) is associated with several important immune-mediated disorders. However, its role in uveitis, an important eye inflammatory disease, is unknown. Here, we investigated the function of IL-33 in the development of experimental autoimmune uveitis (EAU). IL-33 and IL-33 receptor (ST2) were expressed in murine retinal pigment epithelial (RPE) cells in culture, and IL-33 increased the expression of Il33 and Mcp1 mRNA in RPE cells. In situ, IL-33 was highly expressed in the inner nuclear cells of the retina of naïve mice, and its expression was elevated in EAU mice. ST2-deficient mice developed exacerbated EAU compared with WT mice, and administration of IL-33 to WT mice significantly reduced EAU severity. The attenuated EAU in IL-33-treated mice was accompanied by decreased frequency of IFN-γ+ and IL-17(+) CD4+ T cells and reduced IFN-γ and IL-17 production but with increased frequency of IL-5(+) and IL-4(+) CD4 T cells and IL-5 production in the draining lymph node and spleen. Macrophages from the IL-33-treated mice show a significantly higher polarization toward an alternatively activated macrophage phenotype. Our results therefore demonstrate that the endogenous IL-33/ST2 pathway plays an important role in EAU, and suggest that IL-33 represents a potential option for treatment of uveitis., (© 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Programmed necrosis or necroptosis is an inflammatory form of cell death that critically requires the receptor-interacting protein kinase 3 (RIPK3). Here we showed that RIPK3 controls a separate, necrosis-independent pathway of inflammation by regulating cytokine expression in dendritic cells (DCs). Ripk3(-/-) bone-marrow-derived dendritic cells (BMDCs) were highly defective in lipopolysaccharide (LPS)-induced expression of inflammatory cytokines. These effects were caused by impaired NF-κB subunit RelB and p50 activation and by impaired caspase 1-mediated processing of interleukin-1β (IL-1β). This DC-specific function of RIPK3 was critical for injury-induced inflammation and tissue repair in response to dextran sodium sulfate (DSS). Ripk3(-/-) mice exhibited an impaired axis of injury-induced IL-1β, IL-23, and IL-22 cytokine cascade, which was partially corrected by adoptive transfer of wild-type DCs, but not Ripk3(-/-) DCs. These results reveal an unexpected function of RIPK3 in NF-κB activation, DC biology, innate inflammatory-cytokine expression, and injury-induced tissue repair.

IL-33 is a recently characterized IL-1 family member that is proposed to function as an alarmin, or endogenous signal of cellular damage, as well as act as a pleiotropic cytokine. The ability of IL-33 to potentiate both Th1 and Th2 immunity supports its role in pathogen clearance and disease immunopathology. Yet, IL-33 restrains experimental colitis and transplant rejection by expanding regulatory T cells (Treg) via an undefined mechanism. We sought to determine the influence of IL-33 on hematopoietic cells that drives Treg expansion and underlies the therapeutic benefit of IL-33 administration. In this study, we identify a feedback loop in which conventional mouse CD11c(+) dendritic cells (DC) stimulated by IL-33 secrete IL-2 to selectively expand IL-33R(ST2(+))- suppressive CD4(+)Foxp3(+) Treg. Interestingly, this occurs in the absence of classical DC maturation, and DC-derived (innate) IL-2 increases ST2 expression on both DC and interacting Treg. ST2(+) Treg represent an activated subset of Foxp3(+) cells, demonstrated to be ICOS(high)CD44(high) compared with their ST2(-) counterparts. Furthermore, although studies have shown that IL-33-exposed DC promote Th2 responses, we reveal that ST2(+) DC are required for IL-33-mediated in vitro and in vivo Treg expansion. Thus, we have uncovered a relationship between IL-33 and innate IL-2 that promotes the selective expansion of ST2(+) Treg over non-Treg. These findings identify a novel regulatory pathway driven by IL-33 in immune cells that may be harnessed for therapeutic benefit or for robust expansion of Treg in vitro and in vivo., (Copyright © 2014 by The American Association of Immunologists, Inc.)

Interleukin-33 (IL-33) appears to play a crucial role in the expression of allergic diseases, but its cellular source and regulatory mechanisms remain to be fully elucidated. Mast cells, one of the major effecter cell populations in mediating allergy, express high levels of IL-33 receptor, ST2, and have been shown to express IL-33 transcripts. In this study, we aimed to examine the secretion of IL-33 in mast cells and their response to IL-33. We have successfully detected secreted IL-33 from cell supernatants through a modified enzyme-linked immunosorbent assay (ELISA) technique-cell-based ELISA. Activation of bone marrow-derived cultured mast cells (BMMCs) by crosslinkage of an antigen [ovalbumin (OVA)] and OVA-specific IgE mAbs significantly induced the expression of IL-33 transcripts, cytosolic and secreted proteins. In addition, the Toll-like receptor (TLR) 2 and TLR-9 ligands could trigger IL-33 mRNA expression. Exposure of BMMCs to IL-33 significantly increased the levels of IL-13 and IL-6 expression, concomitant with enhanced activation of mitogen-activated protein kinase (MAPKs) (ERK, p38, and JNK) and nuclear factor-kappa B. These results suggest that mouse BMMCs are capable of producing and serving as endogenous sources of IL-33, and that IL-33 plays an important role in regulating mast cell functions.

