concanavalin-a and Colitis

The gut-liver axis is of clinical importance as a potential therapeutic target in a wide range of liver diseases; however, the mechanisms underlying interactions between microbial products and immune responses in the liver remain unknown. In this study, we demonstrated that IL-10-producing macrophages contribute to immune tolerance in the inflamed liver under intestinal barrier disruption in a murine tandem model of dextran sulfate sodium (DSS) colitis and concanavalin A (Con A) hepatitis. Intestinal barrier disruption protected mice from subsequent liver injury, and the severity of colitis directly affected susceptibility to such injury. The protective effect of DSS-Con A was canceled in gut-sterilized mice, suggesting that gut microbiota play a substantial role in this process. Altered gut microbiota and their metabolites, along with a disrupted intestinal barrier, directly gave rise to immunological permissiveness in the inflamed liver. We identified 1-methylnicotinamide (1-MNA) as a candidate metabolite capable of suppressing liver injury with the potential to induce IL-10-producing macrophages. Consistently, expression of nicotinamide N-methyltransferase, which converts nicotinamide to 1-MNA, was upregulated in the liver of DSS-Con A mice, and this effect was abrogated by gut sterilization. Collectively, our results provide a mechanistic insight into the regulation of immunological balance in the liver via the gut-liver axis.

Topics: Animals; Chemical and Drug Induced Liver Injury; Colitis; Concanavalin A; Dextran Sulfate; Disease Models, Animal; Female; Gastrointestinal Microbiome; Hepatitis; Interleukin-10; Liver; Macrophages; Male; Mice; Mice, Inbred C57BL; Niacinamide; T-Lymphocytes

Liver resident macrophages designated Kupffer cells (KCs) form the largest subpopulation of tissue macrophages. KCs are involved in the pathogenesis of liver inflammation. However, the role of KCs in the systemic inflammation is still elusive. In this study, we examined whether KCs are involved in not only intrahepatic inflammation but also extrahepatic systemic inflammation. Administration of clodronate liposomes resulted in the KC deletion and in the suppression of liver injury in T cell-mediated hepatitis by ConA as a local acute inflammation model, while the treatment did not influence dextran sulfate sodium- (DSS-) induced colitis featured by weight loss, intestinal shrink, and pathological observation as an ectopic local acute inflammation model. In contrast, KC deletion inhibited collagen-induced arthritis as a model of extrahepatic, systemic chronical inflammation. KC deleted mice showed weaker arthritic scores, less joint swelling, and more joint space compared to arthritis-induced control mice. These results strongly suggest that KCs are involved in not only intrahepatic inflammatory response but also systemic (especially) chronic inflammation.

Topics: Acute Disease; Animals; Arthritis, Experimental; Chronic Disease; Colitis; Collagen; Concanavalin A; Dextran Sulfate; Disease Models, Animal; Female; Inflammation; Kupffer Cells; Liver; Mice, Inbred C57BL; Mice, Inbred DBA; Mice, Inbred ICR

Leptin-deficient ob/ob mice are resistant to dextran sulfate sodium (DSS)-induced colitis and Concanavalin A (Con A)-induced hepatitis. However, the signal transduction pathways involved have not been identified. The present study investigated the effect of leptin-induced STAT3 signaling in the DSS and Con A models. Mice carrying a leptin receptor (LEPR) gene mutant for Y1138 (s/s mice), with abrogated leptin-induced STAT3 signaling, were compared with wild-type (WT) and LEPR-deficient db/db mice. Administration of DSS to s/s mice resulted in a clinical score and colon shortening of intermediate severity compared with disease induced in WT and db/db mice-the latter group having the lowest disease severity. A comparable degree of inflammatory infiltrate and epithelial damage was observed in the colon of WT and s/s mice, and these parameters were reduced in db/db mice. Levels of IFN-gamma, IL-6, IL-10, and TNF-alpha were comparable in the colon of s/s and db/db mice, and a similar trend was observed for CXCL2. s/s and WT mice developed severe liver disease in response to Con A, whereas db/db mice were protected. However, Con A-induced serum IL-6 and TNF-alpha levels in s/s mice mimicked levels observed in db/db rather than WT mice. In conclusion, lack of leptin-induced STAT3 signaling is associated with reduced cytokine production following DSS and Con A administration, but it appears to sensitize mice to the effects of proinflammatory mediators.

