Motomi Yamazaki

Damaged epithelia regenerated by bone marrow–derived cells in the human gastrointestinal tract

Studies have shown that bone marrow cells have the potential to differentiate into a variety of cell types. Here we show that bone marrow cells can repopulate the epithelia of the human gastrointestinal tract. Epithelial cells of male donor origin were distributed in every part of the gastrointestinal tract of female bone marrow transplant recipients. Donor-derived epithelial cells substantially repopulated the gastrointestinal tract during epithelial regeneration after graft-versus-host disease or ulcer formation. Regeneration of gastrointestinal epithelia with donor-derived cells in humans shows a potential clinical application of bone marrow–derived cells for repairing severely damaged epithelia, not only in the gastrointestinal tract but also in other tissues.

Mucosal T Cells Expressing High Levels of IL-7 Receptor Are Potential Targets for Treatment of Chronic Colitis

The IL-7/IL-7R-dependent signaling pathway plays a crucial role in regulating the immune response in intestinal mucosa. Here we demonstrate the pivotal role of this pathway in the development and treatment of chronic colitis. T cells expressing high levels of IL-7R were substantially infiltrated in the chronic inflamed mucosa of TCR α-chain knockout mice and IL-7 transgenic mice. Transfer of mucosal T cells expressing high levels of IL-7R, but not T cells expressing low levels of IL-7R, from these mice into recombinase-activating gene-2−/− mice induced chronic colitis. Selective elimination of T cells expressing high levels of IL-7R by administrating small amounts of toxin-conjugated anti-IL-7R Ab completely ameliorated established, ongoing colitis. These findings provide evidence that therapeutic approaches targeting mucosal T cells expressing high levels of IL-7R are effective in the treatment of chronic intestinal inflammation and may be feasible for use in the therapy of human inflammatory bowel disease.

Functional expression of costimulatory molecule CD86 on epithelial cells in the inflamed colonic mucosa.

BACKGROUND & AIMS Costimulatory signals are essential for T-cell activation. The aim of this study was to demonstrate expression of costimulatory molecules CD86 and CD80 in human colonic epithelial cells and assess their functional roles in the activation of T cells in inflamed colonic mucosa. METHODS CD86 and CD80 expression was assessed by immunohistochemistry of colonic mucosa, reverse-transcription polymerase chain reaction, and flow cytometric analysis of isolated colonic epithelial cells and cell lines. Costimulatory effect of CD86-expressing colonic epithelial cells on purified CD4(+) T-cell proliferation was also assessed at suboptimal phytohemagglutinin stimulation. RESULTS CD86 and CD80 messenger RNA was detected in isolated colonic epithelial cells from normal and inflamed mucosa of patients with ulcerative colitis. Immunohistochemistry and flow cytometric analysis of colonic epithelial cells confirmed cell surface expression of CD86 protein in inflamed colonic mucosa. Cell surface expression of CD86 protein was increased in the colonic epithelial cell line HT29-18-N2 after interferon gamma stimulation. Purified CD4(+) T cells proliferated in response to suboptimal phytohemagglutinin costimulated with interferon gamma-stimulated HT29-18-N2, and these proliferative responses were efficiently inhibited by the addition of anti-CD86 monoclonal antibody. Costimulatory effect of CD86-expressing colonic epithelial cells isolated from inflamed mucosa of ulcerative colitis was also demonstrated at suboptimal phytohemagglutinin stimulation in CD4(+) T cells. CONCLUSIONS Colonic epithelial cells may act as antigen-presenting cells, and contribute to the activation of T cells through costimulatory molecule CD86 expression in inflamed colonic mucosa.

Interleukin-18 overproduction exacerbates the development of colitis with markedly infiltrated macrophages in interleukin-18 transgenic mice

Background and Aim:  The authors have previously shown that production of interleukin (IL)‐18 was increased in the inflamed mucosa of patients with Crohns disease (CD) and blockade of IL‐18 ameliorated the murine model of CD. This demonstrated that IL‐18 plays a significant role during intestinal inflammation. However, the initial role of IL‐18 during intestinal inflammation was unclear; therefore the susceptibility of IL‐18 transgenic (Tg) mice to acute dextran sulfate sodium (DSS)‐induced colitis was examined.

Interleukin-13 Damages Intestinal Mucosa via TWEAK and Fn14 in Mice—A Pathway Associated With Ulcerative Colitis

BACKGROUND & AIMS TWEAK, a member of the tumor necrosis factor (TNF) superfamily, promotes intestinal epithelial cell injury and signals through the receptor Fn14 following irradiation-induced tissue damage and during development of colitis in mice. Interleukin (IL)-13, an effector of tissue damage in similar models, has been associated with the pathogenesis of ulcerative colitis (UC). We investigated interactions between TWEAK and IL-13 following mucosal damage in mice. METHODS We compared patterns of gene expression in intestinal tissues from wild-type and TWEAK knockout mice following γ-irradiation. Intestinal explants from these mice were used to detect cell damage induced by IL-13 and TNF-α. Levels of messenger RNA for IL-13, TWEAK, and Fn14 were measured in mucosal samples from patients with UC. RESULTS Based on gene expression analysis, TWEAK mediates γ-irradiation-induced epithelial cell cycle arrest and apoptosis. However, TWEAK alone did not induce damage or apoptosis of primary intestinal epithelial cells. On the other hand, exogenous IL-13 activated caspase-3 in naïve intestinal explants; this process required TWEAK, Fn14, and secretion of endogenous TNF-α which was mediated by ADAM17. Conversely, activation of caspase by exogenous TNF-α required IL-13, TWEAK, and Fn14. In mucosa from patients with UC, messenger RNA levels of IL-13, TWEAK, and Fn14 increased with level of disease severity. CONCLUSIONS IL-13-induced damage of intestinal epithelial cells requires TWEAK, its receptor (Fn14), and TNF-α. IL-13, TNF-α, TWEAK, and Fn14 could perpetuate and aggravate intestinal inflammation in patients with UC.

