二氢杨梅素对实验性结肠炎小鼠调节性B细胞的影响

目的 探讨二氢杨梅素(DHM)对实验性结肠炎小鼠的作用及其对调节性B细胞(Breg)的影响.方法 将小鼠随机分为正常组、DSS(葡聚糖硫酸钠)组、DSS+DHM组、DSS+mesalazine组(造模采用4％DSS连续饮用7 d).7 d后,每天分别给予灌胃水(0.2 mL/d)、DHM(40 mg/kg·d)和美沙拉...     

Oral administration of antigens from intestinal flora anaerobic bacteria reduces the severity of experimental acute colitis in BALB/c mice

Homeostasis between indigenous intestinal flora and host response may be broken in inflammatory bowel disease. The present study explores whether repeated oral administration of intestinal flora antigens can protect mice against dextran sodium sulphate (DSS)-induced colitis. Sonicates of Gram-positive, Gram-negative, or anaerobic resident bacteria isolated from mouse intestinal flora were fed to BALB/c mice by gastric gavage, with or without cholera toxin. After four weekly doses of 1 mg of these antigen preparations (or of PBS as control), DSS colitis was induced. One week later colitis was evaluated by clinical scores and histology. Mice fed a pool of the three sonicates had decreased inflammation scores (5 (1-14); median (range)) compared with PBS-fed control animals (15 (7-19); P < 0.05). Decreased inflammation was observed in mice fed anaerobic bacteria antigens (7 (6-11); P < 0.05 versus control), but not in mice fed a pool of Gram-positive and -negative sonicates (16 (12-16)). Inflammation scores of mice fed antigens with cholera toxin were similar to those of PBS-fed control animals. DSS-induced colitis can be suppressed by oral administration of normal intestinal flora antigens containing anaerobes.     

Studies on the mechanisms of mucous cell depletion in experimental colitis

Mucous cell depletion is an important histological discriminator in favour of ulcerative colitis. The mechanism of this change was investigated in guinea pigs with experimental colitis induced by intrarectal instillation of acetic acid or dinitrochlorobenzene (DNCB) in sensitized animals. Both models showed mucous cell depletion. Increased numbers of mucous cells were flash-labelled with tritiated thymidine (3HTdR), indicating an increased pool of proliferating (oligo-) mucous cells; mucous cell production was in fact increased absolutely compared with control animals, and there was a marked increase in the rate of turnover of mucous cells. The results indicate that in colitis there is (i) increased mucous cell production and (ii) an increased rate of mucous cell turnover due to increased loss of older mucous cells. It is concluded that the disease process in experimental colitis leads to premature discharge of mucin, so that mucous cells are no longer recognizable by light microscopy. This explains the increase in mucin production in ulcerative colitis, which occurs even in the presence of significant mucous cell depletion.     

Immunomodulation of experimental colitis: the role of NK1.1 liver lymphocytes and surrogate antigens--bystander effect.