Topics: Animals; Chemical and Drug Induced Liver Injury; Colitis; Concanavalin A; Cytokines; Dextran Sulfate; Inflammation; Mice; Receptors, Leptin; Signal Transduction; STAT3 Transcription Factor

Inflammatory bowel disease (IBD) is a multifactorial disorder with an unknown aetiology. The aim of this study is to employ a murine model of IBD to identify pathways and genes, which may play a key role in the pathogenesis of IBD and could be important for discovery of new disease markers in human disease. Here, we have investigated severe combined immunodeficient (SCID) mice, which upon adoptive transfer with concanavalin A-activated CD4(+) T cells develop inflammation of the colon with predominance in rectum. Mice with increasing level of inflammation was studied. RNA from rectum of transplanted and non-transplanted SCID mice was investigated by a genome-wide gene expression analysis using the Affymetrix mouse expression array 430A (MOE430A) including 22,626 probe sets. A significant change in gene expression (P = 0.00001) is observed in 152 of the genes between the non-transplanted control mice and colitis mice, and among these genes there is an overrepresentation of genes involved in inflammatory processes. Some of the most significant genes showing higher expression encode S100A proteins and chemokines involved in trafficking of leucocytes in inflammatory areas. Classification by gene clustering based on the genes with the significantly altered gene expression corresponds to two different levels of inflammation as established by the histological scoring of the inflamed rectum. These data demonstrate that this SCID T-cell transfer model is a useful animal model for human IBD and can be used for suggesting candidate genes involved in the pathogenesis and for identifying new molecular markers of chronic inflammation in human IBD.

Topics: Adoptive Transfer; Animals; Chromosome Mapping; Cluster Analysis; Colitis; Colon; Concanavalin A; Female; Gene Expression Profiling; Genome; Genome, Human; Humans; Inflammatory Bowel Diseases; Mice; Mice, Inbred BALB C; Mice, SCID; Oligonucleotide Array Sequence Analysis; Reverse Transcriptase Polymerase Chain Reaction; Synteny; T-Lymphocytes

Natural killer T-cell (NKT) regulatory lymphocytes have been shown to behave differently in various immune settings. The aim of the present study was to determine the effect of microenvironmental signaling on NKT polarization and the process of active CD8 and NKT intrahepatic lymphocyte sequestration. In an in vitro assay, double negative (DN) NKT hybridoma cells were incubated with Hep3B hepatoma cells. This caused a significant increase in the secretion of alpha-fetoprotein (AFP) from Hep3B cells. When NKT cells were exposed to beta-glucoslyceramide (beta-GC) prior to incubation, Hep3B cells exhibited increased proliferation, increased IFN secretion, and reduced AFP secretion. In vivo, the adoptive transfer of naïve DN NKT cells into athymic nude-nu mice transplanted with human Hep3B hepatocellular carcinoma (HCC) caused accelerated tumor growth. This effect was inhibited by prior ex vivo exposure of DN NKT lymphocytes to beta-GC. To assess the effect of the immunological environment on NKT cells, immune mediated hepatitis and colitis were induced simultaneously in mice. Induction of TNBS colitis prior to administration of concanavalin A (Con A) hepatitis resulted in an aggravation of the liver damage caused by Con A hepatitis alone. This effect was associated with reduced intrahepatic CD8+ T cell trapping and an increase in intrahepatic NKT cells. The presence of different ligands altered host microenvironment signaling and influenced the fate and polarization of NKT cells and the sequestration of active intrahepatic lymphocytes. These data support the notion that NKT regulatory lymphocytes have an inherent plasticity that may be important for their regulatory function.