Airway inflammation in allergic asthma reflects a threshold response of the innate immune system, including group 2 innate lymphoid cells (ILC2), followed by an adaptive Th2 cell-mediated response. Transcription factor Gata3 is essential for differentiation of both Th2 cells and ILC2. We investigated the effects of enforced Gata3 expression in T cells and ILC2 on the susceptibility of mice to allergic airway inflammation (AAI). We used CD2-Gata3 transgenic (Tg) mice with enforced Gata3 expression driven by the CD2 promoter, which is active both in T cells and during ILC2 development. CD2-Gata3 Tg mice and wild-type (WT) littermates were analyzed in mild models of AAI without adjuvants. Whereas OVA allergen exposure did not induce inflammation in WT controls, CD2-Gata3 Tg mice showed clear AAI and enhanced levels of IL-5 and IL-13 in bronchoalveolar lavage. Likewise, in house dust mite-driven asthma, CD2-Gata3 Tg mice were significantly more susceptible to AAI than WT littermates, whereby both ILC2 and Th2 cells were important cellular sources of IL-5 and IL-13 in bronchoalveolar lavage and lung tissue. Compared with WT littermates, CD2-Gata3 Tg mice contained increased numbers of ILC2, which expressed high levels of IL-33R and contributed significantly to early production of IL-4, IL-5, and IL-13. CD2-Gata3 Tg mice also had a unique population of IL-33-responsive non-B/non-T lymphoid cells expressing IFN-γ. Enforced Gata3 expression is therefore sufficient to enhance Th2 and ILC2 activity, and leads to increased susceptibility to AAI after mild exposure to inhaled harmless Ags that otherwise induce Ag tolerance.

Cytokines are immunomodulating proteins involved in cellular communication. The levels of different cytokines reflect the immune capabilities of a person. In literature both whole blood and peripheral blood mononuclear cells (PBMCs) are used, which might lead to different results. The choice between these different sources is not always explained. The goal of our experiments is to determine the cytokine response of whole blood, PBMCs and polymorphonuclear cells (PMNs) after stimulation with lipopolysaccharide (LPS). We used a multiplex analysis to determine a difference in cytokine secretion patterns. In general, PBMCs demonstrated the highest cytokine production and PMNs have an overall low cytokine production. CCL11 and interleukin-23 (IL-23) (and IL-12p40) were exclusively expressed in whole blood. IL-20, VEGF and GM-CSF were expressed only by PBMCs. This difference in expression could be explained by the bioactive components in serum, presence and interaction with granulocytes or platelets in whole blood, the anticoagulant heparin in whole blood and others. The expression of cytokines by cells is dependent on the microenvironment. Different conditions lead to different results. We recommend a thorough examination of the conditions before performing experiments., (© 2013.)

The aim of the study was to characterize endometrial mRNA transcription, immunolocalization, and protein expression of interleukin (IL) 1alpha, IL1beta, IL6, and IL1RI, IL1RII, and IL6Ralpha/beta in the course of endometrosis during the estrous cycle. Additionally, the influence of IL1alpha, IL1beta, and IL6 on prostaglandin (PG) secretion and PG synthase mRNA transcription in endometrial tissue during endometrosis was investigated. The endometrial samples were obtained at the early (n = 12), mid- (n = 12), and late (n = 12) luteal phases and at the follicular (n = 12) phase of the estrous cycle. Within each of these phases, there were four samples within each category I, II, and III of endometrium, according to the Kenney classification. In experiment 1, transcription of IL1alpha, IL1beta, IL6, and their receptor's (IL1RI, IL1RII, and IL6Ralpha/beta) mRNAs and their immunolocalization and protein expression were determined using real-time PCR and immunohistochemistry, respectively. In Experiment 2, endometrial samples (n = 5 samples within categories I, II, and III) were obtained for tissue culture in the midluteal phase of the estrous cycle. The endometrial tissues were stimulated with IL1alpha (10 ng/ml), IL1beta (10 ng/ml), IL6 (10 ng/ml), and oxytocin (positive control; 10⁻⁷ M) for 24 h. The PG concentration was determined using ELISA. In addition, transcription of PTGS-2, PGES, and PGFS mRNAs was determined using real-time PCR. ILs were found to regulate PG secretion via modulation of PG synthases in equine endometrium. The alterations in IL and the expression of their receptors, and in endometrial secretory functions, were observed during the course of endometrosis, and suggest serious changes in the endometrial microenvironment. The described disturbances may be closely related to impaired endometrial processes responsible for the subfertility or the infertility in endometrosis.

Primary biliary cirrhosis (PBC) is a typical autoimmune disease for which the pathogenesis remains unclear. IL-23 and IL-17 are pro-inflammatory cytokines of the "IL-23/IL-17 axis," which may play a key role in the pathogenesis of autoimmune diseases. In this study, we investigated the expression of IL-23 and IL-17 in the peripheral blood of patients with PBC and its clinical significance. We used quantitative PCR to determine mRNA expressions of IL-23, IL-23 receptor, and IL-17 in peripheral blood mononuclear cells (PBMC) from PBC patients. ELISA's were used to determine patients' serum levels of IL-23 and IL-17. IL-23- and IL-17-producing cells in liver biopsis were also analyzed. Compared to a healthy control group, the mRNA expression levels of IL-23 p19, its corresponding receptor, IL-23R, and IL-17 in PBMC's from PBC patients were significantly increased, and these levels were correlated with PBC disease stages. PBC patients' serum levels of IL-23 and IL-17 were higher than those in a post-hepatic cirrhosis group and a healthy group, and were significantly higher in the early PBC disease stages than in the advanced PBC stages. There were significantly more IL23+ and IL-17+ mononuclear cells in portal areas of liver tissues in advanced stages of this disease than in the early stages. The serum levels of IL-23 and IL-17 in PBC patients were positively correlated with serum GGT levels. Thus, IL-23 and IL-17 may play an important role in the pathogenesis of PBC by promoting inflammation. Because the IL-23 and IL-17 levels in the peripheral blood of PBC patients were increased and were correlated with clinical stages, they may be indices that could be used to clinically monitor PBC., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