The imbalance between Th1 pro-inflammatory and Th2 anti-inflammatory cytokine-producing cells plays a major role in the pathogenesis of inflammatory bowel disease (IBD). Induction of oral tolerance to colitis-extracted proteins was previously shown to down-regulate the anti-colon immune response, thereby alleviating experimental colitis. Immune bystander effect and liver-associated lymphocytes expressing the NK1.1 marker (NK1.1(+) LAL) have been suggested as being important in tolerance induction. The aims of the present study were to determine whether oral administration of inflammatory and non-inflammatory colon-extracted proteins of different species can induce peripheral immune tolerance and alleviate experimental colitis; and to examine the role of NK1.1(+) LAL in oral tolerance induction. Colitis was induced in C57/B6 mice by intracolonic instillation of trinitrobenzene sulphonic acid (TNBS). Mice received six oral doses of colonic proteins extracted from TNBS-colitis colonic wall, or normal colonic wall, from four different species. Standard clinical, macroscopic, and microscopic scores were used for colitis assessment. Serum interferon gamma (IFNgamma) and interleukin 10 (IL10) levels were measured by ELISA. To evaluate the role of NK1.1(+) LAL in maintaining the balance between immunogenic and tolerogenic subsets of cells, their cytotoxicity functions were tested in tolerized and non-tolerized-mice. The administration of mouse-derived colitis-extracted proteins, or of surrogate proteins extracted from normal mouse colon, or from rat or human inflammatory colons, was found to alleviate experimental colitis. Tolerized mice had less diarrhoea; showed a marked reduction of colonic ulceration, intestinal and peritoneal adhesions, wall thickness, and oedema; and demonstrated a significant improvement of all microscopic parameters for colitis. Induction of tolerance led to an increase in IL10 and a decrease in IFNgamma serum levels. NK1.1(+) LAL cytotoxicity function increased markedly in tolerized mice. In contrast, mice fed with proteins extracted from normal rat, rabbit, and human colon, or from rabbit inflammatory colon, developed severe colitis, with a marked increase in IFNgamma and a decrease in IL10 serum levels, and down-regulation of NK1.1(+) LAL function. This study has shown that oral tolerance can be induced in experimental colitis by means of the feeding of surrogate antigens; this alleviates experimental colitis. NK1.1(+) LAL cytotoxicity function is associated with peripheral tolerance induction and may help to maintain the Th1/Th2 immune balance.     

Quantification of mucosal leucocyte endothelial cell interaction by in vivo fluorescence microscopy in experimental colitis in mice.

Leucocyte recruitment to sites of intestinal inflammation is a crucial, multi-step process that leads ultimately to the accumulation of cells in the inflamed tissue. We established a new in vivo model system of experimental colitis to quantify leucocyte-endothelial cell interaction and leucocyte extravasation in the inflamed mucosa of the colon. Furthermore, we investigated the pathophysiological role of ICAM-1 in the intestinal microcirculation in vivo. Using the model of dextran sodium sulphate (DSS)-induced acute and chronic colitis in mice, in vivo microscopy was performed in the colonic submucosal postcapillary venules and the submucosal collecting venules in normal or inflamed murine colonic segments. ICAM-1 expression was blocked by an anti-ICAM-1 monoclonal antibody or by suppressing NF-kappaB activation by gliotoxin. Significant increases in leucocyte adhesiveness (51-fold in postcapillary venules, 30-fold in collecting venules, P < 0.01) and extravasation (6.5-fold) could be demonstrated as early as day 2 of DSS-application in acute colitis (P < 0.01). This was paralleled by increases in both the histological damage scores and myeloperoxidase activities. In chronic dextran sodium sulphate-induced colitis significant increases in leucocyte-endothelium interactions and leucocyte extravasation were observed. Blocking ICAM-1 expression with a monoclonal antibody or gliotoxin, leucocyte sticking and extravasation were significantly down-regulated in vivo compared to controls (> 70%; P < 0.01). This new model system offers the possibility to specifically assess the role of adhesion molecules in the colonic mucosa in vivo as well as to investigate and quantify the effectiveness of experimental therapeutic approaches in acute or chronic intestinal inflammation.     

Aluminum enhances inflammation and decreases mucosal healing in experimental colitis in mice

The increasing incidence of inflammatory bowel diseases (IBDs) in developing countries has highlighted the critical role of environmental pollutants as causative factors in their pathophysiology. Despite its ubiquity and immune toxicity, the impact of aluminum in the gut is not known. This study aimed to evaluate the effects of environmentally relevant intoxication with aluminum in murine models of colitis and to explore the underlying mechanisms. Oral administration of aluminum worsened intestinal inflammation in mice with 2,4,6-trinitrobenzene sulfonic acid- and dextran sodium sulfate-induced colitis and chronic colitis in interleukin 10-negative (IL10^−/−) mice. Aluminum increased the intensity and duration of macroscopic and histologic inflammation, colonic myeloperoxidase activity, inflammatory cytokines expression, and decreased the epithelial cell renewal compared with control animals. Under basal conditions, aluminum impaired intestinal barrier function. In vitro, aluminum induced granuloma formation and synergized with lipopolysaccharide to stimulate inflammatory cytokines expression by epithelial cells. Deleterious effects of aluminum on intestinal inflammation and mucosal repair strongly suggest that aluminum might be an environmental IBD risk factor.     

