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Background: Intestinal epithelial cell (IEC) mitochondrial dysfunction involvement in inflammatory bowel diseases (IBD), including Crohn's disease affecting the small intestine, is emerging in recent studies. As the interface between the self and the gut microbiota, IECs serve as hubs of bidirectional cross-talk between host and luminal microbiota. However, the role of mitochondrial-microbiota interaction in the ileum is largely unexplored. Prohibitin 1 (PHB1), a chaperone protein of the inner mitochondrial membrane required for optimal electron transport chain function, is decreased during IBD. We previously demonstrated that mice deficient in PHB1 specifically in IECs (Phb1 i∆IEC ) exhibited mitochondrial impairment, Paneth cell defects, gut microbiota dysbiosis, and spontaneous inflammation in the ileum (ileitis). Mice deficient in PHB1 in Paneth cells (epithelial secretory cells of the small intestine; Phb1 ∆PC ) also exhibited mitochondrial impairment, Paneth cell defects, and spontaneous ileitis. Here, we determined whether this phenotype is driven by Phb1 deficiency-associated ileal microbiota alterations or direct effects of loss of PHB1 in host IECs., Results: Depletion of gut microbiota by broad-spectrum antibiotic treatment in Phb1 ∆PC or Phb1 i∆IEC mice revealed a necessary role of microbiota to cause ileitis. Using germ-free mice colonized with ileal microbiota from Phb1-deficient mice, we show that this microbiota could not independently induce ileitis without host mitochondrial dysfunction. The luminal microbiota phenotype of Phb1 i∆IEC mice included a loss of the short-chain fatty acid butyrate. Supplementation of butyrate in Phb1-deficient mice ameliorated Paneth cell abnormalities and ileitis. Phb1-deficient ileal enteroid models suggest deleterious epithelial-intrinsic responses to ileal microbiota that were protected by butyrate., Conclusions: These results suggest a mutual and essential reinforcing interplay of gut microbiota and host IEC, including Paneth cell, mitochondrial health in influencing ileitis. Restoration of butyrate is a potential therapeutic option in Crohn's disease patients harboring epithelial cell mitochondrial dysfunction. Video Abstract., (© 2023. The Author(s).)

Introduction: The Unfolded Protein Response, a mechanism triggered by the cell in response to Endoplasmic reticulum stress, is linked to inflammatory responses. Our aim was to identify novel Unfolded Protein Response-mechanisms that might be involved in triggering or perpetuating the inflammatory response carried out by the Intestinal Epithelial Cells in the context of Inflammatory Bowel Disease., Methods: We analyzed the transcriptional profile of human Intestinal Epithelial Cell lines treated with an Endoplasmic Reticulum stress inducer (thapsigargin) and/or proinflammatory stimuli. Several genes were further analyzed in colonic biopsies from Ulcerative Colitis patients and healthy controls. Lastly, we generated Caco-2 cells lacking HMGCS2 by CRISPR Cas-9 and analyzed the functional implications of its absence in Intestinal Epithelial Cells., Results: Exposure to a TLR ligand after thapsigargin treatment resulted in a powerful synergistic modulation of gene expression, which led us to identify new genes and pathways that could be involved in inflammatory responses linked to the Unfolded Protein Response. Key differentially expressed genes in the array also exhibited transcriptional alterations in colonic biopsies from active Ulcerative Colitis patients, including NKG2D ligands and the enzyme HMGCS2. Moreover, functional studies showed altered metabolic responses and epithelial barrier integrity in HMGCS2 deficient cell lines., Conclusion: We have identified new genes and pathways that are regulated by the Unfolded Protein Response in the context of Inflammatory Bowel Disease including HMGCS2 , a gene involved in the metabolism of Short Chain Fatty Acids that may have an important role in intestinal inflammation linked to Endoplasmic Reticulum stress and the resolution of the epithelial damage., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Martín-Adrados, Wculek, Fernández-Bravo, Torres-Ruiz, Valle-Noguera, Gomez-Sánchez, Hernández-Walias, Ferreira, Corraliza, Sancho, Esteban, Rodriguez-Perales, Cruz-Adalia, Nakaya, Salas, Bernardo, Campos-Martín, Martínez-Zamorano, Muñoz-López, Gómez del Moral, Cubero, Blumberg and Martínez-Naves.)

