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Recent advances in human genetics have shed light on the genetic factors contributing to inflammatory diseases, particularly Crohn's disease (CD), a prominent form of inflammatory bowel disease. Certain risk genes associated with CD directly influence cytokine biology and cell-specific communication networks. Current CD therapies primarily rely on anti-inflammatory drugs, which are inconsistently effective and lack strategies for promoting epithelial restoration and mucosal balance. To understand CD's underlying mechanisms, we investigated the link between CD and the FGFR1OP gene, which encodes a centrosome protein. FGFR1OP deletion in mouse intestinal epithelial cells disrupted crypt architecture, resulting in crypt loss, inflammation, and fatality. FGFR1OP insufficiency hindered epithelial resilience during colitis. FGFR1OP was crucial for preserving non-muscle myosin II activity, ensuring the integrity of the actomyosin cytoskeleton and crypt cell adhesion. This role of FGFR1OP suggests that its deficiency in genetically predisposed individuals may reduce epithelial renewal capacity, heightening susceptibility to inflammation and disease., Competing Interests: Declaration of interests R.J.X. is co-founder of Jnana Therapeutics and Celsius Therapeutics, scientific advisory board member at Nestlé, and board director at MoonLake Immunotherapeutics. These organizations had no roles in this study., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Regulatory T (Treg) cells play a critical regulatory role in the immune system by suppressing excessive immune responses and maintaining immune balance. The effective migration of Treg cells is crucial for controlling the development and progression of inflammatory diseases. However, the mechanisms responsible for directing Treg cells into the inflammatory tissue remain incompletely elucidated. In this study, we identified BAF60b, a subunit of switch/sucrose nonfermentable (SWI/SNF) chromatin remodeling complexes, as a positive regulator of Treg cell migration that inhibits the progression of inflammation in experimental autoimmune encephalomyelitis (EAE) and colitis animal models. Mechanistically, transcriptome and genome-wide chromatin-landscaped analyses demonstrated that BAF60b interacts with the transcription factor RUNX1 to promote the expression of CCR9 on Treg cells, which in turn affects their ability to migrate to inflammatory tissues. Our work provides insights into the essential role of BAF60b in regulating Treg cell migration and its impact on inflammatory diseases., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Chronic colonic injury and inflammation pose high risks for field cancerization, wherein injury-associated mutations promote stem cell fitness and gradual clonal expansion. However, the long-term stability of some colitis-associated mutational fields could suggest alternate origins. Here, studies of acute murine colitis reveal a punctuated mechanism of massive, neutral clonal expansion during normal wound healing. Through three-dimensional (3D) imaging, quantitative fate mapping, and single-cell transcriptomics, we show that epithelial wound repair begins with the loss of structural constraints on regeneration, forming fused labyrinthine channels containing epithelial cells reprogrammed to a non-proliferative plastic state. A small but highly proliferative set of epithelial founder progenitor cells (FPCs) subsequently emerges and undergoes extensive cell division, enabling fluid-like lineage mixing and spreading across the colonic surface. Crypt budding restores the glandular organization, imprinting the pattern of clonal expansion. The emergence and functions of FPCs within a critical window of plasticity represent regenerative targets with implications for preneoplasia., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Clostridioides difficile produces toxins that damage the colonic epithelium, causing colitis. Variation in disease severity is poorly understood and has been attributed to host factors and virulence differences between C. difficile strains. We test 23 epidemic ST1 C. difficile clinical isolates for their virulence in mice. All isolates encode a complete Tcd pathogenicity locus and achieve similar colonization densities. However, disease severity varies from lethal to avirulent infections. Genomic analysis of avirulent isolates reveals a 69-bp deletion in the cdtR gene, which encodes a response regulator for binary toxin expression. Deleting the 69-bp sequence in virulent R20291 strain renders it avirulent in mice with reduced toxin gene transcription. Our study demonstrates that a natural deletion within cdtR attenuates virulence in the epidemic ST1 C. difficile isolates without reducing colonization and persistence. Distinguishing strains on the basis of cdtR may enhance the specificity of diagnostic tests for C. difficile colitis., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

This study reveals a previously uncharacterized mechanism to restrict intestinal inflammation via a regulatory RNA transcribed from a noncoding genomic locus. We identified a novel transcript of the lncRNA HOXA11os specifically expressed in the distal colon that is reduced to undetectable levels in colitis. HOXA11os is localized to mitochondria under basal conditions and interacts with a core subunit of complex 1 of the electron transport chain (ETC) to maintain its activity. Deficiency of HOXA11os in colonic myeloid cells results in complex I deficiency, dysfunctional oxidative phosphorylation (OXPHOS), and the production of mitochondrial reactive oxygen species (mtROS). As a result, HOXA11os-deficient mice develop spontaneous intestinal inflammation and are hypersusceptible to colitis. Collectively, these studies identify a new regulatory axis whereby a lncRNA maintains intestinal homeostasis and restricts inflammation in the colon through the regulation of complex I activity., Competing Interests: Declaration of interests This work was supported in part by a sponsored research agreement from Janssen to K.A.F. K.A.F. also serves on an advisory board to Janssen on topics unrelated to the content of this study., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Innate lymphoid cells (ILCs) are considered innate counterparts of adaptive T cells; however, their common and unique transcriptional signatures in pediatric inflammatory bowel disease (pIBD) are largely unknown. Here, we report a dysregulated colonic ILC composition in pIBD colitis that correlates with inflammatory activity, including accumulation of naive-like CD45RA + CD62L - ILCs. Weighted gene co-expression network analysis (WGCNA) reveals modules of genes that are shared or unique across innate and adaptive lymphocytes. Shared modules include genes associated with activation/tissue residency, naivety/quiescence, and antigen presentation. Lastly, nearest-neighbor-based analysis facilitates the identification of "most inflamed" and "least inflamed" lymphocytes in pIBD colon with unique transcriptional signatures. Our study reveals shared and unique transcriptional signatures of colonic ILCs and T cells in pIBD. We also provide insight into the transcriptional regulation of colonic inflammation, deepening our understanding of the potential mechanisms involved in pIBD., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Mesenchymal stem cells (MSCs) are ubiquitous multipotent cells that exhibit significant therapeutic potentials in a variety of disorders. Nevertheless, their clinical efficacy is limited owing to poor survival, low rate of engraftment, and impaired potency upon transplantation. Spheroidal three-dimensional (3D) culture of MSCs (MSC 3D ) has been proven to better preserve their in vivo functional properties. However, the molecular mechanisms underlying the improvement in MSC function by spheroid formation are not clearly understood. NLRP3 inflammasomes, a key component of the innate immune system, have recently been shown to play a role in cell fate decision of MSCs. The present study examined the role of NLRP3 inflammasomes in the survival and potency of MSC spheroids. We found that MSC 3D led to decreased activation of NLRP3 inflammasomes through alleviation of ER stress in an autophagy-dependent manner. Importantly, downregulation of NLRP3 inflammasomes signaling critically contributes to the enhanced survival rate in MSC 3D through modulation of pyroptosis and apoptosis. The critical role of NLRP3 inflammasome suppression in the enhanced therapeutic efficacy of MSC spheroids was further confirmed in an in vivo mouse model of DSS-induced colitis. These findings suggest that 3D culture confers survival and functional advantages to MSCs by suppressing NLRP3 inflammasome activation., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.)

