Reduced susceptibility to dextran sulphate sodium-induced colitis in the interleukin-2 heterozygous (IL-2) mouse

Summary Mice homozygous for an inactivation of the interleukin-2 (IL-2) gene develop a T-cell dependent colitis. Heterozygous (IL-2+/-) mice are clinically healthy but have been shown to express reduced levels of IL-2 in the colon. Splenocytes from the IL-2+/- mice had a poorer proliferative response to polyclonal T-cell activation and these mice have reduced numbers of intestinal regulatory T cells (CD4+ CD25+ cells) when compared to wild type mice. When exposed to dextran sulphate sodium (DSS) IL-2+/- mice showed a markedly reduced susceptibility to DSS-induced colitis. While DSS treatment caused a marked increase in both CD4+ and CD8+ colonic T cells expressing increased levels of IL-2, IL-4, and IL-10 in wild type mice none of these changes were seen in IL-2+/- mice. On the contrary, cytokine expression in intestinal T cells of IL-2+/- mice was actually reduced after DSS treatment. These results suggest that reduced levels of IL-2 leads to attenuated activation and function of intestinal T cells in IL-2+/- mice and a failure to react adequately to DSS exposure.     

Blocking lymphotoxin-beta receptor activation diminishes inflammation via reduced mucosal addressin cell adhesion molecule-1 (MAdCAM-1) expression and leucocyte margination in chronic DSS-induced colitis

The lymphotoxin-beta receptor (LTbetaR) pathway is critical for maintenance of organized lymphoid structures and is involved in the development of colitis. To investigate the mechanisms by which LTbetaR activation contributes to the pathology of chronic inflammation we used a soluble LTbetaR-Ig fusion protein as a competitive inhibitor of LTbetaR activation in the mouse model of chronic colitis induced by oral administration of dextran sulphate sodium. Strong expression of LTbeta which constitutes part of the LTalpha(1)beta(2) ligand complex was detected in colonic tissue of mice with chronic colitis. Treatment with LTbetaR-Ig significantly attenuated the development and histological manifestations of the chronic inflammation and reduced the production of inflammatory cytokines such as TNF, IL-1beta, and IL-6. Moreover, LTbetaR-Ig treatment significantly down-regulated mucosal addressin cell adhesion molecule-1 (MAdCAM-1) expression, leading to reduced leucocyte rolling and sticking in postcapillary and collecting venules and reduced extravasation into the intestinal mucosa as quantified by in vivo fluorescence microscopy. Thus, LTbetaR pathway inhibition ameliorates DSS-induced experimental chronic colitis in mice by MAdCAM-1 down-regulation entailing reduced lymphocyte margination and extravasation into the inflamed mucosa. Therefore, a combined treatment with reagents blocking T cell-mediated perpetuation of chronic inflammation such as LTbetaR-Ig together with direct anti-inflammatory reagents such as TNF inhibitors could constitute a promising treatment strategy for chronic colitis.     

Altered expression of cell adhesion molecules in uninvolved gut in inflammatory bowel disease.

Adhesion of circulating cells to vascular endothelium occurs in the early phase of inflammation, and is mediated by specific cell adhesion molecules. Many such adhesion molecules are increased in inflamed regions of ulcerative colitis (UC) and Crohn's disease (CD) but there is limited knowledge of their expression in the uninvolved gut, adjacent to inflammation. We investigated immunohistochemically the expression of platelet endothelial cell adhesion molecule-1 (PECAM-1), intercellular adhesion molecule-1 (ICAM-1) and lymphocyte function-associated antigen-1 (LFA-1) on resected specimens taken at a distance of 2-4 cm from the inflamed area and without histological signs of inflammation. Compared with normal gut, we found (i) a significant increase of PECAM-1-positive vessels in the mucosa of uninvolved UC (149.0 +/- 24.1 vessels/mm2 (mean +/- s.d.); normal colon = 123.1 +/- 21.6; P = 0.004); (ii) a significant decrease of ICAM-1-positive vessels in uninvolved CD (111.9 +/- 22.6 vessels/mm2; normal ileum = 136.9 +/- 27.6; P = 0.04); and (iii) a moderate but statistically insignificant increase of LFA-1-positive cells in the mucosa of uninvolved UC and Crohn's ileitis. This altered expression of cell adhesion molecules may contribute to the early lesion in inflammatory bowel disease and provide new therapeutic opportunities.     