We have previously reported that, depending on their activation status, mouse γδ T cells can either enhance or inhibit the activity of IL-17(+) autoreactive T cells in experimental autoimmune uveitis. In this study, we showed that γδ T cells in naive C57BL/6 (B6) mouse do not express the IL-23R, whereas in immunized mice, it is expressed on >50% of γδ T cells. In vitro studies showed that IL-23R expression on γδ T cells is modulated by their state of activation, as weakly activated γδ T cells expressed the IL-23R, but highly activated γδ T cells did not. Functional studies showed that IL-23R(+) γδ T cells had the strongest suppressive effect on IL-17(+) autoreactive T cells, and that this effect was inhibited when the IL-23R was blocked by anti-IL-23R Ab or in the presence of excessive amounts of exogenous IL-23. We conclude that the balance between the enhancing and inhibitory effects of γδ T cells is regulated by their level of IL-23R expression. The expression of variable IL-23R levels allows γδ T cells to have different regulatory effects on adaptive immune responses, conceivably as a result of αβ and γδ T cells competing for IL-23.

Coxsackievirus B (CVB) is a common cause of acute and chronic infectious myocarditis and pancreatitis. Th1 cells producing IFN-γ and TNF-α are important for CVB clearance, but they are also associated with the pathogenesis of inflammatory lesions, suggesting that the modulation of Th1 and Th2 balance is likely important in controlling CVB-induced pancreatitis. We investigated the role of IL-33, which is an important recently discovered cytokine for induction of Th2-associated responses, in experimental CVB5 infection. We found that mice deficient in IL-33R, T1/ST2, significantly developed more severe pancreatitis, had greater weight loss, and contained higher viral load compared with wild-type (WT) mice when infected with CVB5. Conversely, WT mice treated with rIL-33 developed significantly lower viral titers, and pancreatitis was attenuated. Mechanistic studies demonstrated that IL-33 enhances the degranulation and production of IFN-γ and TNF-α by CD8(+) T and NK cells, which is associated with viral clearance. Furthermore, IL-33 triggers the production of IL-4 from mast cells, which results in enhanced differentiation of M2 macrophages and regulatory T cells, leading to the attenuation of inflammatory pancreatitis. Adoptively transferred mast cells or M2 macrophages reversed the heightened pancreatitis in the T1/ST2(-/-) mice. In contrast, inhibition of regulatory T cells exacerbated the disease in WT mice. Together, our findings reveal an unrecognized IL-33/ST2 functional pathway and a key mechanism for CVB5-induced pancreatitis. These data further suggest a novel approach in treating virus-induced pancreatitis, which is a major medical condition with unmet clinical needs.

Objectives: The aim of this study was to compare the expression of interleukin (IL)-22, IL-22R and IL-23 in the peri-implant soft tissues between the peri-implantitis patient group (PG) and peri-implant healthy control group (HG)., Methods: The tissues were collected from 12 peri-implantitis patients and eight peri-implant healthy controls. Immunohistochemistry (IHC) and real-time quantitative PCR (qPCR) were performed to analyse the gene expression of IL-22, IL-22R and IL-23p19 in peri-implant soft tissues in the PG and the HG group., Results: The IHC result showed that number of IL-22, IL-22R, and IL-23p19 positive cells increased in PG than in HG (P<0.05). The result of qPCR demonstrated that the expressions of IL-22 messenger RNA (mRNA) and IL-23p19 mRNA were significantly higher in the PG group compared to the HG group (P<0.05). Gene expression of IL-22R mRNA was higher in the PG group; however, there was no statistically significant difference between these two groups (P>0.05)., Conclusions: This study indicates that there is an increased expression level of IL-22 and IL-23 in patients with peri-implantitis, which may induce expression of related pro-inflammatory cytokines and may further have a crucial role in tissue repair and reconstruction in pathogenesis of peri-implantitis., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

TH17 cells (interleukin-17 (IL-17)-producing helper T cells) are highly proinflammatory cells that are critical for clearing extracellular pathogens and for inducing multiple autoimmune diseases. IL-23 has a critical role in stabilizing and reinforcing the TH17 phenotype by increasing expression of IL-23 receptor (IL-23R) and endowing TH17 cells with pathogenic effector functions. However, the precise molecular mechanism by which IL-23 sustains the TH17 response and induces pathogenic effector functions has not been elucidated. Here we used transcriptional profiling of developing TH17 cells to construct a model of their signalling network and nominate major nodes that regulate TH17 development. We identified serum glucocorticoid kinase 1 (SGK1), a serine/threonine kinase, as an essential node downstream of IL-23 signalling. SGK1 is critical for regulating IL-23R expression and stabilizing the TH17 cell phenotype by deactivation of mouse Foxo1, a direct repressor of IL-23R expression. SGK1 has been shown to govern Na(+) transport and salt (NaCl) homeostasis in other cells. We show here that a modest increase in salt concentration induces SGK1 expression, promotes IL-23R expression and enhances TH17 cell differentiation in vitro and in vivo, accelerating the development of autoimmunity. Loss of SGK1 abrogated Na(+)-mediated TH17 differentiation in an IL-23-dependent manner. These data demonstrate that SGK1 has a critical role in the induction of pathogenic TH17 cells and provide a molecular insight into a mechanism by which an environmental factor such as a high salt diet triggers TH17 development and promotes tissue inflammation.