Oral treatment with Hev b 13 ameliorates experimental colitis in mice

Hev b 13 is an allergenic esterase obtained from the rubber tree Hevea brasiliensis, which has been shown recently to induce human mononuclear cells to release interleukin (IL)-10 in vitro. This immunoregulatory cytokine appears to play an important role in preventing inflammation and mucosal damage in animal models of colitis and in Crohn's disease patients. The aim of this study was to evaluate the therapeutic effect of Hev b 13 in mice with 2,4,6-trinitrobenzene sulphonic acid (TNBS)-induced colitis. Two hours following colonic instillation of the haptenizing agent, and daily thereafter for 5 days, Hev b 13 was administered by oral gavage. In mice treated with daily doses of either 0·5 mg/kg or 5·0 mg/kg of Hev b 13, the clinical signs of diarrhoea, rectal prolapse and body weight loss and also histological damage of the distal colon, were reduced significantly, in comparison with water-treated diseased mice. These findings suggest a potent anti-inflammatory activity of Hev b 13; this activity is speculated to be related to its interaction with cells from the immune system.     

Changes in the phosphorylation of claudins during the course of experimental colitis

The phosphorylation of the tight-junction protein claudin causes allosterism, endocytosis and changes in the polarity of the epithelium, thus affecting the barrier function. The phosphorylation status of claudin during the course of colitis has not been demonstrated. In the present study, we found that the phosphorylated claudin-4 and claudin-7 contents were increased in experimental colitis at days 6 and 8, and colonic phosphorylated claudin-6 was found to be increased at day 4 and day 8. Colonic phosphorylated claudin-5 was found to be decreased at day 4 but increased at day 6. These changes were accompanied by increases in intestinal permeability. In T84 cells, phosphorylated claudin-3 was increased at 48 h but decreased at 72 h after lipopolysaccharide (LPS) treatment. Phosphorylated claudin-5 and claudin-7 were decreased 72 h after LPS treatment, while phosphorylated claudin-6 was increased at 72 h after LPS treatment. We conclude that the phosphorylation of colonic claudins was changed during the course of colitis, which may be related to the change in the intestinal barrier function. Cytokine such as LPS was found to affect the phosphorylation of colonic claudins.     

Neither direct nor developmental exposure to bisphenol A alters the severity of experimental inflammatory colitis in mice

Abstract

Bisphenol A (BPA) is a high production volume endocrine disrupting chemical that is widely used in many consumer products and prevalent in human biological fluids. Recent studies suggest that BPA is active even at low levels, raising concern about its potential harm to human health. Given that the main route of exposure to BPA is oral, via the consumption of BPA-tainted foods and beverages, intestinal tissues could be particularly vulnerable to BPA-induced changes. A novel examination is reported here of whether oral exposure to BPA affects inflammatory bowel disease (IBD), an immune-mediated disease of the colon, using a mouse model of inflammatory colitis. In addition to direct exposure, the possible contribution of maternal BPA exposure to disease later in life is explored. It was found that daily oral exposure to BPA at the US Environmental Protection Agency described oral reference dose (50?µg/kg/day), either via direct oral route or through maternal sources (i.e. developmental exposure), did not significantly alter disease outcomes of body weight, survival, or colonic pathology. These observations suggest that oral BPA exposure, at this dose and for this exposure duration, has minimal influence on aspects of the inflammatory response that regulate immune mediated diseases of the gastrointestinal tract.     