Topics: Animals; Benzoates; CD8-Positive T-Lymphocytes; Cell Line; Cell Line, Tumor; Cell Proliferation; Chemical and Drug Induced Liver Injury; Colitis; Concanavalin A; Cytokines; Disease Models, Animal; Glycosphingolipids; Humans; Immunity; Killer Cells, Natural; Ligands; Liver Neoplasms, Experimental; Lymphocyte Subsets; Male; Mice; Mice, Nude; Signal Transduction; Xenograft Model Antitumor Assays

Curcumin (diferulolylmethane) has been shown to have a protective role in mouse models of inflammatory bowel diseases (IBD) and to reduce the relapse rate in human ulcerative colitis (UC), thus making it a potentially viable supportive treatment option. Trinitrobenzene sulfonic acid (TNBS) colitis in NKT-deficient SJL/J mice has been described as Th1-mediated inflammation, whereas BALB/c mice are believed to exhibit a mixed Th1/Th2 response.. We therefore investigated the effect of dietary curcumin in colitis induced in these 2 strains.. In the BALB/c mice, curcumin significantly increased survival, prevented weight loss, and normalized disease activity. In the SJL/J mice, curcumin demonstrated no protective effects. Genomewide microarray analysis of colonic gene expression was employed to define the differential effect of curcumin in these 2 strains. This analysis not only confirmed the disparate responses of the 2 strains to curcumin but also indicated different responses to TNBS. Curcumin inhibited proliferation of splenocytes from naive BALB/c mice but not SJL/J mice when nonspecifically stimulated in vitro with concanavalin A (ConA). Proliferation of CD4(+) splenocytes was inhibited in both strains, albeit with about a 2-fold higher IC(50) in SJL/J mice. Secretion of IL-4 and IL-5 by CD4(+) lymphocytes of BALB/c mice but not SJL/J mice was significantly augmented by ConA and reduced to control levels by curcumin.. The efficacy of dietary curcumin in TNBS colitis varies in BALB/c and SJL/J mouse strains. Although the exact mechanism underlying these differences is unclear, the results suggest that the therapeutic value of dietary curcumin may differ depending on the nature of immune dysregulation in IBD.

Topics: Animals; Colitis; Concanavalin A; Curcumin; Diet; Disease Models, Animal; Inflammatory Bowel Diseases; Lymphocytes; Male; Mice; Mice, Inbred BALB C; Mice, Inbred Strains; Microarray Analysis; Reverse Transcriptase Polymerase Chain Reaction; Species Specificity; Trinitrobenzenesulfonic Acid

Virus-mediated delivery of therapeutic transgenes to the inflamed colon holds a great potential to serve as an effective therapeutic strategy for inflammatory bowel disease, since local, long-term expression of the encoded therapeutic proteins in the colorectal system is potentially achievable. Viral vectors, derived from adeno-associated virus (AAV), should be very useful for such therapeutic strategies, particularly because they can establish long-term expression of transgenes. However, few studies have been carried out to investigate the ability of AAV-based vectors to transduce the inflamed colon.. AAV, derived from adeno-associated virus serotype 2 (AAV2), showed a limited ability to transduce colonic cell lines in vitro when used in free form. No appreciable enhancement of the transduction efficiency was seen when AAV2 particles were attached stably to the surfaces of microbeads and delivered to target cells in the form of AAV2-microbead conjugates. However, the transduction efficiency of these colonic cell lines was enhanced substantially when a lectin, concanavalin A (Con A), was co-attached to the microbead surfaces, to which AAV2 particles had been conjugated. This considerable infectivity enhancement of AAV2-microbead conjugates by the co-attachment of Con A may be derived from the fact that Con A binds to alpha-D-mannosyl moieties that are commonly and abundantly present in cell-surface carbohydrate chains, allowing the conjugates to associate stably with target cells. Intracolonical administration of free AAV2 or AAV2-microbead conjugates without Con A into a mouse colitis model by enema showed very poor transduction of the colonic tissue. In contrast, the delivery of AAV2 in the form of AAV2-microbead conjugates bearing Con A resulted in efficient transduction of the inflamed colon.. AAV2-microbead conjugates bearing Con A can serve as efficient gene transfer agents both for poorly permissive colonic cell lines in vitro and for the inflamed colon in a mouse colitis model. This efficient transduction system for the inflamed colon should be useful for the development of gene therapy strategies for inflammatory bowel disease.