Psoriasis is a T-cell-mediated autoimmune disease involving the skin. Two cytokines, interleukin-12 (IL-12) and IL-23 have been shown to play a pivotal role in the pathogenesis of the disease. Ustekinumab (Stelara) is a therapeutic monoclonal antibody (mAb) targeted against the p40 shared subunit of IL-12 and IL-23. Recently the ability of therapeutic proteins (TP) including mAbs that target either cytokines directly (e.g., Pegasys; peginterferon α-2a) or their respective cell surface receptors [e.g., tocilizumab (Actemra); anti IL-6R] to desuppress cytochrome P450 (P450) enzymes in vitro and in the clinic, has been demonstrated. In the present study the ability of IL-12 and IL-23 to suppress multiple P450 enzymes was investigated in vitro using six separate lots of cultured human hepatocytes. Following exposure of 10 ng/ml IL-12 and IL-23 for 48 hours, either alone or in combination, no change in CYP2B6, 2C9, 2C19, or 3A4 gene expression or functional activity was observed. None of the untreated hepatocyte donors showed appreciable expression of the IL-12 or IL-23 receptors. Similar results were seen with whole human liver samples. Exposure of hepatocytes to IL-12 and/or IL-23, known P450 suppressors (IL-6 and tumor necrosis factor-α) or known P450 inducers (β-naphthoflavone, phenobarbital, and rifampicin) did not appreciably alter the expression of the IL-12 and IL-23 receptors either. Finally, in contrast to the positive control IL-6, expression of the acute phase C-reactive protein was unaltered following IL-12 and/or IL-23 treatment. Together, these data suggest a negligible propensity for IL-12 or IL-23 to directly alter P450 enzymes in human hepatocytes.

Hidden low grade inflammation underlines various cardio-metabolic diseases. This type of inflammation is triggered by abnormal reaction to unwanted self products or deregulation of cellular response to cytokines. In the case of atherosclerosis hidden inflammation is induced by modified lipoproteins and develops under control of different cytokines including IL-4 and TGFβ. The key innate immune cells reacting on these factors are monocytes and macrophages. It is well established that monocytes represent a heterogeneous cell population that easily reacts to pathologic changes in the organism. The best studied marker which expression on monocytes is changed in inflammatory conditions is CD16. Although this marker was shown to be associated with various pathologic conditions its specificity is highly questionable. There is an urgent need of identification of new monocyte-expressed biomarkers that may help not only to detect hidden inflammation but also to determine its type. Our analysis of type 2 activation of human monocytes resulted in identification of 3 novel biomarkers for hidden type 2 inflammation. These include stabilin-1, FOXQ1 and IL17RB. These markers are expressed by monocytes/macrophages under stimulation with IL-4 or its combination with TGFβ - 2 cytokines playing important role in atherogenesis. Stabilin-1 was demonstrated on the monocytes in patients with hyperholisterolemia and in macrophages within atherosclerotic lesions. Association of FoxQ1 and IL17RB with atherosclerosis can be deduced from published data but requires experimental confirmation. Functions of all three proteins suggest that they are not only diagnostic markers, but also involved in atherogenesis and can be used as targets for novel therapeutic approaches.

Interleukin-20 (IL-20) is a proinflammatory cytokine involved in rheumatoid arthritis, atherosclerosis, and osteoporosis. However, little is known about the role of IL-20 in oral cancer. We explored the function of IL-20 in the tumor progression of oral cancer. IL-20 expression levels in tumorous and nontumorous oral tissue specimens from 40 patients with four different stages oral cancer were analyzed with immunohistochemistry (IHC) staining and quantitative real-time PCR (qRT-PCR). Expression of IL-20 and its receptor subunits was higher in clinical oral tumor tissue than in nontumorous oral tissue. The role of IL-20 was examined in two oral cancer cell lines (OC-3 and OEC-M1). In vitro, IL-20 promoted TNF-α, IL-1β, MCP-1, CCR4, and CXCR4 and increased proliferation, migration, reactive oxygen species (ROS) production, and colony formation of oral cancer cells via activated STAT3 and AKT/JNK/ERK signals. To evaluate the therapeutic potential of anti-IL-20 monoclonal antibody 7E for treating oral cancer, an ex vivo tumor growth model was used. In vivo, 7E reduced tumor growth and inflammation in oral cancer cells. In conclusion, IL-20 promoted oral tumor growth, migration, and tumor-associated inflammation. Therefore, IL-20 may be a novel target for treating oral cancer, and anti-IL-20 monoclonal antibody 7E may be a feasible therapeutic.