Alteration in expression of beta 2 integrins on lamina propria lymphocytes in ulcerative colitis and Crohn's disease

We have previously demonstrated by immunohistochemistry that mucosal expression of beta 2 integrins was enhanced in Crohn's disease and ulcerative colitis as compared to normal controls. We aimed, therefore, to determine whether there was a corresponding alteration in the expression of CD11a/CD18 (LFA-1), the primary lymphocyte beta 2 integrin, among the principal subsets of lamina propria lymphocytes (LPLs). Accordingly, LPLs were extracted from surgical resection specimens derived from patients with Crohn's colitis, ulcerative colitis, and from noninflamed controls. Following immunofluorescent staining, three-color flow-cytometry analysis identified LPLs on the basis of CD45 side scatter gating, which in turn, were further subdivided into CD4(+), CD8(+), and CD19(+) cells to account for the predominant T and B cells in the lamina propria. Expression patterns of CD11a, the alpha-subunit of LFA-1; CD18, the beta-subunit of LFA-1; and alpha d, a novel alpha-subunit of the beta 2 integrin family were assessed for each of these lymphocyte subsets. In Crohn's disease and ulcerative colitis there was an increased mean percentage expression of CD4(+) cells and CD11a(+) cells compared with noninflamed controls. CD11a was more likely to be expressed on CD4(+) cells in both Crohn's disease and ulcerative colitis and compared with controls and less expressed on CD19(+) cells. It is likely that an influx of CD4(+)11a(+) cells into the lamina propria accounted for these changes. These results suggest that although currently there is great interest in harnessing alpha 4 beta 7 in treatment of inflammatory bowel disease, further consideration should be given to the role of CD11a in these disease states.     

Colonic histopathology in untreated celiac sprue or refractory sprue: Is it lymphocytic colitis or colonic lymphocytosis?

Colonic histopathology in some patients with untreated celiac sprue and refractory sprue has been said to be indistinguishable from lymphocytic colitis, but there have been no objective comparisons on which this is based. The purpose of this study was to determine the prevalence and to characterize the nature of colonic histopathology at the time of diagnosis in patients with celiac or refractory sprue. Colonoscopic biopsy specimens obtained at the time of diagnosis from 16 patients with celiac sprue, six patients with refractory sprue, nine patients with lymphocytic colitis, and five normal controls were analyzed blindly by histological and morphometric methods, quantitating the number and specific subtypes of inflammatory cells within the lamina propria and epithelium. Immunoperoxidase staining of intraepithelial lymphocytes with a monoclonal antibody to CD8 also was performed. Three of 16 patients with untreated celiac sprue (19%) were thought to have colonic histological abnormalities, which by morphometry consisted of slightly increased numbers of lymphocytes in the surface epithelium and lamina propria, many of which were CD8-positive. These abnormalities were distinguishable from lymphocytic colitis by the lack of increased overall lamina propria cellularity and surface epithelial abnormalities, and by fewer intraepithelial lymphocytes. In refractory sprue, colonic histological abnormalities were more frequent than in celiac sprue, occurring in four of six patients (67%), more pronounced, and identical to those in the lymphocytic colitis syndrome. However, colonic intraepithelial lymphocytes in lymphocytic colitis were mostly CD8-positive, whereas those in the colitis of refractory sprue rarely were. Mild colonic lymphocytosis in patients with untreated celiac sprue should be distinguished from lymphocytic colitis by the lack of surface epithelial abnormalities, the lack of increased cellularity of the lamina propria, and the lack of ongoing watery diarrhea after treatment with a gluten-free diet. In contrast, colonic histopathology in refractory sprue is indistinguishable from lymphocytic colitis, although immunohistochemical differences do exist.     

Phenotypically distinct subsets of CD4+ T cells induce or protect from chronic intestinal inflammation in C. B-17 scid mice

CD4⁺ T cells in the mouse can be subdivided into two fractionsbased on the level of expression of the CD45RB determinant.Previous studies have shown that these subsets are functionallydistinct. We have further characterized the properties of thesesubpopulations in vivo by injecting them into C. B-17 scid mice.The animals restored with the CD45RB^highCD4⁺ T cell populationdeveloped a lethal wasting disease with severe mononuclear cellinfiltrates into the colon and elevated levels of IFN- mRNA.In contrast, animals restored with the reciprocal CD45RB^lowsubset or with unfractionated CD4⁺ T cells did not develop thewasting or colitis. Importantly, the co-transfer of the CD45RB^lowpopulation with the CD45RB^high population prevented the wastingdisease and colitis. These data indicate that important regulatoryinteractions occur between the CD45RB^high and CD45RB^lowCD4⁺T cell subsets and that disruption of this mechanism has fatalconsequences.     