小檗碱对TNBS诱导的小鼠结肠炎的治疗作用研究

本研究探讨小檗碱(BBR)对2,4,6-三硝基苯磺酸(TNBS)诱导的小鼠克罗恩病(CD)模型的免疫调节作用。通过TNBS诱导小鼠造成CD模型,观察BBR对CD小鼠的疗效,并进一步探讨BBR治疗CD的作用及其机制。实验小鼠随机分为4组,分别为对照组、TNBS组(CD模型组)、CD模型BBR高剂量治疗组和CD模型BBR低剂量治疗组。于造模后疾病恢复期取各组脾脏和结肠,观察肠道大体损伤及病理情况,检测了结肠局部的炎症因子水平和脾脏中T细胞亚群数量变化、相关的细胞因子和转录因子的表达。BBR组与TNBS组相比,体重恢复时间提前、速度加快;结肠大体损伤及病理评分下降,结肠匀浆上清炎症因子的水平亦下降;脾脏中Th1、Th17及Treg细胞亚群的比例均有下降,相应的转录因子也有不同水平的降低。结果表明BBR治疗对TNBS诱导的小鼠结肠炎有效,其可通过免疫调节和抗炎症作用而治疗CD。     

根尖牙乳头干细胞治疗葡聚糖硫酸钠诱导的实验性肠炎

文题释义： 根尖牙乳头干细胞：从根尖未发育完全年轻恒牙的根尖牙乳头组织中分离得到的一组具有自我更新、克隆形成和多向分化能力的间充质干细胞群。它在牙根形成和发育中起重要作用,是构建组织工程牙齿的种子细胞。 炎性肠病：主要包括克罗恩病和溃疡性结肠炎2种类型,是以肠道炎症和组织损伤为特征的慢性疾病。炎性肠病主要流行于欧美国家,在中国发病率呈增长趋势,其发病机制尚未完全阐明。在临床上其病情常有反复、迁延不愈,且有癌变风险,仍缺乏有效的治疗手段。目前有研究表明促炎性细胞因子和抗炎性细胞因子间失衡导致肠上皮损伤是炎性肠病的重要病理机制。 背景：根尖牙乳头干细胞在牙根形成和发育中起重要作用,但是关于根尖牙乳头干细胞免疫调节作用方面的研究并不充分。 目的：探讨根尖牙乳头干细胞对葡聚糖硫酸钠诱导的实验性肠炎的治疗作用。 方法：将24只C57/BL6小鼠随机平分为4组：正常对照组、模型对照组、根尖牙乳头干细胞组、FasL敲低根尖牙乳头干细胞组;除了正常对照组,其余3组小鼠饮用3%葡聚糖硫酸钠溶液诱导建立急性实验性肠炎模型,并在诱导开始第3天分别腹腔注射PBS、人根尖牙乳头干细胞(2×10⁶个细胞)及FasL敲减根尖牙乳头干细胞(2×10⁶个细胞)。诱导第10天处死小鼠,通过体质量及结肠长度测量、结肠组织病理学分析、结肠组织肿瘤坏死因子α、白细胞介素6、白细胞介素1β水平来评估结肠炎的严重程度,流式细胞分析比较各组小鼠肠系膜淋巴结调节性T细胞(Tregs)水平。 结果与结论：根尖牙乳头干细胞移植对葡聚糖硫酸钠诱导的小鼠肠炎具有治疗作用,小鼠肠炎症状、体质量减轻状况明显改善,结肠的组织病理学评分及3项炎症因子水平显著降低,肠系膜淋巴结中Tregs水平显著上调。FasL基因敲减后根尖牙乳头干细胞丧失其治疗效果;因此 Fas-FasL通路在根尖牙乳头干细胞发挥免疫调控机制中发挥了重要的作用。 ORCID: 0000-0001-6717-998X(顾永春) 中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程     