Background: Recently, a new subset of T helper (Th) cell that predominantly secret cytokine interleukin-22 (IL-22) is identified, termed Th22 cells. The Th22 subset has been demonstrated to be involved in immunity and tissue inflammation. However, the existence of Th22 cells and role of IL-22 in acute viral myocarditis (AVMC) remain unknown., Methods: BALB/c mice were intraperitoneally (i.p) infected with CVB3 for establishing AVMC models. Control mice were treated with phosphate-buffered saline (PBS) i.p. On day 14 post injection, frequencies of splenic Th22 cells were determined, productions of IL-22 and expressions of IL-22R (IL-22 receptor) were measured. To further investigate the effects of IL-22, AVMC mice treated with Anti-IL-22 neutralizing antibody were explored. The severity of AVMC were monitored; the frequencies of Th22 cells, the expressions of IL-22 and IL-22R were investigated; in addition to IFN-γ, inflammatory cytokines IL-17, TNF-α, IL-6 as well as IL-1β, were evaluated. Cardiac viral replication were detected., Results: Compared with control group, significant elevations of circulating Th22 cells and IL-22, cardiac protein and mRNA of IL-22, and IL-22R1 were demonstrated in AVMC group. Treatment of AVMC mice with Anti-IL-22 Ab exacerbated the severity of viral myocarditis, verified by lower survival rate, higher HW/BW ratios and cardiac pathological scores. Anti-IL-22 Ab decreased the frequencies of Th22 cells and the levels of IL-22, and increased the expressions of cardiac IL-22R1. Up-regulations of IL-17, IL-6 and TNF-α, down-regulations of IFN-γ proteins and gene expressions in the plasma and myocardium, were observed in Anti-IL-22 Ab group. Furthermore, neutralization of IL-22 significantly promoted cardiac viral replication., Conclusions: Our data indicate that the increased frequencies of IL-22-producing Th22 cells may play an important role in the pathogenesis of CVB3-induced mice AVMC, IL-22 may act as an myocardium-protective cytokine via the IL-22-IL-22R pathway, and suggest that targeting the Th22 cell and IL-22-IL-22R pathway could provide new therapeutic modalities for the treatment of CVB3-induced AVMC.

IL-31 is a T cell-derived cytokine that signals via a heterodimeric receptor composed of IL-31Rα and oncostatin M receptor β. Although several studies have aimed to investigate IL-31-mediated effects, the biological functions of this cytokine are currently not well understood. IL-31 expression correlates with the expression of IL-4 and IL-13 and is associated with atopic dermatitis in humans, indicating that IL-31 is involved in Th2-mediated skin inflammation. Because dendritic cells are the main activators of Th cell responses, we posed the question of whether dendritic cells express the IL-31R complex and govern immune responses triggered by IL-31. In the current study, we report that primary human CD1c(+) as well as monocyte-derived dendritic cells significantly upregulate the IL-31Rα receptor chain upon stimulation with IFN-γ. EMSAs, chromatin immunoprecipitation assays, and small interfering RNA-based silencing assays revealed that STAT1 is the main transcription factor involved in IFN-γ-dependent IL-31Rα expression. Subsequent IL-31 stimulation resulted in a dose-dependent release of proinflammatory mediators, including TNF-α, IL-6, CXCL8, CCL2, CCL5, and CCL22. Because these cytokines are crucially involved in skin inflammation, we hypothesize that IL-31-specific activation of dendritic cells may be part of a positive feedback loop driving the progression of inflammatory skin diseases.

Cytokine components of Th17 pathway play vital roles in human psoriasis. Although much is known about TCR αβ T cells in psoriasis, the role of unconventional T cells, including γδ T cells, is unclear. In this study, using an IL-23 skin injection model of psoriasiform dermatitis in mice, we demonstrate that IL-22, IL-17A, and the IL-23R were highly enriched in a population of CCR6(+), TCR γδ-low expressing (GDL) T cells that accumulated in the epidermis after IL-23 injections. GDL cells were distinct from resident TCR γδ-high, Vγ3(+),CCR6(-) T cells in the epidermis that did not change appreciably in numbers following IL-23 injection. Large numbers of CCR6(+) cells were detected at or above the level of the epidermal basement membrane by confocal microscopy 5 d after repeated IL-23 injections at the same time GDL cells increased in numbers in the epidermis. TCR δ-deficient mice (lacking γδ T cells) exhibited decreased ear swelling and downregulated expression of IL-22 and IL-17A in the epidermis following IL-23 injection. Our data suggest that a subset of γδ T cells play a critical role in IL-23-mediated psoriasiform dermatitis.

Foxp3- and ROR gamma t-expressing cells are involved in acquired immune responses. The change in Foxp3 and ROR gamma t expression in lung tissue and their role in emphysema has not been studied for COPD patients and normal smokers. In the present study, Foxp3 and ROR gamma t were assessed using real-time quantitative polymerase chain reaction and western blotting, and the expression and distribution of Foxp3, IL-17, IL-23R and CCR6 were measured by immunohistochemistry in peripheral lung tissue (10 smokers with COPD, 10 smokers and 10 nonsmokers with normal lung function). Foxp3 expression was lower and ROR gamma t expression was higher in COPD patients when compared with smokers and nonsmokers (all P values were less than 0.001). The ratios of Foxp3/ROR gamma t mRNA and protein were positively correlated to FEV1%pred and negatively correlated to the mean alveoli area. Foxp3(+) cell numbers were decreased, while the number of IL-17(+) cells, IL-23R(+) cells and CCR6(+) cells were increased in the lung alveolar walls of COPD patients compared with normal smokers and nonsmokers (all P values were less than 0.001). The IL-17(+) cell numbers were positively correlated to both CCR6(+) and IL-23R(+) cells. Our data show a decreased Foxp3 expression and an increased ROR gamma t expression in COPD patients and normal smokers that parallels the aggravation of the disease. The IL-17(+)-cell-related cytokines receptors CCR6 and IL-23R had an association with the mechanism of IL-17(+) cell number increasing, which will provide a new immuno-therapeutic target for COPD., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Objective: Fibroblast growth factors (FGF) axis, and in particular FGF2 axis, is an important mitogenic stimulus in endometrial carcinogenesis. hSef is a key inhibitory regulator of FGF signaling and aberrant hSef expression is reported to be present in various human carcinomas. The objective of this study was to investigate the role of hSef in the growth and proliferation of endometrial adenocarcinoma cells and to explore the mechanism that may be involved., Methods: Using western blot analysis, we determined the expression of hSef in Ishikawa cells under different conditions. Using luciferase reporter assays and western blot analysis, we detected the effect of hSef on MAPK/ERK-mediated FGF2 signaling. Using MTT, cell counting and colony formation assays, we analyzed the growth and proliferation of Ishikawa cells under different conditions., Results: We found that the hSef expression was positively regulated by FGF2-induced MAPK/ERK signaling and inversely, hSef expression efficiently inhibited the activity of FGF2-induced MAPK/ERK signaling, indicating the presence of hSef-mediated negative feedback mechanism for FGF signaling in endometrial cancer cells. In addition, we found that MAPK/ERK signaling was essential for the growth and proliferation of endometrial cancer cells in vitro, and hSef expression significantly reduced the cell proliferation., Conclusions: hSef expression can inhibit the growth and proliferation of endometrial cancer cells via acting on the FGF2/MAPK/ERK signaling., (Copyright © 2011 Elsevier Inc. All rights reserved.)