Estrogen receptor β (ERβ) and NOD-like receptor family pyrin domain containing 6 (NLRP6) are highly expressed in intestinal tissues. Loss of ERβ and NLRP6 exacerbate colitis in mouse models; however, the underlying mechanisms are incompletely understood. Here, we report that ERβ directly activates the NLRP6 gene expression via binding to estrogen responsive element of Nlrp6 gene promoter. ERβ also physically interacts with the NLRP6 nucleotide-binding domain and promotes NLRP6 inflammasome assembly. The ERβ-NLRP6 axis then interacts with multiple autophagy-related proteins, including ULK1, BECN1, ATG16L1, LC3B, and p62, and affects the autophagosome biogenesis and autophagic flux. Finally, NLRP6-mediated autophagy suppresses the inflammatory response by promoting the K48-linked polyubiquitination of ASC, Casp-1 p20, IL-1β, TNF-α, and prohibitin-2. Thus, ERβ-NLRP6 direct an anti-inflammatory response by promoting autophagy. Our work uncovers an ERβ-NLRP6-autophagy pathway as a regulatory mechanism that maintains intestinal epithelial cell homeostasis and facilitates tissue repair in colitis., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

Feces are enriched with microRNAs (miRNAs) that shape the gut microbiota. These miRNAs are differentially expressed in the feces of healthy and diseased subjects. However, whether fecal miRNAs in subjects with inflammatory bowel diseases are involved in regulating microbiota composition and whether they have any beneficial effects remains unknown. Here, we studied the fecal microbiome composition and miRNA abundance in mice with dextran sulfate sodium (DSS)-induced colitis and mice at the recovery phase to explore different miRNAs expressed, their relations with microbial abundance, and their effects on colitis. We found that miR-142a-3p expression was significantly increased in the feces of mice recovered from colitis and that it could alleviate disease symptoms in mice treated with DSS in a microbiome-dependent manner. Specifically, miR-142a-3p promoted the growth of Lactobacillus reuteri, which had a high abundance in the feces of mice recovered from colitis, by regulating transcripts of polA and locus tag LREU_RS03575. Moreover, L. reuteri, as well as its metabolite reuterin, could alleviate DSS-induced disease symptoms. These results highlight the role of fecal miR-142a-3p in the prevention of colitis. We propose that the feces of subjects who have recovered from diseases might be enriched with miRNAs with preventive effects against those diseases., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.)

We lack a holistic understanding of the genetic programs orchestrating embryonic colon morphogenesis and governing damage response in the adult. A window into these programs is the transcriptomes of the epithelial and mesenchymal cell populations in the colon. Performing unbiased single-cell transcriptomic analyses of the developing mouse colon at different embryonic stages (embryonic day 14.5 [E14.5], E15.5, and E18.5), we capture cellular and molecular profiles of the stages before, during, and after the appearance of crypt structures, as well as in a model of adult colitis. The data suggest most adult lineages are established by E18.5. We find embryonic-specific gene expression profiles and cell populations that reappear in response to tissue damage. Comparison of the datasets from mice and human colitis suggests the processes are conserved. In this study, we provide a comprehensive single-cell atlas of the developing mouse colon and evidence for the reactivation of embryonic genes in disease., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Both genetic predisposition and environmental factors appear to play a role in inflammatory bowel disease (IBD) development. Genetic studies in humans have linked the interleukin (IL)-23 signaling pathway with IBD, but the environmental factors contributing to disease have remained elusive. Here, we show that the azo dyes Red 40 and Yellow 6, the most abundant food colorants in the world, can trigger an IBD-like colitis in mice conditionally expressing IL-23, or in two additional animal models in which IL-23 expression was augmented. Increased IL-23 expression led to generation of activated CD4 + T cells that expressed interferon-γ and transferred disease to mice exposed to Red 40. Colitis induction was dependent on the commensal microbiota promoting the azo reduction of Red 40 and generation of a metabolite, 1-amino-2-naphthol-6-sulfonate sodium salt. Together these findings suggest that specific food colorants represent novel risk factors for development of colitis in mice with increased IL-23 signaling., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Neutralization of tumor necrosis factor (TNF) represents a widely used therapeutic strategy for autoimmune diseases including inflammatory bowel disease (IBD). However, the fact that many patients with IBD are non-responsive to anti-TNF therapies suggests the need for a better understanding of TNF signaling in IBD. Here, we show that co-deletion of TNF receptor 1 (TNFR1, Tnfrsf1a) in the Il10 -/- spontaneous colitis model exacerbates disease, resulting in very-early-onset inflammation after weaning. The disease can be interrupted by treatment with antibiotics. The single deletion of TNFR1 induces subclinical colonic epithelial dysfunction and mucosal immune abnormalities, including accumulation of neutrophils and depletion of B cells. During the pre-disease period (before weaning), both Tnfr1 -/- and Il10 -/- Tnfr1 -/- animals exhibit impaired expression of pro-inflammatory cytokines compared with wild-type and Il10 -/- controls, respectively. Collectively, these results demonstrate the net anti-inflammatory functions of TNF/TNFR1 signaling through the regulation of colonic immune homeostasis in early life., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Checkpoint blockade with antibodies specific for the PD-1 and CTLA-4 inhibitory receptors can induce durable responses in a wide range of human cancers. However, the immunological mechanisms responsible for severe inflammatory side effects remain poorly understood. Here we report a comprehensive single-cell analysis of immune cell populations in colitis, a common and severe side effect of checkpoint blockade. We observed a striking accumulation of CD8 T cells with highly cytotoxic and proliferative states and no evidence of regulatory T cell depletion. T cell receptor (TCR) sequence analysis demonstrated that a substantial fraction of colitis-associated CD8 T cells originated from tissue-resident populations, explaining the frequently early onset of colitis symptoms following treatment initiation. Our analysis also identified cytokines, chemokines, and surface receptors that could serve as therapeutic targets for colitis and potentially other inflammatory side effects of checkpoint blockade., Competing Interests: Declaration of Interests K.W.W. serves on the scientific advisory board of TCR2 Therapeutics, T-Scan Therapeutics, and Nextechinvest and receives sponsored research funding from Novartis. He is a scientific co-founder of Immunitas Therapeutics. M.D. receives research funding from Novartis and is on the Scientific Advisory Board for Neoleukin Therapeutics. S.K.D. receives research funding from Novartis, Bristol-Myers Squibb, and Eli Lilly. F.S.H. receives research funding from Bristol-Myers Squibb and Novartis; he also consults for Merck, EMD Serono, Novartis, Takeda, Genentech/Roche, Compass Therapeutics, Apricity, Aduro, Sanofi, Pionyr, 7 Hills Pharma, Verastem, Torque, Rheos Kairos, Psioxus Therapeutics, Amgen, and Pieris Pharmaceutical. O.R. receives research support from Merck and is a speaker for activities supported by educational grants from BMS and Merck. He is a consultant for Merck, Celgene, Five Prime, GSK, GFK, Bayer, Roche/Genentech, Puretech, Imvax, and Sobi. In addition, he has patent “Methods of using pembrolizumab and trebananib” pending. J.A.N. has a provisional patent application for spatial quantification of immune cell infiltration in the tumor microenvironment. R.J.S. has received research funding from Merck and Amgen., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Gastrointestinal enterochromaffin cells regulate bone and gut homeostasis via serotonin (5-hydroxytryptamine [5-HT]) production. A recent report suggested that gut microbes regulate 5-HT levels; however, the precise underlying molecular mechanisms are unexplored. Here, we reveal that the cation channel Piezo1 in the gut acts as a sensor of single-stranded RNA (ssRNA) governing 5-HT production. Intestinal epithelium-specific deletion of mouse Piezo1 profoundly disturbed gut peristalsis, impeded experimental colitis, and suppressed serum 5-HT levels. Because of systemic 5-HT deficiency, conditional knockout of Piezo1 increased bone formation. Notably, fecal ssRNA was identified as a natural Piezo1 ligand, and ssRNA-stimulated 5-HT synthesis from the gut was evoked in a MyD88/TRIF-independent manner. Colonic infusion of RNase A suppressed gut motility and increased bone mass. These findings suggest gut ssRNA as a master determinant of systemic 5-HT levels, indicating the ssRNA-Piezo1 axis as a potential prophylactic target for treatment of bone and gut disorders., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Inflammatory bowel disease (IBD) is a chronic inflammatory disease associated with increased risk of gastrointestinal cancers. We whole-genome sequenced 446 colonic crypts from 46 IBD patients and compared these to 412 crypts from 41 non-IBD controls from our previous publication on the mutation landscape of the normal colon. The average mutation rate of affected colonic epithelial cells is 2.4-fold that of healthy colon, and this increase is mostly driven by acceleration of mutational processes ubiquitously observed in normal colon. In contrast to the normal colon, where clonal expansions outside the confines of the crypt are rare, we observed widespread millimeter-scale clonal expansions. We discovered non-synonymous mutations in ARID1A, FBXW7, PIGR, ZC3H12A, and genes in the interleukin 17 and Toll-like receptor pathways, under positive selection in IBD. These results suggest distinct selection mechanisms in the colitis-affected colon and that somatic mutations potentially play a causal role in IBD pathogenesis., Competing Interests: Declaration of Interests C.A.A. is a paid consultant for Genomics plc and Bristol-Myers Squibb. All other authors declare no competing interests., (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)