Post-translational modifications in intestinal epithelial cells (IECs) allow for precise control in intestinal homeostasis, the breakdown of which may precipitate the pathological damage and inflammation in inflammatory bowel disease. The O-linked β-N-acetylglucosamine (O-GlcNAc) modification on intracellular proteins controls diverse biological processes; however, its roles in intestinal homeostasis are still largely unexplored. Here, we found that levels of protein O-GlcNAcylation and the expression of O-GlcNAc transferase (OGT), the enzyme adding the O-GlcNAc moiety, were reduced in IECs in human IBD patients. Deletion of OGT specifically in IECs resulted in disrupted epithelial barrier, microbial dysbiosis, Paneth cell dysfunction, and intestinal inflammation in mice. Using fecal microbiota transplantation in mice, we demonstrated that microbial dysbiosis although was insufficient to induce spontaneous inflammation but exacerbated chemical-induced colitis. Paneth cell-specific deletion of OGT led to Paneth cell dysfunction, which might predispose mice to chemical-induced colitis. On the other hand, the augmentation of O-GlcNAc signaling by inhibiting O-GlcNAcase, the enzyme removing O-GlcNAcylation, alleviated chemical-induced colitis. Our data reveal that protein O-GlcNAcylation in IECs controls key regulatory mechanisms to maintain mucosal homeostasis., (© 2018 The Authors. Published under the terms of the CC BY 4.0 license.)

Inflammatory bowel disease (IBD) results from a continuum of complex interactions between a quartet of host-derived and external elements that involve various aspects of the intestinal microbiota, the immune system that is centered around the intestinal epithelial cell barrier, the genetic composition of the host and specific environmental factors. Recent studies into the complexity of these arrangements increasingly support the syndromic nature of this disorder and involve a wide range of interacting biologic pathways that affect innate immunity, adaptive immunity, endoplasmic reticulum stress and autophagy as well as metabolic pathways associated with cellular homeostasis. It is further likely that all of the aforementioned host factors including the microbiota, which is as much a part of ourselves as is any organ system, are under the influence of yet-to-be-understood environmental factors that predispose to and precipitate IBD. Notwithstanding the importance of genetic predisposition, these environmental influences are no doubt central to disease pathogenesis in light of the rapid emergence of IBD throughout the world and assumption of disease in migrating populations from low to high risk environments. It can thus be anticipated that environmental factors that modify the risk for development of IBD have the common attribute of affecting the relationship between the commensal microbiota and the immune system in a manner that intersects with the functionally relevant immuno-genetic pathways, and potentially modifies them through epigenetic effects, in a manner that are uniquely operative within a particular syndromic context of IBD and occur sequentially and in a reiterative fashion, perhaps beginning in early life., (© 2016 S. Karger AG, Basel.)

The human host and the intestinal microbiota co-exist in a mutually beneficial relationship, which contributes to host and microbial metabolism as well as maturation of the host's immune system, among many other pathways (Tremaroli and Backhed, 2012; Hooper et al., 2012). At mucosal surfaces, and particularly in the intestine, the commensal microbiota provides 'colonization resistance' to invading pathogens and maintains homeostasis through microbial regulation of mucosal innate and adaptive immunity (Renz et al., 2012). Recent evidence suggests that natural killer T cells (NKT cells), a subgroup of lipid-reactive T cells, play central roles in bidirectional interactions between the host and the commensal microbiota, which govern intestinal homeostasis and prevent inflammation. Here, we provide a brief overview of recently identified pathways of commensal microbial regulation of NKT cells, discuss feedback mechanisms of NKT cell-dependent control of microbial colonization and composition, and highlight the critical role of host-microbial cross-talk for prevention of NKT cell-dependent mucosal inflammation., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Purpose of Review: The purpose of this study is to provide an overview of the role of endoplasmic reticulum (ER) stress and the unfolded protein response (UPR) in inflammatory bowel disease (IBD)., Recent Findings: Human genetic studies have identified several UPR-related genes and autophagy-related genes as IBD risk loci. Impairment of each branch of the UPR causes spontaneous enteritis or creates higher susceptibility for intestinal inflammation in model systems. Deficiency of either UPR or autophagy in small intestinal epithelial cells promotes each other's compensatory engagement, which is especially prominent in Paneth cells such that, in the absence of both, severe spontaneous enteritis emerges., Summary: Interactions between the UPR and autophagy exhibit critical synergistic interactions within the intestinal epithelium and especially Paneth cells that are of considerable importance to the maintenance of homeostasis. When dysfunctional in the Paneth cell, spontaneous inflammation can emerge that may extend beyond the epithelium providing direct experimental evidence that subsets of Crohn's disease may emanate from primary Paneth cell disturbances.

