Yoh Ishiguro

Mucosal proinflammatory cytokine production correlates with endoscopic activity of ulcerative colitis

Abstract: Proinflammatory cytokines are believed to be involved in the pathogenesis of ulcerative colitis (UC). The aim of this study was to clarify the profiles of proinflammatory cytokine production in patients with UC in terms of disease intractability, endoscopic findings, and host response to lipopolysaccharide (LPS) stimulation. Colonic mucosal tissues were obtained from patients with active UC (n = 15, including 4 patients with intractable disease) and inactive UC (n = 7), non-inflammatory bowel disease (IBD) colitis (n = 11), and controls (n = 20). Organ culture was performed, and the amounts of four cytokines (described below) in the culture media were determined by enzyme-linked immunosorbent assay (ELISA). LPS stimulation enhanced interleukin (IL)-1β, IL-8, and IL-6 production in colonic specimens from all groups, but enhanced tumor necrosis factor (TNF)-α production only in active UC specimens. Levels of IL-6, IL-8, and TNF-α were significantly higher in active UC than in non-IBD colitis, and the production of all three of these cytokines was correlated to the endoscopic grade of inflammation. The production of these cytokines was also significantly higher in patients with intractable disease receiving corticosteroids than in patients with non-intractable disease receiving corticosteroids. These results suggest that enhanced production of mucosal proinflammatory cytokines may be implicated in the pathogenesis of UC.

Macrophage migration inhibitory factor contributes to the development of acute dextran sulphate sodium-induced colitis in Toll-like receptor 4 knockout mice

Toll‐like receptor 4 (TLR4), which recognizes lipopolysaccharides, plays an important role in the innate immune response. In this study, we investigated the role of TLR4 in the development of experimental colitis with regard to the biological actions of macrophage migration inhibitory factor (MIF) using TLR4 null (–/–) mice. TLR4–/– mice were given 2% dextran sulphate sodium (DSS) in drinking water to induce colitis, which was clinically and histologically as severe as that seen in wild‐type (WT) mice. The level of tumour necrosis factor (TNF)‐α in colon tissues was increased in WT mice but unchanged in TLR4–/– mice. The level of myeloperoxidase (MPO) activity in colon tissues was increased by DSS administration in both TLR4–/– and WT mice. The expression of MIF was up‐regulated in the colons of TLR4–/– mice with acute DSS‐induced colitis. An anti‐MIF antibody significantly suppressed colitis and elevation of matrix metalloproteinase‐13 in TLR4–/– mice. The current results obtained from TLR4–/– mice provide evidence that MIF plays a critical role in the development of acute DSS‐induced colitis.

Cyclosporine regulates intestinal epithelial apoptosis via TGF-β-related signaling

Cyclosporine is a potent immunomodulator and has a beneficial effect in the treatment of ulcerative colitis (UC). We analyzed the mechanism of the effects of cyclosporine on the regulation of epithelial apoptosis via TGF-beta-related signaling, because the balance between the apoptosis and regeneration of epithelial cells seems to be a key factor to maintain the intestinal homeostasis. For this purpose, colitis was induced by treatment of 4% dextran sulfate sodium (DSS), and the effect of treatment with cyclosporine and anti-TGF-beta antibody was assessed. Treatment with cyclosporine ameliorated body weight loss, mucosal destruction, and epithelial apoptosis in DSS-induced colitis. Cyclosporine was shown to upregulate the expression of TGF-beta in the colonic tissue, enhance the expression of p-Smad2 and cFLIP in epithelial cells, and inhibit caspase-8 activity but not caspase-1 or -9. Upregulation of cFLIP in the colonic epithelial cells, amelioration of body weight loss, and mucosal destruction by cyclosporine were attenuated by anti-TGF-beta antibody treatment. These results indicated that cyclosporine could have a protective role against epithelial apoptosis associated with upregulation of TGF-beta-related signaling.

Salmon cartilage proteoglycan suppresses mouse experimental colitis through induction of Foxp3+ regulatory T cells

Proteoglycans (PGs) are complex glycohydrates which are widely distributed in extracellular matrix (ECM). PGs are involved in the construction of ECM, cell proliferation and differentiation. ECM components are involved in transduction of proinflammatory responses, but it is still unknown whether PGs are involved in inflammatory response. In this study, we investigated the effect of PG extracted from salmon cartilage on the progression of experimental colitis-induced in severe combined immunodeficiency mice by cell transfer from interleukin-10 (IL-10)-/- mice. IL-10-/- cell-transferred mice showed weight loss, colon shortening and histological appearance of mild colitis. Daily oral administration of PG attenuated the clinical progression of colitis in a dose-dependent manner. Colitis-induced mice showed the elevated expression of IFN-γ, IL-12, TNF-α, IL-21, IL-23p19, IL-6, IL-17A and retinoic acid-related orphan receptor γt (RORγt) in lamina propria mononuclear cells (LPMCs) and oral administration of PG suppressed the expression of these factors. Conversely, expression of Foxp3 that induces CD4+CD25+ regulatory T cells in LPMCs was enhanced by PG administration. These findings suggested that salmon PG attenuated the progression of colitis due to suppression of inflammatory response by enhancement of regulatory T cell induction.