IRF5 polymorphisms are associated with multiple immune-mediated diseases, including ulcerative colitis. IRF5 contributions are attributed to its role in myeloid lineages. How T cell-intrinsic IRF5 contributes to inflammatory outcomes is not well understood. We identify a previously undefined key role for T cell-intrinsic IRF5. In mice, IRF5 in CD4 + T cells promotes Th1- and Th17-associated cytokines and decreases Th2-associated cytokines. IRF5 is required for the optimal assembly of the TCR-initiated signaling complex and downstream signaling at early times, and at later times binds to promoters of Th1- and Th17-associated transcription factors and cytokines. IRF5 also regulates chemokine receptor-initiated signaling and, in turn, T cell migration. In vivo, IRF5 in CD4 + T cells enhances the severity of experimental colitis. Importantly, human CD4 + T cells from high IRF5-expressing disease-risk genetic carriers demonstrate increased chemokine-induced migration and Th1/Th17 cytokines and reduced Th2-associated and anti-inflammatory cytokines. These data demonstrate key roles for T cell-intrinsic IRF5 in inflammatory outcomes., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)

Regulatory T cells (Tregs) are pivotal for immune suppression. Cellular metabolism is important for Treg homeostasis and function. However, the exact role of mitochondrial respiration in Tregs remains elusive. Mitochondrial transcription factor A (Tfam) is essential for mitochondrial respiration and controls mitochondrial DNA replication, transcription, and packaging. Here, we show that genetic ablation of Tfam in Tregs impairs Treg maintenance in non-lymphoid tissues in the steady state and in tumors. Tfam-deficient Tregs have reduced proliferation and Foxp3 expression upon glucose deprivation in vitro. Tfam deficiency preferentially affects gene activation in Tregs through regulation of DNA methylation, with enhanced methylation in the TSDR of the Foxp3 locus. Deletion of Tfam in Tregs affects Treg homing and stability, resulting in tissue inflammation in colitis, but enhances tumor rejection. Thus, our work reveals a critical role of Tfam-mediated mitochondrial respiration in Tregs to regulate inflammation and anti-tumor immunity., (Copyright © 2019 The Author(s). Published by Elsevier Inc. All rights reserved.)

Several large-scale genome-wide association studies genetically linked IRGM to Crohn's disease and other inflammatory disorders in which the IRGM appears to have a protective function. However, the mechanism by which IRGM accomplishes this anti-inflammatory role remains unclear. Here, we reveal that IRGM/Irgm1 is a negative regulator of the NLRP3 inflammasome activation. We show that IRGM expression, which is increased by PAMPs, DAMPs, and microbes, can suppress the pro-inflammatory responses provoked by the same stimuli. IRGM/Irgm1 negatively regulates IL-1β maturation by suppressing the activation of the NLRP3 inflammasome. Mechanistically, we show that IRGM interacts with NLRP3 and ASC and hinders inflammasome assembly by blocking their oligomerization. Further, IRGM mediates selective autophagic degradation of NLRP3 and ASC. By suppressing inflammasome activation, IRGM/Irgm1 protects from pyroptosis and gut inflammation in a Crohn's disease experimental mouse model. This study for the first time identifies the mechanism by which IRGM is protective against inflammatory disorders., (Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.)