Results of recent genetic and immunologic studies have brought to the forefront several biologic pathways that allow for a better understanding of the mechanisms of tissue homeostasis, on the one hand, and inflammatory bowel disease (IBD) on the other. The explosion of research activity as a result of these newly identified targets is bringing the pathogenesis of these complex disorders into focus as well as creating new therapeutic opportunities. The greatest advances with perhaps the largest impact on our understanding of the etiology of Crohn's disease are those related to bacterial sensing, such as through nucleotide-binding oligomerization domain-containing protein 2 (NOD2) and its relationships to autophagy and the unfolded protein response as a consequence of endoplasmic reticulum stress. Interestingly, it appears as though these pathways, which are rooted in microbial sensing and regulation, are interrelated. Genetic studies have also renewed interest in previously studied pathways in IBD, such as the formation and function of the inflammasome and its relationship to interleukin (IL) 1-beta signaling. With the recent success of therapeutic agents designed to block tumor necrosis factor, the IL-12/23 pathways, and lymphocyte homing, insights have been gained into the biologic relevance and impact of these various inflammatory pathways in IBD. In this review, the exciting recent advances in these biologic pathways of IBD are discussed, particularly in light of their therapeutic relevance.

The unfolded protein response (UPR) is a signaling pathway from the endoplasmic reticulum (ER) to the nucleus that protects cells from the stress caused by misfolded or unfolded proteins [1, 2]. As such, ER stress is an ongoing challenge for all cells given the central biologic importance of secretion as part of normal physiologic functions. This is especially the case for cells that are highly dependent upon secretory function as part of their major duties. Within mucosal tissues, the intestinal epithelium is especially dependent upon an intact UPR for its normal activities [3]. This review will discuss the UPR and the special role that it provides in the functioning of the intestinal epithelium and, when dysfunctional, its implications for understanding mucosal homeostasis and intestinal inflammation, as occurs in inflammatory bowel disease (IBD)., (Copyright © 2011 Elsevier Inc. All rights reserved.)

The intestinal epithelium forms a highly active functional interface between the relatively sterile internal body surfaces and the enormously complex and diverse microbiota that are contained within the lumen. Genetic models that allow for manipulation of genes specifically in the intestinal epithelium have provided an avenue to understand the diverse set of pathways whereby intestinal epithelial cells (IECs) direct the immune state of the mucosa associated with homeostasis versus either productive or non-productive inflammation as occurs during enteropathogen invasion or inflammatory bowel disease (IBD), respectively. These pathways include the unfolded protein response (UPR) induced by stress in the endoplasmic reticulum (ER), autophagy, a self-cannibalistic pathway important for intracellular bacterial killing and proper Paneth cell function as well as the interrelated functions of NOD2/NF-κB signaling which also regulate autophagy induction. Multiple genes controlling these IEC pathways have been shown to be genetic risk factors for human IBD. This highlights the importance of these pathways not only for proper IEC function but also suggesting that IECs may be one of the cellular originators of organ-specific and systemic inflammation as in IBD.