As a novel family of cell surface receptors, triggering receptors expressed on myeloid cells (TREMs) play an important role in inflammatory responses. However, the role of TREMs in the ocular immune system remains unknown. In this study, we examined the expression and function of TREM-1 in Pseudomonas aeruginosa keratitis, one of the most common sight-threatening ocular diseases. TREM-1 was significantly increased in human corneas after P. aeruginosa infection. Consistent with TREM-1 expression at the human ocular surface, TREM-1 levels (mRNA and protein) were also elevated in the infected corneas of C57BL/6 (B6) mice at 1, 3, and 5 days postinfection. To determine whether TREM-1 dictates the outcome of P. aeruginosa keratitis in susceptible mice, TREM-1 signaling in B6 mice was blocked with a soluble mTREM-1/Fc fusion protein. The results indicated that blockade of TREM-1 reduced the severity of corneal disease, polymorphonuclear neutrophil infiltration, Th1/proinflammatory cytokine expression and Toll-like receptor (TLR) activation but enhanced the production of Th2 cytokines, murine β-defensin 2 (mBD2), single Ig interleukin-1R-related molecule (SIGIRR), and ST2. Furthermore, we also used agonistic anti-mTREM-1 antibody to activate TREM-1 signaling in B6 mice and found that TREM-1 activation resulted in worsened disease and earlier corneal perforation in infected B6 mouse corneas and elevated production of proinflammatory cytokines and TLR signaling molecules but reduced expression of mBD2, SIGIRR, and ST2. To the best of our knowledge, this study provides the first evidence that TREM-1 functions as an inflammatory amplifier in P. aeruginosa keratitis by modulating TLR signaling and Th1/Th2 responses.

Background: IL-25 is thought to participate in allergic inflammation by propagating T(h)2-type responses., Objective: To address the hypothesis that allergen provocation increases expression of IL-25 and its receptor IL-25R in the asthmatic bronchial mucosa and skin dermis of atopic subjects., Methods: Sequential single and double immunostaining was used to evaluate the numbers and phenotypes of IL-25 and IL-25R immunoreactive cells in bronchial biopsies from mild atopic subjects with asthma (n = 10) before and 24 hours after allergen inhalation challenge and skin biopsies from atopic subjects (n = 10) up to 72 hours after allergen subepidermal injection., Results: IL-25 immunoreactivity was expressed by a majority of epidermal cells in both organs at baseline and was not further augmented by challenge. IL-25R immunoreactive cells were rare in the epidermis before or after challenge. Allergen challenge was associated with significantly (P < .01) increased expression of IL-25 and IL-25R immunoreactivity in the submucosa of both organs. IL-25 immunoreactivity colocalized with eosinophils, mast cells, and endothelial cells, whereas IL-25R immunoreactivity colocalized with eosinophils, mast cells, endothelial cells, and T lymphocytes. In both organs, correlations were observed between increases in IL-25 expression and the magnitudes of the late-phase allergen-induced clinical responses., Conclusion: Allergen provocation induces functionally relevant, increased expression of IL-25 and its receptor in the asthmatic bronchial mucosa and dermis of sensitized atopic subjects. In addition to T cells, eosinophils, mast cells, and endothelial cells are potential sources and targets of IL-25 in the course of allergic inflammation., (Copyright © 2011 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.)

Aplastic anaemia (AA) is considered as an immune-mediated bone marrow failure syndrome. The mechanism is involved with a variety of immune molecules including interferon-γ (IFN-γ), tumour necrosis factor-α (TNF-α) and interleukins (ILs). IL-27 is a novel member of the IL-12 family, which mediates T cell response and enhances the production of IFN-γ. However, little is known about the role of IL-27 in the development of AA. This study investigated the role of IL-27 and its receptor IL-27R in the pathogenesis of AA. Both the mRNA expression of IL-27/IL-27R subunits in the bone marrow mononuclear cells (BMMNCs) and the levels of IL-27 in the marrow plasma in AA were found to be higher than in controls. Increased IL-27 correlated with the disease severity of AA. Subsequently, we stimulated marrow T lymphocytes with recombinant human (rh)IL-27 and found that rhIL-27 enhanced the production of TNF-α and IFN-γ in both CD4(+) and CD8(+) T lymphocytes from AA patients. We also detected increased TNF-α and IFN-γ in the supernatants of BMMNCs from AA patients after IL-27 stimulation. In conclusion, our data suggest that elevated IL-27 and IL-27-induced TNF-α and IFN-γ overproduction might be involved in the pathogenesis of AA., (© 2011 Blackwell Publishing Ltd.)