β2-integrins promote neutrophil recruitment to infected tissues and are crucial for host defense. Neutrophil recruitment is defective in leukocyte adhesion deficiency type-1 (LAD1), a condition caused by mutations in the CD18 (β2-integrin) gene. Using a model of Citrobacter rodentium (CR)-induced colitis, we show that CD18 -/- mice display increased intestinal damage and systemic bacterial burden, compared to littermate controls, ultimately succumbing to infection. This phenotype is not attributed to defective neutrophil recruitment, as it is shared by CXCR2 -/- mice that survive CR infection. CR-infected CD18 -/- mice feature prominent upregulation of IL-17 and downregulation of IL-22. Exogenous IL-22 administration, but not endogenous IL-17 neutralization, protects CD18 -/- mice from lethal colitis. β2-integrin expression on macrophages is mechanistically linked to Rac1/ROS-mediated induction of noncanonical-NLRP3 (nucleotide-binding domain, leucine-rich-containing family, pyrin domain-containing-3) inflammasome-dependent IL-1β production, which promotes ILC3-derived IL-22. Therefore, β2-integrins are required for protective IL-1β-dependent IL-22 responses in colitis, and the identified mechanism may underlie the association of human LAD1 with colitis., (Copyright © 2019 The Author(s). Published by Elsevier Inc. All rights reserved.)

Inflammatory bowel disease (IBD) results from a dysregulated interaction between the microbiota and a genetically susceptible host. Genetic studies have linked TNFSF15 polymorphisms and its protein TNF-like ligand 1A (TL1A) with IBD, but the functional role of TL1A is not known. Here, we found that adherent IBD-associated microbiota induced TL1A release from CX3CR1 + mononuclear phagocytes (MNPs). Using cell-specific genetic deletion models, we identified an essential role for CX3CR1 + MNP-derived TL1A in driving group 3 innate lymphoid cell (ILC3) production of interleukin-22 and mucosal healing during acute colitis. In contrast to this protective role in acute colitis, TL1A-dependent expression of co-stimulatory molecule OX40L in MHCII + ILC3s during colitis led to co-stimulation of antigen-specific T cells that was required for chronic T cell colitis. These results identify a role for ILC3s in activating intestinal T cells and reveal a central role for TL1A in promoting ILC3 barrier immunity during colitis., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Intestinal mesenchymal cells play essential roles in epithelial homeostasis, matrix remodeling, immunity, and inflammation. But the extent of heterogeneity within the colonic mesenchyme in these processes remains unknown. Using unbiased single-cell profiling of over 16,500 colonic mesenchymal cells, we reveal four subsets of fibroblasts expressing divergent transcriptional regulators and functional pathways, in addition to pericytes and myofibroblasts. We identified a niche population located in proximity to epithelial crypts expressing SOX6, F3 (CD142), and WNT genes essential for colonic epithelial stem cell function. In colitis, we observed dysregulation of this niche and emergence of an activated mesenchymal population. This subset expressed TNF superfamily member 14 (TNFSF14), fibroblastic reticular cell-associated genes, IL-33, and Lysyl oxidases. Further, it induced factors that impaired epithelial proliferation and maturation and contributed to oxidative stress and disease severity in vivo. Our work defines how the colonic mesenchyme remodels to fuel inflammation and barrier dysfunction in IBD., (Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.)

The adaptor protein CARD9 links detection of fungi by surface receptors to the activation of the NF-κB pathway. Mice deficient in CARD9 exhibit dysbiosis and are more susceptible to colitis. Here we examined the impact of Card9 deficiency in the development of colitis-associated colon cancer (CAC). Treatment of Card9 -/- mice with AOM-DSS resulted in increased tumor loads as compared to WT mice and in the accumulation of myeloid-derived suppressor cells (MDSCs) in tumor tissue. The impaired fungicidal functions of Card9 -/- macrophages led to increased fungal loads and variation in the overall composition of the intestinal mycobiota, with a notable increase in C. tropicalis. Bone marrow cells incubated with C. tropicalis exhibited MDSC features and suppressive functions. Fluconazole treatment suppressed CAC in Card9 -/- mice and was associated with decreased MDSC accumulation. The frequency of MDSCs in tumor tissues of colon cancer patients correlated positively with fungal burden, pointing to the relevance of this regulatory axis in human disease., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Roquin proteins preclude spontaneous T cell activation and aberrant differentiation of T follicular helper (Tfh) or T helper 17 (Th17) cells. Here we showed that deletion of Roquin-encoding alleles specifically in regulatory T (Treg) cells also caused the activation of conventional T cells. Roquin-deficient Treg cells downregulated CD25, acquired a follicular Treg (Tfr) cell phenotype, and suppressed germinal center reactions but could not protect from colitis. Roquin inhibited the PI3K-mTOR signaling pathway by upregulation of Pten through interfering with miR-17∼92 binding to an overlapping cis-element in the Pten 3' UTR, and downregulated the Foxo1-specific E3 ubiquitin ligase Itch. Loss of Roquin enhanced Akt-mTOR signaling and protein synthesis, whereas inhibition of PI3K or mTOR in Roquin-deficient T cells corrected enhanced Tfh and Th17 or reduced iTreg cell differentiation. Thereby, Roquin-mediated control of PI3K-mTOR signaling prevents autoimmunity by restraining activation and differentiation of conventional T cells and specialization of Treg cells., (Copyright © 2017 Elsevier Inc. All rights reserved.)

In mammalian cells, histone deacetylase (HDAC) and Sirtuin (SIRT) are two families responsible for removing acetyl groups from acetylated proteins. Here, we describe protein deacetylation coupled with deacetylimination as a function of lysyl oxidase (LOX) family members. LOX-like 3 (Loxl3) associates with Stat3 in the nucleus to deacetylate and deacetyliminate Stat3 on multiple acetyl-lysine sites. Surprisingly, Loxl3 N-terminal scavenger receptor cysteine-rich (SRCR) repeats, rather than the C-terminal oxidase catalytic domain, represent the major deacetylase/deacetyliminase activity. Loxl3-mediated deacetylation/deacetylimination disrupts Stat3 dimerization, abolishes Stat3 transcription activity, and restricts cell proliferation. In Loxl3-/- mice, Stat3 is constitutively acetylated and naive CD4 + T cells are potentiated in Th17/Treg cell differentiation. When overexpressed, the SRCR repeats from other LOX family members can catalyze protein deacetylation/deacetylimination. Thus, our findings delineate a hitherto-unknown mechanism of protein deacetylation and deacetylimination catalyzed by lysyl oxidases., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Inflammation, which is a highly regulated host response against danger signals, may be harmful if it is excessive and deregulated. Ideally, anti-inflammatory therapy should autonomously commence as soon as possible after the onset of inflammation, should be controllable by a physician, and should not systemically block beneficial immune response in the long term. We describe a genetically encoded anti-inflammatory mammalian cell device based on a modular engineered genetic circuit comprising a sensor, an amplifier, a "thresholder" to restrict activation of a positive-feedback loop, a combination of advanced clinically used biopharmaceutical proteins, and orthogonal regulatory elements that linked modules into the functional device. This genetic circuit was autonomously activated by inflammatory signals, including endogenous cecal ligation and puncture (CLP)-induced inflammation in mice and serum from a systemic juvenile idiopathic arthritis (sIJA) patient, and could be reset externally by a chemical signal. The microencapsulated anti-inflammatory device significantly reduced the pathology in dextran sodium sulfate (DSS)-induced acute murine colitis, demonstrating a synthetic immunological approach for autonomous anti-inflammatory therapy., (Copyright © 2017 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.)