Retinoic acid-inducible gene-I is constitutively expressed and involved in IFN-γ-stimulated CXCL9-11 production in intestinal epithelial cells

Retinoic acid-inducible gene-I (RIG-I) is a member of the DExH/D family proteins, and plays an important role in antiviral response via interferon-stimulated genes (ISGs) and type 1 IFN. In this study, the roles of RIG-I in the epithelial cells in the cross-talk between type 2 IFN and inducible chemokines production are high-lighted. The results showed that RIG-I was constitutively expressed in normal surface epithelia lining the colonic mucosa. RIG-I was constitutively expressed in the epithelial cell lines HT-29, and IFN-gamma and TNF-alpha enhanced the RIG-I expression in a dose-dependent manner. IFN-gamma was shown to stimulate CXCL9-11 production, and RNA interference against RIG-I resulted in significant decrease of IFN-gamma-induced CXCL9-11 productions. These results suggest that RIG-I play an important role in the cross-talk between inflammatory cytokines and immune cell trafficking. In conclusion, RIG-I might regulate the gut barrier function in homeostatic and inflammatory conditions.

Multiple Accumulation of Vδ2+ γδ T-Cell Clonotypes in Intestinal Mucosa from Patients with Crohn's Disease

The γδT cells have been known to play an important role in the regulation of the mucosal immune system, but the relationship between these cells and the pathogenesis of Crohns disease (CD) has remained obscure. We now demonstrate the T-cell receptor (TCR) Vδ2 gene transcripts characterize antigenic immune response in the intestinal mucosa from patients with CD. TCR Vδ2 gene transcripts of six patients with CD and six controls were subcloned and 20 randomly selected clones from each sample were subjected to nucleotide sequencing. Sequence analysis demonstrated that the different clones in the intestinal mucosa of CD were significantly increased (11.833 ± 0.946) compared to controls (7.167 ± 1.470) (P = 0.0374). The presence of intraindividual dominance and oligoclonality of TCR Vδ2 gene transcripts in normal controls appears reflect positive selection and expansion of specific γδ T cells in normal controls. By contrast TCR Vδ2 gene transcripts in the intestinal mucosa of CD can express different clonotypes. We conclude that accumulation of multiple Vδ2+ γδ T-cell clonotypes are involved in the pathogenesis of CD.

The Presentation of Haptenated Proteins and Activation of T Cells in the Mesenteric Lymph Nodes by Dendritic Cells in the TNBS Colitis Rat

Abstract: We described the role of dendritic cells (DCs) in aspects of T cell activation at the mesenteric lymph nodes (MLN) in trinitrobenzene sulfonic acid (TNBS)‐induced colitis. An antigenic‐immune response is induced at the MLN, and dendritic cells are the affected cell type. Cross‐linking and GRO/CINC‐1 have synergistic effects for DC maturation.

Virulence factor p60 of Listeria monocytogenes modulates innate immunity by inducing tumor necrosis factor α

We investigated the effect of p60, a virulence factor of Listeria monocytogenes, on host immune response in vitro and in vivo. Administration of p60 before a sublethal infection with L. monocytogenes enhanced innate host resistance in naïve mice. Mouse macrophage RAW264.7 cells produced tumor necrosis factor (TNF)-alpha in response to stimulation with recombinant p60. Toll-like receptor 4 may be involved in TNF-alpha production from RAW264.7 cells and enhanced host resistance induced by p60 administration. Our findings demonstrated that p60 modulates innate immune responses against L. monocytogenes infection.

Transforming growth factor-β regulates susceptibility of epithelial apoptosis in murine model of colitis

Abstract: Transforming growth factor (TGF)‐beta has a key role in intestinal homeostasis. Our present data suggest that TGF‐beta, which was constitutively expressed by lamina propria mononuclear cells and epithelium, affected epithelial cells. Abnormal suppression of TGF‐beta could enhance the sensitivity of epithelial cells to apoptosis associated with interferon‐gamma in DSS‐induced colitis.

Macrophage Migration Inhibitory Factor and Activator Protein-1 in Ulcerative Colitis

Abstract: Macrophage migration inhibitory factor (MIF) is a cytokine that has potent antisteroid effects. We determined that MIF is involved in the glucocorticoid‐resistant inflammatory process of ulcerative colitis (UC), and the altered AP‐1 signal is a potent therapeutic target for refractory UC.