Topics: Animals; Cell Line; Colitis; Colon; Concanavalin A; Dependovirus; Drug Carriers; Genetic Therapy; Genetic Vectors; Mice; Microspheres; Signal Transduction; Transfection

The role of leptin in the immune system has been well established. While adipocytes represent the major source, leptin production by lymphocytes, infiltrating at the site of inflammation, was recently demonstrated. However, the significance of this locally released leptin remains unresolved. In the present study, two models in which absence of leptin-signalling is associated with protection were employed: the model of ConA-induced hepatitis and the CD4(+)CD45Rb(high) transfer model of colitis. For the ConA model, scid mice were reconstituted with either WT or leptin-deficient (ob/ob) CD4(+) T cells. Eight weeks post transfer, ConA was injected and serum ALT, TNFalpha, leptin as well as liver mononuclear cell activation and histological signs of inflammation were evaluated. No difference between recipients of WT or ob/ob cells was observed for any of the parameters evaluated. In the second model, either WT or ob/ob CD4(+)CD45Rb(high) cells were transferred into scid mice. No histological differences were detected, although recipients of ob/ob cells showed higher weight loss compared to recipients of WT cells. Spontaneous production of IL-6 from colon cultures obtained from recipients of ob/ob cells was reduced compared to recipients of WT cells, whereas stimulation of lamina propria lymphocytes with leptin resulted in a higher IFNgamma release in recipients of ob/ob cells compared to recipients of WT cells. In conclusion, the present study provides evidence that T cell-derived leptin does not play a major role in the regulation of the inflammatory process, indicating that the adipose tissue is the critical player in the immune-modulating effects of leptin.

Topics: Animals; Apoptosis; CD4-Positive T-Lymphocytes; Colitis; Colon; Concanavalin A; Hepatitis; Intestinal Mucosa; Leptin; Leukocyte Common Antigens; Leukocytes, Mononuclear; Liver; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; Mice, SCID

OX40 is a member of the TNFR superfamily, and is found predominantly on activated CD4-positive T cells. In vitro an OX40-IgG fusion protein inhibits mitogen- and Ag-driven proliferation and cytokine release by splenocytes and lymph node T cells. In contrast, an OX40 ligand-IgG fusion protein enhanced proliferative responses. In normal mice, OX40-positive cells are observed only in lymphoid tissues, including Peyer's patches of the gut. In mice with hapten-induced colitis or IL-2 knockout mice with spontaneous colitis, OX40-positive cells are found infiltrating the lamina propria. Administration of the OX40-IgG fusion protein to mice with ongoing colitis (but not the OX40 ligand-IgG) ameliorated disease in both mouse models of inflammatory bowel disease. This was evidenced by a reduction in tissue myeloperoxidase; reduced transcripts for TNF-alpha, IL-1, IL-12, and IFN-gamma; and a reduction in the T cell infiltrate. Targeting OX40 therefore shows considerable promise as a new strategy to inhibit ongoing T cell reactions in the gut.

Topics: Animals; CHO Cells; Colitis; Concanavalin A; Cricetinae; Cytokines; Epitopes, T-Lymphocyte; Female; Humans; Immunoglobulin G; Inflammatory Bowel Diseases; Injections, Intraperitoneal; Interleukin-2; Ligands; Lymphocyte Activation; Membrane Glycoproteins; Mice; Mice, Inbred BALB C; Mice, Inbred C3H; Mice, Knockout; OX40 Ligand; Receptors, OX40; Receptors, Tumor Necrosis Factor; Recombinant Fusion Proteins; T-Lymphocyte Subsets; Tumor Necrosis Factor Receptor Superfamily, Member 7; Tumor Necrosis Factors

Altered interleukin 2 (IL-2) production has been implicated in the pathogenesis of inflammatory bowel diseases.. The temporal relationship between IL-2, prostaglandin E2 (PGE2) production, and mucosal injury was evaluated by isolated colonic lamina propria mononuclear cells (LPMC), using the trinitrobenzene sulfonic acid model of rat colitis.. Spontaneous LPMC IL-2 activity was significantly increased in chronic (5 weeks) but not acute (5 days) or resolved colitis groups. IL-2 activity after concanavalin A activation was highest in the groups with resolved and chronic colitis. PGE2 production was significantly increased in LPMC cultures in acute or chronic colitis as well as the ethanol control groups but not the resolved colitis group. The addition of indomethacin to LPMC cultures decreased PGE2 levels in all groups, whereas IL-2 activity increased only for the chronic and resolved colitis groups. No correlation was found between PGE2 and IL-2 production by LPMC.. In this experimental model, LPMC IL-2 production varied according to the severity and duration of the inflammation. Increased PGE2 production does not appear to be responsible for the IL-2 alterations in colitis.