Myeloid-derived suppressor cells (MDSCs) are well known regulators of regulatory T cells (Treg cells); however, the direct regulation of MDSCs by Treg cells has not been well characterized. We find that colitis caused by functional deficiency of Treg cells leads to altered expansion and reduced function of MDSCs. During differentiation of MDSCs in vitro from bone marrow cells, Treg cells enhanced MDSC function and controlled their differentiation through a mechanism involving transforming growth factor-β (TGF-β). TGF-β-deficient Treg cells were not able to regulate MDSC function in an experimentally induced model of colitis. Finally, we evaluated the therapeutic effect of TGF-β-mediated in-vitro-differentiated MDSCs on colitis. Adoptive transfer of MDSCs that differentiated with TGF-β led to better colitis prevention than the transfer of MDSCs that differentiated without TGF-β. Our results demonstrate an interaction between Treg cells and MDSCs that contributes to the regulation of MDSC proliferation and the acquisition of immunosuppressive functions., (Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.)

High mucosal and fecal concentrations of the antimicrobial siderophore-binding peptide Lipocalin-2 (Lcn2) are observed in inflammatory bowel disease. However, Lcn2 function in chronic intestinal inflammation remains unclear. Here, we demonstrate that Lcn2 protects from early-onset colitis and spontaneous emergence of right-sided colonic tumors resulting from IL-10 deficiency. Exacerbated inflammation in Lcn2(-/-)/Il10(-/-) mice is driven by IL-6, which also controls tumorigenesis. Lcn2(-/-)/Il10(-/-) mice exhibit profound alterations in gut microbial composition, which contributes to inflammation and tumorigenesis, as demonstrated by the transmissibility of the phenotype and protection conferred by antibiotics. Specifically, facultative pathogenic Alistipes spp. utilize enterobactin as iron source, bloom in Lcn2(-/-)/Il10(-/-) mice, and are sufficient to induce colitis and right-sided tumors when transferred into Il10(-/-) mice. Our results demonstrate that Lcn2 protects against intestinal inflammation and tumorigenesis associated with alterations in the microbiota., (Copyright © 2016 Elsevier Inc. All rights reserved.)

CARD9 is a central component of anti-fungal innate immune signaling via C-type lectin receptors, and several immune-related disorders are associated with CARD9 alterations. Here, we used a rare CARD9 variant that confers protection against inflammatory bowel disease as an entry point to investigating CARD9 regulation. We showed that the protective variant of CARD9, which is C-terminally truncated, acted in a dominant-negative manner for CARD9-mediated cytokine production, indicating an important role for the C terminus in CARD9 signaling. We identified TRIM62 as a CARD9 binding partner and showed that TRIM62 facilitated K27-linked poly-ubiquitination of CARD9. We identified K125 as the ubiquitinated residue on CARD9 and demonstrated that this ubiquitination was essential for CARD9 activity. Furthermore, we showed that similar to Card9-deficient mice, Trim62-deficient mice had increased susceptibility to fungal infection. In this study, we utilized a rare protective allele to uncover a TRIM62-mediated mechanism for regulation of CARD9 activation., (Copyright © 2015 Elsevier Inc. All rights reserved.)

The intestinal epithelial barrier plays a critical role in the mucosal immunity. However, it remains largely unknown how the epithelial barrier is maintained after damage. Here we show that growth factor FGF2 synergized with interleukin-17 (IL-17) to induce genes for repairing of damaged epithelium. FGF2 or IL-17 deficiency resulted in impaired epithelial proliferation, increased pro-inflammatory microbiota outgrowth, and consequently worse pathology in a DSS-induced colitis model. The dysregulated microbiota in the model induced transforming growth factor beta 1 (TGFβ1) expression, which in turn induced FGF2 expression mainly in regulatory T cells. Act1, an essential adaptor in IL-17 signaling, suppressed FGF2-induced ERK activation through binding to adaptor molecule GRB2 to interfere with its association with guanine nucleotide exchange factor SOS1. Act1 preferentially bound to IL-17 receptor complex, releasing its suppressive effect on FGF2 signaling. Thus, microbiota-driven FGF2 and IL-17 cooperate to repair the damaged intestinal epithelium through Act1-mediated direct signaling cross-talk., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Indoleamine 2,3-dioxygenase 1 (Ido1) is a rate-limiting enzyme that catalizes the degradation of tryptophan along the kynurenine pathway. Here, we show that Ido1 activity sustains an immunostimulatory potential through inhibition of interleukin (Il)10. In atherosclerosis, Ido1-dependent inhibition of Il10 translates into disease exacerbation. The resistance of Ido1-deficient mice to enhanced immune activation is broken in Ido1/Il10 double-deficient mice, which show exaggerated immune responses and develop severe spontaneous colitis. We demonstrate that Ido1 activity is required for the regulation of Il10 and that kynurenic acid (Kna), an Ido1-derived metabolite, is responsible for reduced Il10 production through activation of a cAMP-dependent pathway and inhibition of Erk1/2 phosphorylation. Resupplementation of Ido1-deficient mice with Kna limits Il10 expression and promotes atherosclerosis. In human atherosclerotic lesions, increased levels of Kna are associated with an unstable plaque phenotype, and its blood levels predict death and recurrent myocardial infarction in patients with coronary artery disease., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Regulatory T (Treg) cells are essential for maintenance of immune homeostasis. Here we found that hydrogen sulfide (H2S) was required for Foxp3(+) Treg cell differentiation and function and that H2S deficiency led to systemic autoimmune disease. H2S maintained expression of methylcytosine dioxygenases Tet1 and Tet2 by sulfhydrating nuclear transcription factor Y subunit beta (NFYB) to facilitate its binding to Tet1 and Tet2 promoters. Transforming growth factor-β (TGF-β)-activated Smad3 and interleukin-2 (IL-2)-activated Stat5 facilitated Tet1 and Tet2 binding to Foxp3. Tet1 and Tet2 catalyzed conversion of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) in Foxp3 to establish a Treg-cell-specific hypomethylation pattern and stable Foxp3 expression. Consequently, Tet1 and Tet2 deletion led to Foxp3 hypermethylation, impaired Treg cell differentiation and function, and autoimmune disease. Thus, H2S promotes Tet1 and Tet2 expression, which are recruited to Foxp3 by TGF-β and IL-2 signaling to maintain Foxp3 demethylation and Treg-cell-associated immune homeostasis., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Plasmacytoid dendritic cells (pDCs) are primary producers of type I interferon (IFN) in response to viruses. The IFN-producing capacity of pDCs is regulated by specific inhibitory receptors, yet none of the known receptors are conserved in evolution. We report that within the human immune system, receptor protein tyrosine phosphatase sigma (PTPRS) is expressed specifically on pDCs. Surface PTPRS was rapidly downregulated after pDC activation, and only PTPRS(-) pDCs produced IFN-α. Antibody-mediated PTPRS crosslinking inhibited pDC activation, whereas PTPRS knockdown enhanced IFN response in a pDC cell line. Similarly, murine Ptprs and the homologous receptor phosphatase Ptprf were specifically co-expressed in murine pDCs. Haplodeficiency or DC-specific deletion of Ptprs on Ptprf-deficient background were associated with enhanced IFN response of pDCs, leukocyte infiltration in the intestine and mild colitis. Thus, PTPRS represents an evolutionarily conserved pDC-specific inhibitory receptor, and is required to prevent spontaneous IFN production and immune-mediated intestinal inflammation., (Copyright © 2015 Elsevier Inc. All rights reserved.)