Increasing evidence has emerged that supports an important intersection between 3 fundamental cell biologic pathways in the pathogenesis of inflammatory bowel disease. These include the intersection between autophagy, as revealed by the original identification of ATG16L1 and IRGM as major genetic risk factors for Crohn's disease, and intracellular bacterial sensing, as shown by the importance of NOD2 in autophagy induction upon bacterial entry into the cell. A pathway closely linked to autophagy and innate immunity is the unfolded protein response, initiated by endoplasmic reticulum stress due to the accumulation of misfolded proteins, which is genetically related to ulcerative colitis and Crohn's disease (XBP1 and ORMDL3). Hypomorphic ATG16L1, NOD2, and X box binding protein-1 possess the common attribute of profoundly affecting Paneth cells, specialized epithelial cells at the bottom of intestinal crypts involved in antimicrobial function. Together with their functional juxtaposition in the environmentally exposed intestinal epithelial cell, their remarkable functional convergence on Paneth cells and their behavior in response to environmental factors, including microbes, these 3 pathways are of increasing importance to understanding the pathogenesis of inflammatory bowel disease. Moreover, in conjunction with studies that model deficient nuclear factor-κB function, these studies suggest a central role for altered intestinal epithelial cell function as one of the earliest events in the development of inflammatory bowel disease., (Copyright © 2011 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Patients with inflammatory bowel disease (IBD) have increased numbers of human tryptase-β (hTryptase-β)-positive mast cells (MCs) in the gastrointestinal tract. The amino acid sequence of mouse mast cell protease (mMCP)-6 is most similar to that of hTryptase-β. We therefore hypothesized that this mMCP, or the related tryptase mMCP-7, might have a prominent proinflammatory role in experimental colitis. The dextran sodium sulfate (DSS) and trinitrobenzene sulfonic acid (TNBS) colitis models were used to evaluate the differences between C57BL/6 (B6) mouse lines that differ in their expression of mMCP-6 and mMCP-7 with regard to weight loss, colon histopathology, and endoscopy scores. Microarray analyses were performed, and confirmatory real-time PCR, ELISA, and/or immunohistochemical analyses were carried out on a number of differentially expressed cytokines, chemokines, and matrix metalloproteinases (MMPs). The mMCP-6-null mice that had been exposed to DSS had significantly less weight loss as well as significantly lower pathology and endoscopy scores than similarly treated mMCP-6-expressing mice. This difference in colitis severity was confirmed endoscopically in the TNBS-treated mice. Evaluation of the distal colon segments revealed that numerous proinflammatory cytokines, chemokines that preferentially attract neutrophils, and MMPs that participate in the remodeling of the ECM were all markedly increased in the colons of DSS-treated WT mice relative to untreated WT mice and DSS-treated mMCP-6-null mice. Collectively, our data show that mMCP-6 (but not mMCP-7) is an essential MC-restricted mediator in chemically induced colitis and that this tryptase acts upstream of many of the factors implicated in IBD.

Background: Breakdown of the gut mucosal barrier during chronic HIV infection allows translocation of bacterial products such as lipopolysaccharides (LPS) from the gut into the circulation. Microbial translocation also occurs in inflammatory bowel disease (IBD). IBD serological markers are useful in the diagnosis of IBD and to differentiate between Crohn's disease (CD) and ulcerative colitis (UC). Here, we evaluate detection of IBD serological markers in HIV-infected patients with advanced disease and their relationship to HIV disease markers., Methods: IBD serological markers (ASCA, pANCA, anti-OmpC, and anti-CBir1) were measured by ELISA in plasma from AIDS patients (n = 26) with low CD4 counts (<300 cells/µl) and high plasma LPS levels, and results correlated with clinical data. For meta-analysis, relevant data were abstracted from 20 articles., Results: IBD serological markers were detected in approximately 65% of AIDS patients with evidence of microbial translocation. An antibody pattern consistent with IBD was detected in 46%; of these, 75% had a CD-like pattern. Meta-analysis of data from 20 published studies on IBD serological markers in CD, UC, and non-IBD control subjects indicated that IBD serological markers are detected more frequently in AIDS patients than in non-IBD disease controls and healthy controls, but less frequently than in CD patients. There was no association between IBD serological markers and HIV disease markers (plasma viral load and CD4 counts) in the study cohort., Conclusions: IBD serological markers may provide a non-invasive approach to monitor HIV-related inflammatory gut disease. Further studies to investigate their clinical significance in HIV-infected individuals are warranted.

Purpose of Review: To provide an overview of the emerging role of cellular stress responses in inflammatory bowel disease (IBD)., Recent Findings: The unfolded protein response (UPR) is a primitive cellular pathway that is engaged when responding to endoplasmic reticulum stress and regulates autophagy. Highly secretory cells such as Paneth cells and goblet cells in the intestines are particularly susceptible to endoplasmic reticulum stress and are exceedingly dependent upon a properly functioning UPR to maintain cellular viability and homeostasis. Primary genetic abnormalities within the components of the UPR (e.g. XBP1, ARG2, ORMDL3), genes that encode proteins reliant upon a robust secretory pathway (e.g. MUC2, HLAB27) and environmental factors that create disturbances in the UPR (e.g. microbial products and inflammatory cytokines) are important factors in the primary development and/or perpetuation of intestinal inflammation., Summary: Endoplasmic reticulum stress is an important new pathway involved in the development of intestinal inflammation associated with IBD and likely other intestinal inflammatory disorders.