Topics: Acute Disease; Animals; Chronic Disease; Colitis; Concanavalin A; Dinoprostone; Disease Models, Animal; Female; Inflammation; Interleukin-2; Intestinal Mucosa; Lymphocyte Activation; Lymphocytes; Organ Size; Rats; Rats, Sprague-Dawley; Reference Values; Spleen; Trinitrobenzenesulfonic Acid

We have examined the T-cell functional capabilities of human intestinal mononuclear cells isolated from surgically obtained normal and inflammatory bowel disease intestinal specimens. Intestinal mononuclear cells have T cells present which respond to the mitogenic lectins E-PHA, Con A, and PWM and to foreign cell-surface antigens in the allogeneic-mixed leukocyte reaction. Intestinal mononuclear cells from ulcerative colitis patients exhibit a significantly decreased responsiveness in comparison to normal intestinal mononuclear cells with regard to both mitogenesis and the allogeneic-mixed leukocyte reaction; however, these defects may be secondary to the severity of the disease process that led to intestinal resection or the therapy which patients had received. Although both normal and inflammatory bowel disease intestinal mononuclear cells exhibit responsiveness in the allogeneic-mixed leukocyte reaction, thus indicating recognition of and sensitization by foreign cell-surface determinants, intestinal mononuclear cells do not subsequently kill the sensitizing cells in cell-mediated lympholysis. Therefore, the subclass of T cells which mediates cell-mediated lympholysis may either be absent from intestinal mononuclear cells or nonfunctional, while the subclass of T cells which responds in the allogeneic-mixed leukocyte reaction is both present and functional. This observation adds to the evidence of major functional differences between intestinal and peripheral blood mononuclear cells. Therefore, it will be necessary to better understand the factors regulating the effector capabilities of intestinal mononuclear cells before delineation of immunopathologic events in these tissues.

Topics: Adult; Colitis; Concanavalin A; Cytotoxicity, Immunologic; Female; Humans; Intestinal Mucosa; Lymphocyte Activation; Lymphocyte Culture Test, Mixed; Phytohemagglutinins; Prednisone; Rosette Formation; T-Lymphocytes

Dinitrochlorobenzene-induced colitis in guinea-pigs may be immunologically mediated: animals must be presensitised to dinitrochlorobenzene to develop colitis, sensitivity can be passively transferred by lymphocytes and the injury can be mitigated by immunosuppression. In this study, we examined lamina propria lymphocytes isolated from colons of animals with dinitrochlorobenzene-induced colitis, and appropriate controls. Lamina propria lymphocytes from colitis animals have a greater percentage of rabbit erythrocyte-rosetting cells (T cells) (20.1 +/- 3.0 vs 2.3 +/- 0.8, p less than .01) and a greater capacity to mediate mitogen-induced cellular cytotoxicity with phytohaemagglutinin than lamina propria lymphocytes from normal colon (% specific cytoxicity = 29.4 +/- 8.7 vs 5.0 +/- 4.5, P less than .005). There was no difference in the percentage of rosetting cells or cytotoxicity index of spleen or mesenteric lymph node lymphocytes between the colitis animals and controls. These data suggest that there are changes in the distribution and functional characteristics of lamina propria lymphocytes which correlate with mucosal cell injury in the dinitrochlorobenzene-colitis model.

Topics: Animals; Colitis; Colon; Concanavalin A; Cytotoxicity, Immunologic; Dinitrochlorobenzene; Guinea Pigs; Leukocyte Count; Lymph Nodes; Lymphocytes; Male; Mesentery; Phytohemagglutinins; Rosette Formation; Spleen; T-Lymphocytes