The interleukin-17 (IL-17) family of cytokines has emerged as a critical player in inflammatory diseases. Among them, IL-25 has been shown to be important in allergic inflammation and protection against parasitic infection. Here we have demonstrated that IL-17B, a poorly understood cytokine, functions to inhibit IL-25-driven inflammation. IL-17B and IL-25, both binding to the interleukin-17 receptor B (IL-17RB), were upregulated in their expression after acute colonic inflammation. Individual inhibition of these cytokines revealed opposing functions in colon inflammation: IL-25 was pathogenic but IL-17B was protective. Similarly opposing phenotypes were observed in Citrobacter rodentium infection and allergic asthma. Moreover, IL-25 was found to promote IL-6 production from colon epithelial cells, which was inhibited by IL-17B. Therefore, our data demonstrate that IL-17B is an anti-inflammatory cytokine in the IL-17 family., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Missense mutations in the nucleotide-binding oligomerization domain (NOD)-like receptor pyrin domain containing family of gene 12 (Nlrp12) are associated with periodic fever syndromes and atopic dermatitis in humans. Here, we have demonstrated a crucial role for NLRP12 in negatively regulating pathogenic T cell responses. Nlrp12(-/-) mice responded to antigen immunization with hyperinflammatory T cell responses. Furthermore, transfer of CD4(+)CD45RB(hi)Nlrp12(-/-) T cells into immunodeficient mice led to more severe colitis and atopic dermatitis. NLRP12 deficiency did not, however, cause exacerbated ascending paralysis during experimental autoimmune encephalomyelitis (EAE); instead, Nlrp12(-/-) mice developed atypical neuroinflammatory symptoms that were characterized by ataxia and loss of balance. Enhanced T-cell-mediated interleukin-4 (IL-4) production promotes the development of atypical EAE disease in Nlrp12(-/-) mice. These results define an unexpected role for NLRP12 as an intrinsic negative regulator of T-cell-mediated immunity and identify altered NF-κB regulation and IL-4 production as key mediators of NLRP12-associated disease., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Systemic 1,25(OH)2D3 treatment ameliorating murine inflammatory bowel diseases (IBD) could not be applied to patients because of hypercalcemia. We tested the hypothesis that increasing 1,25(OH)2D3 synthesis locally by targeting delivery of the 1α-hydroxylase gene (CYP27B1) to the inflamed bowel would ameliorate IBD without causing hypercalcemia. Our targeting strategy is the use of CD11b(+)/Gr1(+) monocytes as the cell vehicle and a macrophage-specific promoter (Mac1) to control CYP27B1 expression. The CD11b(+)/Gr1(+) monocytes migrated initially to inflamed colon and some healthy tissues in dextran sulfate sodium (DSS) colitis mice; however, only the migration of monocytes to the inflamed colon was sustained. Adoptive transfer of Gr1(+) monocytes did not cause hepatic injury. Infusion of Mac1-CYP27B1-modified monocytes increased body weight gain, survival, and colon length, and expedited mucosal regeneration. Expression of pathogenic Th17 and Th1 cytokines (interleukin (IL)-17a and interferon (IFN)-α) was decreased, while expression of protective Th2 cytokines (IL-5 and IL-13) was increased, by the treatment. This therapy also enhanced tight junction gene expression in the colon. No hypercalcemia occurred following this therapy. In conclusion, we have for the first time obtained proof-of-principle evidence for a novel monocyte-based adoptive CYP27B1 gene therapy using a mouse IBD model. This strategy could be developed into a novel therapy for IBD and other autoimmune diseases.

Nonpathogenic-invasive Escherichia coli (InvColi) bacteria are suitable for genetic transfer into mammalian cells and may act as a vehicle for RNA Interference (RNAi) in vivo. Cyclooxygenase-2 (COX-2) is overexpressed in ulcerative colitis (UC) and Crohn's disease (CD), two inflammatory conditions of the colon and small intestine grouped as inflammatory bowel disease (IBD). We engineered InvColi strains for anti-COX-2 RNAi (InvColi(shCOX2)), aiming to investigate the in vivo feasibility of a novel COX-2 silencing strategy in a murine model of colitis induced by dextran sulfate sodium (DSS). Enema administrations of InvColi(shCOX2) in DSS-treated mice led to COX-2 downregulation, colonic mucosa preservation, reduced colitis disease activity index (DAI) and increased mice survival. Moreover, DSS/InvColi(shCOX2)-treated mice showed lower levels of circulating pro-inflammatory cytokines and a reduced colitis-associated shift of gut microbiota. Considering its effectiveness and safety, we propose our InvColi(shCOX2) strategy as a promising tool for molecular therapy in intestinal inflammatory diseases.