Inflammatory bowel disease (IBD) has long been known to arise from the interplay between host and environmental factors. From this, a picture is currently emerging in which IBD is likely the result of a continuum of diseases that range from mono- and oligogenically inherited familial forms at one extreme to sporadic forms at the other extreme, which are polygenic in origin and strongly influenced by environmental factors and especially those of infectious origin. The recent expansion of knowledge on the genetic underpinning of IBD has revealed several converging and inter-related functional host pathways that are central to the pathogenesis of these disorders. These include pathways such as autophagy, intracellular bacterial sensing and the unfolded protein response, which play specific roles at the interface between the host and the highly complex microbial communities within the intestines. As such they focus on the functional relationship between the intestinal epithelium and the unique microbial and immune environments along its luminal and abluminal surfaces. Thus, the genetic and environmental factors which are relevant to IBD seem to have the common property of influencing disease by virtue of their specific impact upon the functional relationship between these microbial communities and the intestinal immune system., (Copyright © 2010 S. Karger AG, Basel.)

Insights into inflammatory bowel disease (IBD) are advancing rapidly owing to immunologic investigations of a plethora of animal models of intestinal inflammation, ground-breaking advances in the interrogation of diseases that are inherited as complex genetic traits, and the development of culture-independent methods to define the composition of the intestinal microbiota. These advances are bringing a deeper understanding to the genetically determined interplay between the commensal microbiota, intestinal epithelial cells, and the immune system and the manner in which this interplay might be modified by relevant environmental factors in the pathogenesis of IBD. This review examines these interactions and, where possible, potential lessons from IBD-directed, biologic therapies that may allow for elucidation of pathways that are central to disease pathogenesis in humans.

The intestinal epithelial cell (IEC) is increasingly recognized to play a prominent role as an important intermediary between the commensal microbiota and the intestinal immune system. Moreover, it is now recognized that intestinal inflammation in inflammatory bowel disease (IBD) may arise primarily from IEC dysfunction due to unresolved endoplasmic reticulum (ER) stress as a consequence of genetic disruption of X box binding protein-1 function. In addition to primary (genetic) abnormalities of the unfolded protein response, a variety of secondary (inflammation and environmental) factors are also likely to be important regulators of ER stress. ER stress pathways are also well known to regulate (and be regulated by) autophagy pathways. Therefore, the host's ability to manage ER stress is likely to be a major pathway in the pathogenesis of intestinal inflammation that arises primarily from the IEC. Herein we discuss ER stress in the IEC as both an originator and perpetuator of intestinal inflammation in IBD.

The unfolded protein response as a consequence of endoplasmic reticulum (ER) stress has recently been implicated as a novel mechanism that may lead to inflammatory bowel disease (IBD). Impairment of proper ER stress resolution in highly secretory Paneth and, to a lesser extent, goblet cells within the epithelium can primarily lead to intestinal inflammation. An inability to manage ER stress may not only be a primary originator of intestinal inflammation as exemplified by genetic polymorphisms in XBP1 that are associated with IBD but also a perpetuator of inflammation when ER stress is induced secondarily to inflammatory mediators or microbial factors. Furthermore, ER stress pathways may interact with other processes that lead to IBD, notably autophagy.

Over the past decade a major hypothesis has emerged for the etiopathogenesis of inflammatory bowel disease (IBD). This hypothesis proposes that IBD represents a dysregulated mucosal immune response to antigens derived from the commensal microbiota in a genetically susceptible host that initially derives from innate immune abnormalities leading to an excessive proinflammatory cytokine derived from CD4+ T cells (T-helper 1, T-helper 2, and T-helper 17 cytokines) over and above the response that is normally associated with tolerance and immunoregulation derived from T-regulatory cells. Given that the genetic predisposition has increasingly been recognized to affect the regulation of innate and adaptive immunity, intestinal epithelial cell physiologic barrier function and the potential inappropriate access of antigens to the mucosal immune system through this dysfunctional barrier function, a key point in understanding IBD pathophysiology is to understand the immunoregulatory pathways associated with the intestinal immune system as they apply to IBD. Therefore, immunogenetic pathways associated with innate and adaptive immunity, the cytokines secreted by innate and adaptive immune cells, the epithelial factors and leukocyte factors that are associated with inflammation and structures on the endothelium that regulate the recruitment of leukocytes define potential pathways that may be amenable to therapeutic manipulation in IBD.