Sarcoidosis is a complex granulomatous inflammatory disorder that shares several clinical and pathogenic features with inflammatory bowel disease (IBD). Postulating a common genetic basis of inflammatory diseases, we tested 106 single-nucleotide polymorphisms (SNPs) that are known or have been suggested to be associated with IBD for a potential association with sarcoidosis and its acute and chronic subphenotypes. We genotyped 1,996 German sarcoidosis patients, comprising 648 acutely and 1,161 chronically affected individuals, 2,622 control subjects, and 342 German trios with affected offspring using SNPlex™ technology. The nonsynonymous SNP rs11209026 (Arg381Gln) in the interleukin (IL)-23 receptor (IL23R) gene was associated with chronic sarcoidosis (OR 0.63; p = 5.58×10(-5)), which was supported by the result of a transmission disequilibrium test analysis in the independent family sample (OR 0.50; p = 0.031). Marker rs12035082 located at chromosome 1q24.3 was found to be associated with the acute subphenotype (OR 1.36; p = 6.80×10(-7)) and rs916977 (HERC2 locus; OR 1.30; p = 4.49×10(-5)) was associated with sarcoidosis. Our results highlight the potential importance of the IL-23 signalling pathway for the development of chronic sarcoidosis. The finding links sarcoidosis pathogenesis to other inflammatory conditions and may contribute to new hypotheses on disease mechanisms.

CD4 Th cells producing the proinflammatory cytokine IL-17 (Th17) have been implicated in a number of inflammatory arthritides including the spondyloarthritides. Th17 development is promoted by IL-23. Ankylosing spondylitis, the most common spondyloarthritis (SpA), is genetically associated with both HLA-B27 (B27) and IL-23R polymorphisms; however, the link remains unexplained. We have previously shown that B27 can form H chain dimers (termed B27(2)), which, unlike classical HLA-B27, bind the killer-cell Ig-like receptor KIR3DL2. In this article, we show that B27(2)-expressing APCs stimulate the survival, proliferation, and IL-17 production of KIR3DL2(+) CD4 T cells. KIR3DL2(+) CD4 T cells are expanded and enriched for IL-17 production in the blood and synovial fluid of patients with SpA. Despite KIR3DL2(+) cells comprising a mean of just 15% of CD4 T in the peripheral blood of SpA patients, this subset accounted for 70% of the observed increase in Th17 numbers in SpA patients compared with control subjects. TCR-stimulated peripheral blood KIR3DL2(+) CD4 T cell lines from SpA patients secreted 4-fold more IL-17 than KIR3DL2(+) lines from controls or KIR3DL2(-) CD4 T cells. Strikingly, KIR3DL2(+) CD4 T cells account for the majority of peripheral blood CD4 T cell IL-23R expression and produce more IL-17 in the presence of IL-23. Our findings link HLA-B27 with IL-17 production and suggest new therapeutic strategies in ankylosing spondylitis/SpA.

Th17 CD4 cells are critical to inflammation. Their secretion of IL-17 drives inflammation in human diseases, including inflammatory bowel disease. Differentiation of mature Th17 cells depends on stimulation with IL-6, TGF-β, and IL-21 and the induction of RORγt, but IL-23 is essential to Th17 phenotype, stability, and function. Induction of Th17 cells can be antagonized by IL-4 or IFN-γ, but mechanisms through which terminal differentiation can be inhibited have not been identified. Human IL-23Rα (HuIL23Rα)-chain mRNA transcripts exist that lack exon 9 ("Δ9"); these are translated to a truncated receptor containing the entire external domain. This soluble variant of the HuIL23Rα-chain antagonizes Th17 maturation. It is secreted and present at low levels in the blood. It represents 10% of HuIL23Rα-chain mRNA, binds IL-23 in solution, and inhibits the phosphorylation of STAT3 caused by IL-23. In in vitro Th17 cell differentiation experiments, Δ9 inhibits the production of the Th17-associated cytokines IL-17A and IL-17F. Δ9 does not bind IL-12; thus, it is a specific inhibitor of IL-23 and a modulator of Th17 cells. Our results indicate that this soluble form of HuIL23Rα likely functions to regulate Th17 activity.