In cancer, immune cells can play conflicting roles, either protective, by elimination of tumor cells during immune surveillance, or detrimental, by promoting carcinogenesis during inflammation. We report here that the thymus-specific serine protease (TSSP), which is involved in CD4(+) T cell maturation in the thymus, exerts a tumor suppressor activity. Mice genetically deficient for TSSP are highly prone to spontaneous cancer development. The absence of TSSP also increases the rate of induced colitis-associated colorectal (CAC) tumor formation, through exacerbated colon inflammation. Adoptive transfer of T cells in various combinations (CD4(+) and CD8(+) from wild-type and/or knockout mice) into T cell-deficient mice showed that the TSSP-deficient CD4(+) T cell compartment promotes tumor development, associated with high levels of the cytokine IL-17A. Inhibition of IL-17A during CAC tumor formation prevents the increased carcinogenesis and colic immune disequilibrium observed in TSSP-deficient mice. Therefore, our data demonstrate that antitumoral immune surveillance requires thymic TSSP-driven production of CD4(+) T cells contributing to inflammatory balance., (Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.)

LGR5 and BMI1 mark intestinal stem cells in crypt base columnar cells and +4 position cells, respectively, but characterization of functional markers in these cell populations is limited. ID1 maintains the stem cell potential of embryonic, neural, and long-term repopulating hematopoietic stem cells. Here, we show in both human and mouse intestine that ID1 is expressed in cycling columnar cells, +4 position cells, and transit-amplifying cells in the crypt. Lineage tracing revealed ID1+ cells to be self-renewing, multipotent stem/progenitor cells that are responsible for the long-term renewal of the intestinal epithelium. Single ID1+ cells can generate long-lived organoids resembling mature intestinal epithelium. Complete knockout of Id1 or selective deletion of Id1 in intestinal epithelium or in LGR5+ stem cells sensitizes mice to chemical-induced colon injury. These experiments identify ID1 as a marker for intestinal stem/progenitor cells and demonstrate a role for ID1 in maintaining the potential for repair in response to colonic injury.

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.

The β-galactoside-binding protein galectin-9 is critical in regulating the immune response, but the mechanism by which it functions remains unclear. We have demonstrated that galectin-9 is highly expressed by induced regulatory T cells (iTreg) and was crucial for the generation and function of iTreg cells, but not natural regulatory T (nTreg) cells. Galectin-9 expression within iTreg cells was driven by the transcription factor Smad3, forming a feed-forward loop, which further promoted Foxp3 expression. Galectin-9 increased iTreg cell stability and function by directly binding to its receptor CD44, which formed a complex with transforming growth factor-β (TGF-β) receptor I (TGF-βRI), and activated Smad3. Galectin-9 signaling was further found to regulate iTreg cell induction by dominantly acting through the CNS1 region of the Foxp3 locus. Our data suggest that exogenous galectin-9, in addition to being an effector molecule for Treg cells, acts synergistically with TGF-β to enforce iTreg cell differentiation and maintenance., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The intestinal epithelium harbors large populations of activated and memory lymphocytes, yet these cells do not cause tissue damage in the steady state. We investigated how intestinal T cell effector differentiation is regulated upon migration to the intestinal epithelium. Using gene loss- and gain-of-function strategies, as well as reporter approaches, we showed that cooperation between the transcription factors T-bet and Runx3 resulted in suppression of conventional CD4(+) T helper functions and induction of an intraepithelial lymphocyte (IEL) program that included expression of IEL markers such as CD8αα homodimers. Interferon-γ sensing and T-bet expression by CD4(+) T cells were both required for this program, which was distinct from conventional T helper differentiation but shared by other IEL populations, including TCRαβ(+)CD8αα(+) IELs. We conclude that the gut environment provides cues for IEL maturation through the interplay between T-bet and Runx3, allowing tissue-specific adaptation of mature T lymphocytes., (Copyright © 2014 Elsevier Inc. All rights reserved.)

CUEDC2, a CUE-domain-containing protein, modulates inflammation, but its involvement in tumorigenesis is still poorly understood. Here, we report that CUEDC2 is a key regulator of macrophage function and critical for protection against colitis-associated tumorigenesis. CUEDC2 expression is dramatically upregulated during macrophage differentiation, and CUEDC2 deficiency results in excessive production of proinflammatory cytokines. The level of CUEDC2 in macrophages is modulated by miR- 324-5p. We find that Cuedc2 KO mice are more susceptible to dextran-sodium-sulfate-induced colitis, and macrophage transplantation results suggest that the increased susceptibility results from the dysfunction of macrophages lacking CUEDC2. Furthermore, we find that Cuedc2 KO mice are more prone to colitis-associated cancer. Importantly, CUEDC2 expression is almost undetectable in macrophages in human colon cancer, and this decreased CUEDC2 expression is associated with high levels of interleukin-4 and miR-324-5p. Thus, CUEDC2 plays a crucial role in modulating macrophage function and is associated with both colitis and colon tumorigenesis., (Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.)

The adoptive transfer of regulatory T cells (Tregs) offers a promising strategy to combat pathologies that are characterized by aberrant immune activation, including graft rejection and autoinflammatory diseases. Expression of a chimeric antigen receptor (CAR) gene in Tregs redirects them to the site of autoimmune activity, thereby increasing their suppressive efficiency while avoiding systemic immunosuppression. Since carcinoembryonic antigen (CEA) has been shown to be overexpressed in both human colitis and colorectal cancer, we treated CEA-transgenic mice that were induced to develop colitis with CEA-specific CAR Tregs. Two disease models were employed: T-cell-transfer colitis as well as the azoxymethane-dextran sodium sulfate model for colitis-associated colorectal cancer. Systemically administered CEA-specific (but not control) CAR Tregs accumulated in the colons of diseased mice. In both model systems, CEA-specific CAR Tregs suppressed the severity of colitis compared to control Tregs. Moreover, in the azoxymethane-dextran sodium sulfate model, CEA-specific CAR Tregs significantly decreased the subsequent colorectal tumor burden. Our data demonstrate that CEA-specific CAR Tregs exhibit a promising potential in ameliorating ulcerative colitis and in hindering colorectal cancer development. Collectively, this study provides a proof of concept for the therapeutic potential of CAR Tregs in colitis patients as well as in other autoimmune inflammatory disorders.