Inflammatory bowel disease (IBD) has been attributed to aberrant mucosal immunity to the intestinal microbiota. The transcription factor XBP1, a key component of the endoplasmic reticulum (ER) stress response, is required for development and maintenance of secretory cells and linked to JNK activation. We hypothesized that a stressful environmental milieu in a rapidly proliferating tissue might instigate a proinflammatory response. We report that Xbp1 deletion in intestinal epithelial cells (IECs) results in spontaneous enteritis and increased susceptibility to induced colitis secondary to both Paneth cell dysfunction and an epithelium that is overly reactive to inducers of IBD such as bacterial products (flagellin) and TNFalpha. An association of XBP1 variants with both forms of human IBD (Crohn's disease and ulcerative colitis) was identified and replicated (rs35873774; p value 1.6 x 10(-5)) with novel, private hypomorphic variants identified as susceptibility factors. Hence, intestinal inflammation can originate solely from XBP1 abnormalities in IECs, thus linking cell-specific ER stress to the induction of organ-specific inflammation.

Purpose of Review: Adaptive immune mechanisms are inherently involved in the pathogenesis of inflammatory bowel disease. This review summarizes the main discoveries made in 2007 within this field., Recent Findings: CD4 T cells secreting interleukin-17 (T helper type 17) cells have emerged as a key effector population driving colitis in animal models previously associated with exaggerated T helper type 1 responses. With regard to T-regulatory cells, a novel suppressive cytokine (interleukin-35) and induction of apoptosis as a means to exert suppression have been identified. The importance of specific chemokine receptors and integrins in effector and T-regulatory cell function in colitis has been recognized., Summary: An improved understanding of adaptive immune mechanisms, on which manifold genetic and environmental traits might converge, and which ultimately mediate intestinal inflammation in inflammatory bowel disease, holds promise for novel effective therapeutic intervention.

Natural killer T (NKT) cells are a small subset of unconventional T cells that recognize lipid antigens presented by the nonclassical major histocompatibility complex (MHC) class I molecule CD1d. NKT cells are involved in the host response to a variety of microbial pathogens and likely commensals. In the intestine, invariant and noninvariant NKT cells can be found among intraepithelial lymphocytes and in the lamina propria. Activation of intestinal NKT cells by CD1d-expressing intestinal epithelial cells and professional antigen-presenting cells may contribute to induction of oral tolerance and protection from mucosal infections. On the other hand, sustained and uncontrolled activation of NKT cells may play a pivotal role in the pathogenesis of inflammatory bowel disease. Here we review the current literature on intestinal NKT cells and their function in the intestine in health and disease.

Natural killer T (NKT) cells are a subset of lymphocytes that express cell surface molecules of both conventional T cells and natural killer cells and share the features of both innate and adaptive immune cells. NKT cells have been proposed to make both protective and pathogenic contributions to inflammatory bowel diseases (IBD). On the one hand, recent studies have shown that these cells are involved in the maintenance of mucosal homeostasis. On the other, NKT cells were shown to play a pathogenic role in human ulcerative colitis. Similar contrasting data have been generated in murine models of IBD. Whether the apparent differences in NKT response patterns depend on variations in NKT antigens and/or on the presence of specific subsets of mucosal NKT cells remains to be elucidated. In this article we review the current literature on intestinal NKT cells and their roles in IBD pathogenesis. Specifically, the nomenclature, NKT antigens, and immune mechanisms of NKT cells within the intestinal mucosa are discussed.

Inflammatory bowel disease is an immune-mediated intestinal inflammatory condition that is associated with an increase in autoantibodies that bind to epithelial cells. However, it is unknown whether the epithelial cell-derived products that are recognized by such autoantibodies are involved in the pathogenic process. Through a combined antigen-screening approach utilizing humoral and cellular immune responses, we identify herein an epithelial lectin, galectin-4, that specifically stimulates IL-6 production by CD4(+) T cells. Interestingly, the reactivity of CD4(+) T cells to galectin-4 is precisely elicited under intestinal inflammatory conditions. The galectin-4-mediated production of IL-6 is MHC class II independent and induced by PKCtheta-associated pathway through the immunological synapse. The galectin-4-mediated stimulation of CD4(+) T cells is shown to exacerbate chronic colitis and delay the recovery from acute intestinal injury. These studies identify the presence of an immunogenic, endogenous lectin in the intestine and dissect the biological role of lectin/CD4(+) T cell interactions under inflammatory conditions.