A synthetic mimetic of CD4 is able to suppress disease in a rodent model of immune colitis

CD4+ mucosal T cells mediate the intestinal inflammation in Crohns disease and may serve as an important target for immune intervention. Here we assessed the therapeutic effect of a synthetic mimetic of CD4 designed to mimic both the sequence and conformation of the complementarity‐determining region 3 of murine CD4 V1 domain (rD‐mPGPtide) in a mouse colitis model using immunization with 2,4,6‐trinitrobenzene sulfonic acid (TNB). i.  v. administration of the rD‐mPGPtide but not control scrambled peptide could suppress severe inflammation in the chronic colitis mouse model. After treatment with the rD‐mPGPtide, a striking improvement of diarrhea and acute wasting disease was observed with decreased mortality. Serum anti‐TNB antibody titers, CD45RBlowCD4+ T cells in the lamina propria and IFN‐γ mRNA expression in the mucosa were significantly decreased with the rD‐mPGPtide treatment. Anti‐CD4 antibody also suppressed disease by depletion of CD45RBhighCD4+ T cells in the colonic mucosa. The observation that the synthetically engineered analogue of murine CD4 inhibits inflammation in a rodent disease model by different mechanisms than anti‐CD4 antibody suggests that a human version of this peptide has potential therapeutic utility in CD4+ mucosal T cell‐mediated intestinal inflammation in Crohns disease.

Microbiota-derived lactate accelerates colon epithelial cell turnover in starvation-refed mice

Oral food intake influences the morphology and function of intestinal epithelial cells and maintains gastrointestinal cell turnover. However, how exactly these processes are regulated, particularly in the large intestine, remains unclear. Here we identify microbiota-derived lactate as a major factor inducing enterocyte hyperproliferation in starvation-refed mice. Using bromodeoxyuridine staining, we show that colonic epithelial cell turnover arrests during a 12- to 36-h period of starvation and increases 12-24 h after refeeding. Enhanced epithelial cell proliferation depends on the increase in live Lactobacillus murinus, lactate production and dietary fibre content. In the model of colon tumorigenesis, mice exposed to a carcinogen during refeeding develop more aberrant crypt foci than mice fed ad libitum. Furthermore, starvation after carcinogen exposure greatly reduced the incidence of aberrant crypt foci. Our results indicate that the content of food used for refeeding as well as the timing of carcinogen exposure influence the incidence of colon tumorigenesis in mice.

Human intestinal epithelial cell-derived interleukin (IL)-18, along with IL-2, IL-7 and IL-15, is a potent synergistic factor for the proliferation of intraepithelial lymphocytes

Intestinal epithelial cell (IEC)‐derived cytokines, such as stem cell factor (SCF), interleukin (IL)‐7 and IL‐15 are known to be required for the development of intestinal intraepithelial lymphocytes (IELs). A newly described cytokine, IL‐18, has also been shown to be produced by intestinal epithelial cells. To demonstrate the functional effects of IL‐18 on human IELs, we assessed IL‐18/IL‐18 receptor expression in IEC/IEL and proliferation following stimulation of intestinal IELs by IL‐18. IL‐18 transcripts were detected both in freshly isolated human colonic epithelial cells and in various colonic epithelial cell lines. IL‐18 protein was also detected by ELISA and flow cytometric analysis using antihuman IL‐18‐specific monoclonal antibody (MoAb). Furthermore, IELs constitutively expressed the IL‐18 receptor in addition to the IL‐2 and IL‐7 receptors. More importantly, IL‐18 augmented significant proliferative responses of IEL in combination with IL‐2, IL‐7 and IL‐15 both in the presence and in absence of anti‐CD3 MoAb. These results suggest that IL‐18 might play a crucial role in the proliferation and maintenance of intestinal IELs.

Th1-mediated intestinal inflammation in Crohn’s disease may be induced by activation of lamina propria lymphocytes through synergistic stimulation of interleukin-12 and interleukin-18 without T cell receptor engagement

Th1-mediated intestinal inflammation in Crohns disease may be induced by activation of lamina propria lymphocytes through synergistic stimulation of interleukin-12 and interleukin-18 without T cell receptor engagement

Regulatory effect of interleukin-4 and interleukin-13 on colon cancer cell adhesion

To assess the role of interleukin-4 (IL-4) and interleukin-13 (IL-13) in colon cancer cell–cell adhesion, we investigated the effect of both cytokines in human colon cancer cell line, colo205 cell–cell adhesion. IL-4 receptor was expressed on the cell surface of colo205, and recombinant IL-4 inhibited colo205 cell–cell adhesion in a dose-dependent fashion without inhibiting cell proliferation. Flow cytometric analysis revealed that monoclonal antibodies (mAbs) directed against E-cadherin and carcinoembryonic antigen (CEA) inhibited colo205 cell–cell adhesion and IL-4 significantly inhibited the expression of E-cadherin and CEA. IL-13 also inhibited colo205 cell–cell adhesion. These results indicated that IL-4 and IL-13 inhibited colon cancer cell–cell adhesion by down-regulation of E-cadherin and CEA molecules. We then investigated the expression of both cytokines from freshly isolated colon cancer tumour-infiltrating lymphocytes (TILs). With reverse transcription-polymerase chain reaction and flow cytometric analysis, we demonstrated that colon TILs expressed IL-4 and IL-13 mRNA and protein. These results suggest that Th 2 type cytokines IL-4 and IL-13 locally-produced from TILs may regulate colon cancer adhesion by down-regulation of adhesion molecules.