Immune cells sense microbial products through Toll-like receptors (TLR), which trigger host defense responses including type 1 interferons (IFNs) secretion. A coding polymorphism in the protein tyrosine phosphatase nonreceptor type 22 (PTPN22) gene is a susceptibility allele for human autoimmune and infectious disease. We report that Ptpn22 selectively regulated type 1 IFN production after TLR engagement in myeloid cells. Ptpn22 promoted host antiviral responses and was critical for TLR agonist-induced, type 1 IFN-dependent suppression of inflammation in colitis and arthritis. PTPN22 directly associated with TNF receptor-associated factor 3 (TRAF3) and promotes TRAF3 lysine 63-linked ubiquitination. The disease-associated PTPN22W variant failed to promote TRAF3 ubiquitination, type 1 IFN upregulation, and type 1 IFN-dependent suppression of arthritis. The findings establish a candidate innate immune mechanism of action for a human autoimmunity "risk" gene in the regulation of host defense and inflammation., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Dendritic cell (DC) activation is essential for the induction of immune defense against pathogens, yet needs to be tightly controlled to avoid chronic inflammation and exaggerated immune responses. Here, we identify a mechanism of immune homeostasis by which adaptive immunity, once triggered, tempers DC activation and prevents overreactive immune responses. T cells, once activated, produced Protein S (Pros1) that signaled through TAM receptor tyrosine kinases in DCs to limit the magnitude of DC activation. Genetic ablation of Pros1 in mouse T cells led to increased expression of costimulatory molecules and cytokines in DCs and enhanced immune responses to T cell-dependent antigens, as well as increased colitis. Additionally, PROS1 was expressed in activated human T cells, and its ability to regulate DC activation was conserved. Our results identify a heretofore unrecognized, homeostatic negative feedback mechanism at the interface of adaptive and innate immunity that maintains the physiological magnitude of the immune response., (Copyright © 2013 Elsevier Inc. All rights reserved.)

A20 is an anti-inflammatory protein linked to multiple human autoimmune diseases and lymphomas. A20 possesses a deubiquitinating motif and a zinc finger, ZF4, that binds ubiquitin and supports its E3 ubiquitin ligase activity. To understand how these activities mediate A20's physiological functions, we generated two lines of gene-targeted mice, abrogating either A20's deubiquitinating activity (Tnfaip3(OTU) mice) or A20's ZF4 (Tnfaip3(ZF4) mice). Both Tnfaip3(OTU) and Tnfaip3(ZF4) mice exhibited increased responses to TNF and sensitivity to colitis. A20's C103 deubiquitinating motif restricted both K48- and K63-linked ubiquitination of receptor interacting protein 1 (RIP1). A20's ZF4 was required for recruiting A20 to ubiquitinated RIP1. A20(OTU) proteins and A20(ZF4) proteins complemented each other to regulate RIP1 ubiquitination and NFκB signaling normally in compound mutant Tnfaip3(OTU/ZF4) cells. This complementation involved homodimerization of A20 proteins, and we have defined an extensive dimerization interface in A20. These studies reveal how A20 proteins collaborate to restrict TNF signaling., (Copyright © 2013 Elsevier Inc. All rights reserved.)

The tumor suppressor p53 is frequently mutated in human cancer. Common mutant p53 (mutp53) isoforms can actively promote cancer through gain-of-function (GOF) mechanisms. We report that mutp53 prolongs TNF-α-induced NF-κB activation in cultured cells and intestinal organoid cultures. Remarkably, when exposed to dextran sulfate sodium, mice harboring a germline p53 mutation develop severe chronic inflammation and persistent tissue damage, and are highly prone to inflammation-associated colon cancer. This mutp53 GOF is manifested by rapid onset of flat dysplastic lesions that progress to invasive carcinoma with mutp53 accumulation and augmented NF-κB activation, faithfully recapitulating features frequently observed in human colitis-associated colorectal cancer (CAC). These findings might explain the early appearance of p53 mutations in human CAC., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Inflammatory bowel disease is an important risk factor for colorectal cancer. We show that sphingosine-1-phosphate (S1P) produced by upregulation of sphingosine kinase 1 (SphK1) links chronic intestinal inflammation to colitis-associated cancer (CAC) and both are exacerbated by deletion of Sphk2. S1P is essential for production of the multifunctional NF-κB-regulated cytokine IL-6, persistent activation of the transcription factor STAT3, and consequent upregulation of the S1P receptor, S1PR1. The prodrug FTY720 decreased SphK1 and S1PR1 expression and eliminated the NF-κB/IL-6/STAT3 amplification cascade and development of CAC, even in Sphk2(-/-) mice, and may be useful in treating colon cancer in individuals with ulcerative colitis. Thus, the SphK1/S1P/S1PR1 axis is at the nexus between NF-κB and STAT3 and connects chronic inflammation and CAC., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Mice lacking junctional adhesion molecule A (JAM-A, encoded by F11r) exhibit enhanced intestinal epithelial permeability, bacterial translocation, and elevated colonic lymphocyte numbers, yet do not develop colitis. To investigate the contribution of adaptive immune compensation in response to increased intestinal epithelial permeability, we examined the susceptibility of F11r(-/-)Rag1(-/-) mice to acute colitis. Although negligible contributions of adaptive immunity in F11r(+/+)Rag1(-/-) mice were observed, F11r(-/-)Rag1(-/-) mice exhibited increased microflora-dependent colitis. Elimination of T cell subsets and cytokine analyses revealed a protective role for TGF-β-producing CD4(+) T cells in F11r(-/-) mice. Additionally, loss of JAM-A resulted in elevated mucosal and serum IgA that was dependent upon CD4(+) T cells and TGF-β. Absence of IgA in F11r(+/+)Igha(-/-) mice did not affect disease, whereas F11r(-/-)Igha(-/-) mice displayed markedly increased susceptibility to acute injury-induced colitis. These data establish a role for adaptive immune-mediated protection from acute colitis under conditions of intestinal epithelial barrier compromise., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Colitis results from breakdown of homeostasis between intestinal microbiota and the mucosal immune system, with both environmental and genetic influencing factors. Flagellin receptor TLR5-deficient mice (T5KO) display elevated intestinal proinflammatory gene expression and colitis with incomplete penetrance, providing a genetically sensitized system to study the contribution of microbiota to driving colitis. Both colitic and noncolitic T5KO exhibited transiently unstable microbiotas, with lasting differences in colitic T5KO, while their noncolitic siblings stabilized their microbiotas to resemble wild-type mice. Transient high levels of proteobacteria, especially enterobacteria species including E. coli, observed in close proximity to the gut epithelium were a striking feature of colitic microbiota. A Crohn's disease-associated E. coli strain induced chronic colitis in T5KO, which persisted well after the exogenously introduced bacterial species had been eliminated. Thus, an innate immune deficiency can result in unstable gut microbiota associated with low-grade inflammation, and harboring proteobacteria can drive and/or instigate chronic colitis., (Copyright © 2012 Elsevier Inc. All rights reserved.)