Background & Aims: Peroxisome proliferator-activated receptor gamma (PPAR gamma) is one of the nuclear receptors that plays a central role in adipocyte differentiation and insulin sensitivity. PPAR gamma has also recently been recognized as an endogenous regulator of intestinal inflammation. However, its levels are decreased during chronic inflammation in human and mice, thus limiting PPAR gamma ligand therapy during established disease. We sought to determine whether this decrease in PPAR gamma could be counteracted by a gene therapy approach., Methods: We characterized PPAR gamma levels in experimental colitis associated with dextran sodium sulfate administration to mice. In this model, the therapeutic benefits of PPAR gamma gene therapy using a replication-deficient adenovirus vector expressing PPAR gamma (Ad-PPAR gamma) was assessed., Results: PPAR gamma protein levels were decreased in whole colonic tissue, lamina propria lymphocytes, and peritoneal exudate cells during the course of colitis. PPAR gamma gene delivery using Ad-PPAR gamma restored responsiveness to a PPAR gamma ligand, resulting in marked amelioration of tissue inflammation associated with the colitis, which included attenuation of intercellular adhesion molecule-1, cyclooxygenase-2 and tumor necrosis factor-alpha expression., Conclusions: Our results suggest that gene delivery of PPAR gamma can be used to restore and/or enhance endogenous anti-inflammatory processes that are normally operative in mammalian tissues such as in the colon.

Background: It has recently been suggested that Crohn's Disease (CD) is associated with an exaggerated T helper 1 cytokine response as manifest by increased production of interleukin-12 (IL-12) and interferon-gamma (IFN-gamma). Epstein-Barr virus-induced gene 3 (EBI3) encodes a 34-kDa glycoprotein that is 27% identical to the p40 unit of IL-12 and has recently been reported to be up-regulated in ulcerative colitis (UC)., Aim: To determine whether mucosal expression of IL-12 p40 or EBI3 correlates with inflammatory bowel disease (IBD)., Patients/methods: mRNA expression in colonic mucosa from patients with UC, Crohn's disease (CD) and non-IBD controls was measured by reverse-transcribed real-time polymerase chain reaction (PCR)., Results: EBI3 was significantly increased in both involved and uninvolved colonic mucosa in patients with UC. Although IL-12 p40 was increased in some patients with CD relative to non-IBD controls, the increase was not statistically significant. However, 5-aminosalicylic acid (5-ASA) use was significantly correlated with reduced IL-12 p40 levels in the patients with CD, but not in UC cases. A similar reduction was not seen in 5-ASA-treated UC patients., Conclusion: IL-12 p40 expression in CD is heterogeneous. In contrast, expression of the IL-12 p40 homologue, EBI3, is up-regulated in nearly all UC cases and in a subset of CD.

In addition to the classical MHC class I and class II molecules, human intestinal epithelial cells also express nonclassical MHC class I-like molecules on their cell surface. CD1d is a non-polymorphic MHC-like molecule whose expression is mainly localized to the epithelial cells of the gastrointestinal tract. The biochemical structure of CD1d on intestinal epithelial cells (IECs) exists in two forms: a 37-kD nonglycosylated, beta 2-microglobulin (beta 2M) independent and a 48-kD glycosylated, beta 2M dependent form. Immunolocalization studies suggest that the 37-kD nonglycosylated form of CD1d is limited to the apical cell surface whereas the 48-50-kD glycosylated, beta 2M dependent form of CD1d is expressed both on the apical and the basolateral surfaces. The beta 2M association with CD1d seems to be important in regulating the pattern of glycosylation and the localization of CD1d within the cell based upon studies of the structure of CD1d in a transfected model cell line and in polarized epithelial cell monolayers. The functional role of intestinal CD1d remains unknown. However, based upon in vitro studies of the antigens presented by human CD1d and mouse CD1d, CD1d expressed on IECs likely presents a very hydrophobic glycolipid molecule possibly from the cell wall of bacteria or host cells. The processed-lipid antigen presented by CD1d may then involve a yet-to-be-identified subpopulation of the resident, oligoclonal alpha beta TCR CD8+ intestinal intraepithelial lymphocyte (iIEL) T cells. Subsequently, these T cells would be very important in regulating the local immune response by producing cytokines and recruiting other immune modulating cells to destroy infected cells, regenerate normal IECs, and possibly downregulate activated T cells to maintain mucosal integrity.