T-cell function is dependent on store-operated Ca(2+) influx that is activated by the stromal interaction molecules (STIM) 1 and 2. We show that mice with T-cell-specific deletion of STIM1 or STIM2 are protected from EAE, a mouse model of multiple sclerosis (MS). While STIM1- and STIM2-deficient T cells could be successfully primed by autoantigen, they failed to produce the proinflammatory cytokines IL-17 and IFN-γ. STIM1-deficient T cells showed reduced expression of IL-23R, required for Th17 cell homeostasis, and had impaired chemokine-dependent T-cell migration caused by a lack of chemokine-induced Ca(2+) influx. Autoantigen-specific STIM1- or STIM2-deficient T cells failed to expand and accumulate in the CNS and lymph nodes following adoptive transfer to passively induce EAE, suggesting that autoantigen-specific restimulation or homeostasis of STIM1- and STIM2-deficient T cells are impaired. Combined deletion of both STIM1 and STIM2, previously shown to impair Treg development and function, completely protected mice from EAE. This indicates that, in the absence of Ca(2+) influx, autoreactive T cells are severely dysfunctional rendering Treg dispensable for the prevention of CNS inflammation. Our findings demonstrate that both STIM1 and STIM2 are critical for T-cell function and autoimmunity in vivo., (Copyright © 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Unlabelled: Interleukin-22 (IL-22), a recently identified member of the IL-10 family of cytokines that is produced by Th17 and natural killer cells, plays an important role in controlling bacterial infection, homeostasis, and tissue repair. Here, we tested the effect of IL-22 on alcohol-induced liver injury in a murine model of chronic-binge ethanol feeding. Feeding male C57BL/6 mice with a Lieber-DeCarli diet containing 5% ethanol for 10 days, followed by a single dose of ethanol (5 g/kg body weight) by gavage, induces significant fatty liver and liver injury with peak serum levels of approximately 250 IU/L alanine aminotransferase and 420 IU/L aspartate aminotransferase 9 hours after gavage. Moreover, chronic-binge ethanol administration increases expression of hepatic and serum inflammatory cytokines and hepatic oxidative stress. Using this model, we demonstrate that treatment with IL-22 recombinant protein activates hepatic signal transducer and activator of transcription 3 (STAT3) and ameliorates alcoholic fatty liver, liver injury, and hepatic oxidative stress. Administration with IL-22 adenovirus also prevents alcohol-induced steatosis and liver injury. Deletion of STAT3 in hepatocytes abolishes the hepatoprotection provided by IL-22 in alcoholic liver injury. In addition, IL-22 treatment down-regulates the hepatic expression of fatty acid transport protein, but up-regulates several antioxidant, antiapoptotic, and antimicrobial genes. Finally, expression of IL-22 receptor 1 is up-regulated whereas IL-22 is undetectable in the livers from mice with chronic-binge ethanol feeding or patients with alcoholic hepatitis., Conclusion: Chronic-binge ethanol feeding may be a useful model to study the early stages of alcoholic liver injury. IL-22 treatment could be a potential therapeutic option to ameliorate alcoholic liver disease, due to its antioxidant, antiapoptotic, antisteatotic, proliferative, and antimicrobial effects with the added benefit of potentially few side effects.

Introduction: Th17 cells have been implicated in the pathogenesis of rheumatoid arthritis (RA). The aim of this study was to systematically analyse the phenotype, cytokine profile and frequency of interleukin-17 (IL-17) producing CD4-positive T cells in mononuclear cells isolated from peripheral blood, synovial fluid and synovial tissue of RA patients with established disease, and to correlate cell frequencies with disease activity., Methods: Flow cytometry was used to analyse the phenotype and cytokine production of mononuclear cells isolated from peripheral blood (PBMC) (n = 44), synovial fluid (SFMC) (n = 14) and synovium (SVMC) (n = 10) of RA patients and PBMC of healthy controls (n = 13)., Results: The frequency of IL-17-producing CD4 T cells was elevated in RA SFMC compared with RA PBMC (P = 0.04). However, the frequency of this population in RA SVMC was comparable to that in paired RA PBMC. The percentage of IL-17-producing CD4 T cells coexpressing tumor necrosis factor alpha (TNFα) was significantly increased in SFMC (P = 0.0068). The frequency of IFNγ-producing CD4 T cells was also significantly higher in SFMC than paired PBMC (P = 0.042). The majority of IL-17-producing CD4 T cells coexpressed IFNγ. IL-17-producing CD4 T cells in RA PBMC and SFMC exhibited very little IL-22 or IL-23R coexpression., Conclusions: These findings demonstrate a modest enrichment of IL-17-producing CD4 T cells in RA SFMC compared to PBMC. Th17 cells in SFMC produce more TNFα than their PBMC counterparts, but are not a significant source of IL-22 and do not express IL-23R. However, the percentage of CD4 T cells which produce IL-17 in the rheumatoid joint is low, suggesting that other cells may be alternative sources of IL-17 within the joints of RA patients.

Objective: IL-23 is a pro-inflammatory cytokine proposed to be central to the development of autoimmune disease. We investigated whether IL-23, together with the downstream mediator IL-17A, was present and functional in RA in humans., Methods: RA synovial cells were cultured in the presence or absence of antibodies directed against IL-23p19 or -23R and -17. IL-23, -12, -17, and their receptors, and IL-6, -1beta and TNF-alpha were measured by ELISA and/or PCR., Results: Small amounts of cell-associated IL-23 (median 110 pg/ml) were detected in RA synovial cultures, and found to be functional as IL-23R blockade resulting in a significant inhibition of TNF-alpha (57%), IL-1beta (51%) and IL-6 (30%). However, there was a considerable variability between individual patient samples, and anti-IL-23p19 was found to be considerably less effective. IL-17A protein was detected in approximately 40% of the supernatants and IL-17A blockade, in IL-17A-producing cultures, resulted in a small but significant inhibition of TNF-alpha (38%), IL-1beta (23%) and IL-6 (22%). Addition of recombinant IL-23 to cultures had a variable effect on the spontaneous production of endogenous IL-17A with enhancement observed in some but not all cultures, suggesting that either the low levels of endogenous IL-23 are sufficient to support cytokine production and/or that the relevant Th17 cells were not present., Conclusions: These results suggest that although IL-23 may have pathogenic activity in a proportion of patients with late-stage RA, it is not abundantly produced in this inflammatory tissue, nor does it have a dominant role in all patient tissues analysed.