Imbalanced secondary mucosal antioxidant response in inflammatory bowel disease

Intestinal mucosal damage in the inflammatory bowel diseases (IBD) Crohn's disease (CD) and ulcerative colitis (UC) involves reactive oxygen metabolites (ROMs). ROMs are neutralized by endogenous antioxidant enzymes in a carefully balanced two-step pathway. Superoxide dismutases (SODs) convert superoxide anion to hydrogen peroxide (H(2)O(2)), which is subsequently neutralized to water by catalase (CAT) or glutathione peroxidase (GPO). Remarkably changed expression levels of the three isoforms of SOD in paired non-inflamed and inflamed mucosae from CD and UC patients have been previously reported in comparison to normal control mucosa. Most notable was the strong up-regulation of Mn-SOD in inflamed epithelium. It was hypothesized that in order to provide optimal protection against ROM-mediated damage, these changes should be coordinately counterbalanced by an increased H(2)O(2)-neutralizing capacity. Therefore, the same tissue samples were used to assess the levels, activities, and/or localization of the most prominent mucosal H(2)O(2)-related antioxidants CAT, GPO, glutathione (GSH), myeloperoxidase (MPO), and metallothionein (MT). Quantitative measurements showed that in both CD and UC patients, intestinal inflammation was associated with increased activities of CAT, GPO, and MPO, whereas the mucosal GSH content was unaffected and the concentration of MT was decreased. Despite this overall increase in mucosal H(2)O(2)-metabolizing enzyme capacity, immunohistochemical analysis revealed a differentially disturbed antioxidant balance in IBD epithelium and lamina propria. In the lamina propria, the risk of direct H(2)O(2)-mediated damage seemed to be restrained by the increasing numbers of CAT- and MPO-positive monocytes/macrophages and neutrophils that infiltrated the inflamed areas. On the other hand, MPO overexpression might increase the lamina propria levels of hypochlorous acid, a stable ROM with multiple pro-inflammatory effects. In the epithelium, the number of cells that expressed CAT remained unchanged during inflammation and GPO was found in only a very low and constant number of epithelial cells. In addition, the inflamed epithelium displayed decreased expression of the hydroxyl radical (OH(*)) scavenger MT. In view of the high epithelial SOD levels in inflamed IBD epithelium, it is speculated that the efficient removal of excess H(2)O(2) is hampered in these cells, thereby increasing not only the risk of detrimental effects of H(2)O(2) directly, but also those of its extremely reactive derivatives such as OH(*). Taken together, the results suggest an imbalanced and inefficient endogenous antioxidant response in the intestinal mucosa of IBD patients, which may contribute to both the pathogenesis and the perpetuation of the inflammatory processes.     

基于Nrf2/HO-1通路的中医药干预溃疡性结肠炎研究进展

溃疡性结肠炎（UC）是临床常见的一种慢性消化系统疾病,其病情反复且治疗难度大,发病机制复杂,与氧化应激反应相关。核转录因子E₂相关因子2（Nrf2）是抗氧化反应中的重要因子,可调控其下游血红素加氧-1（HO-1）的表达,发挥维持机体氧化还原稳态的作用。UC的病程中Nrf2与HO-1的生物活性及含量水平均下降,组织抗氧化和抗炎能力减弱,肠上皮细胞损伤,肠黏膜屏障破坏。目前西医临床上主要以控制炎症和缓解症状治疗为主,虽有一定疗效,但存在停药后易复发、长期用药不良反应多等问题。研究表明,中医药治疗手段丰富,治疗方法灵活,在UC的防治上有广阔的应用前景。近年来,以Nrf2/HO-1通路为切入点,中医药领域开展了大量关于UC中该信号的基础和临床实验,结果表明Nrf2/HO-1通路是中医药治疗UC的重要潜在靶点。基于虚实夹杂的病因病机,中医药以清热解毒燥湿、活血化瘀止痛、益气补中温里和标本兼治等法调控Nrf2/HO-1通路,提高组织抗氧化应激能力,维持促炎因子与抗炎因子平衡,缓解炎症反应,发挥对UC的治疗作用。该文总结和分析了中医药靶向Nrf2/HO-1信号通路干预UC的机制和作用,以期为科研人员更为全面认识中医药对UC中Nrf2/HO-1通路的作用机制提供帮助,旨在推动今后中医药合理运用于临床UC的防治。