Inflammatory bowel disease is a chronic inflammatory condition of the gastrointestinal tract that manifests as ulcerative colitis and Crohn disease. Both these clinical entities result from interrelated genetic and environmental factors that may be channeled through an abnormality in mucosal immune function, possibly due to dysregulated or excessive T helper cell (T(H))1 (Crohn disease) or T(H)2 (ulcerative colitis) responses. This article reviews current knowledge of the role of immune factors in inflammatory bowel disease and the potential therapeutic strategies that target the pathways of T(H)1- or T(H)2-induced inflammation.

Animal models of inflammatory bowel disease (IBD) have been useful in the identification of those immune responses uniquely involved in IBD pathogenesis and in defining the important roles of environmental influences, such as normal luminal bacterial flora and the genetic composition of the host, in modifying IBD-associated inflammation. Recent studies have focused particular attention on CD4+ T cells which produce excessive quantities either of Th1 cytokines (IFN-gamma and TNF) directed by IL-12 or of a Th2 cytokine (IL-4), relative to the production of suppressive cytokines such as IL-10 and transforming growth factor beta. Such insights will be extremely beneficial in the development of novel approaches to the control of IBD-type inflammation, such as the use of anticytokine therapies and gene therapy, and finally, in the identification of the genetic abnormalities and the antigens driving the inflammation that underlies the human disease.

T cell activation, as defined by expression of relevant cell surface molecules, such as the interleukin-2 receptor (CD25), is increased in many chronic relapsing diseases, including inflammatory bowel disease (IBD). These T cells are generally activated through contact of their clonotypic T cell receptor (TCR) with a peptide antigen presented by a major histocompatibility complex molecule. One of the putative antigenic contact sites for the TCR is the third complementarity determining region (CDR3) of the TCR beta-chain variable region (TCRBV). Therefore, analysis of the TCRBV CDR3 provides insight into the diversity of antigens encountered by a given T cell population. This study evaluated the TCRBV CDR3 usage of the activated intestinal lymphocytes from human subjects with IBD, diverticulitis (inflammatory control), and a normal tissue control. Public patterns, as demonstrated by shared TCRBV CDR3 amino acid sequences of activated intestinal T cell subpopulations, were observed. In particular, a public pattern of TCRBV22, a conserved valine in the fifth position, and use of TCRBJ2S1 or TCRBJ2S5 was present in three of four Crohn's disease subjects while not present in the ulcerative colitis subjects. However, the private patterns of TCRBV CDR3 region amino acid sequences were far more striking and easily demonstrated in all individuals studied, including a normal noninflammatory control. Thus we conclude that selective antigenic pressures are prevalent among an individual's activated intestinal lymphocytes.

It is increasingly recognized that chronic Ag exposure may lead to clonal expansions of T cells, including those within the peripheral blood. Inflammatory bowel disease is a chronic, multisystemic disease of unknown origin that predominantly affects the intestine. We sought to determine whether clonal expansions of T cells are present in the peripheral blood of patients with inflammatory bowel disease by an examination of TCR usage. Positively selected CD4+ and CD8+ peripheral blood T cells were isolated from subjects with active ulcerative colitis, Crohn's disease, and diverticulitis and from normal controls. Analysis of complementarity determining region 3 lengths of 24 TCR-beta chain V region families from CD4+ and CD8+ peripheral blood T cells showed a skewed distribution in the three inflammatory groups, consistent with expansion of T cell clones, in comparison to the normally distributed pattern observed among the control donors. Random sequencing of the PCR amplification products of CD4+ peripheral blood T cells from the subjects with ulcerative colitis, Crohn's disease, and diverticulitis revealed reiterative TCR-beta chain sequences that were not found in the normal donors. In subjects with Crohn's disease, the reiterative TCR-beta chain sequences from the CD4+ peripheral blood T cells were persistent over at least a 1-yr period. The persistently expanded TCR-beta chain sequences of CD4+ peripheral blood T cells were identifiable in genomic DNA isolated from archival tissue of intestine from subjects with Crohn's disease and ulcerative colitis by Southern blotting and direct DNA sequencing. An identical twin pair, concordant for Crohn's disease, shared the same reiterative TCR-beta chain sequences in their CD4+ peripheral blood T cells. These studies show that chronic intestinal inflammation is associated with expansions of CD4+ peripheral blood T cells. Furthermore, in inflammatory bowel disease these T cell clonal expansions are persistent and shared among HLA-identical individuals, implicating a response to specific, persistent, and stimulating Ags in these diseases.