Efficacy of recombinant granulocyte colony-stimulating factor (rhG-CSF) in experimental colitis

Inflammatory bowel disease is associated with mucosal neutrophil recruitment and activatation, mediated in part by arachidonic acid metabolites. G-CSF attenuates the immune response to sepsis and ameliorates glycogen storage disease Ib-related colitis. These actions may be effected through the shedding of neutrophil adhesion molecules, or inhibition of proinflammatory mediator synthesis. Immune complex colitis was used to evaluate the effect of rhG-CSF on colonic mucosal inflammation, neutrophil recruitment and the generation of eicosanoids. Immune complex colitis was induced in White New Zealand rabbits. Animals were pretreated with rhG-CSF either 24 h before induction, or at induction, with dosages of 50 and 200 μg/kg. rhG-CSF caused a time- and dose-dependent neutrophilia in all animals. Pretreatment with rhG-CSF resulted in increased tissue myeloperoxidase levels, despite a histologically similar mucosal polymorphonuclear cell infiltrate between treated and control colitis groups. Leukotriene B₄ (LTB₄) and thromboxane B₂ (TXB₂) dialysis fluid levels were lower in treated animals, in particular in the groups receiving two doses (LTB₄: both P < 0.01; TXB₂: both P < 0.01. Prostaglandin E₂ (PGE₂) levels in dialysis fluid of the rhG-CSF-treated animals showed no difference from controls. In this model of experimental colitis, high-dose therapy with G-CSF resulted in a marked decrease of proinflammatory mediators, but mucosal generation of the protective PGE₂ was preserved. These results suggest that prolonged high-dose therapy with G-CSF may have anti-inflammatory effects in colitis.     

Neutralization of tumour necrosis factor (TNF) but not of IL-1 reduces inflammation in chronic dextran sulphate sodium-induced colitis in mice

The cytokines TNF and IL-1 have been implicated as mediators of the inflammatory processes in patients with inflammatory bowel disease (IBD). To investigate the role of these cytokines in mucosal inflammation we used anti-cytokine strategies in a mouse model of acute and chronic colitis. Mice which received 5% dextran sulphate sodium (DSS) in their drinking water showed signs of acute colitis on day 4, with severe weight loss and bloody diarrhoea. Chronic colitis was established after four cycles of feeding 5% DSS for 7 days and water for 10 days, with the mice showing diarrhoea but no weight loss. In acute colitis, treatment with anti-IL-1 reagents, anti-TNF MoAb, or dexamethasone (DEX) led to aggravation. By contrast, in chronic colitis, treatment of mice with several IL-1 activity-inhibiting reagents failed to show significant effects, whereas anti-TNF MoAb or DEX significantly reduced the colitis. We conclude that in acute colitis IL-1 and TNF are beneficial, whereas in chronic colitis, TNF but not IL-1 seems to play a major role in perpetuation of chronic inflammation.     

鹅肌肽降低高尿酸小鼠尿酸

目的探讨在实验性高尿酸小鼠模型中鹅肌肽对高尿酸的缓解作用,以及对肾功能相关指标的影响。方法采用酵母膏联合氧嗪酸钾及腺嘌呤的方法建立模型,4周之后采集血清、尿液、肾脏等标本,检测血尿酸、肌酐、尿素氮及尿酸清除率等指标。利用HE染色的方法观察肾脏形态学上的变化情。结果成功建立实验性高尿酸小鼠模型,鹅肌肽对高尿酸小鼠的干预后,其中血尿酸明显降低,尿酸清除率升高,而且具有浓度依赖性。结论鹅肌肽主要通过促进尿酸排泄,缓解高尿酸血症的作用。     

Transgenic over-expression of macrophage migration inhibitory factor renders mice markedly more susceptible to experimental colitis

Enhanced production of macrophage migration inhibitory factor (MIF) is recognized in patients with inflammatory bowel disease (IBD) and mice with experimental colitis; however, the precise molecular function of MIF in colitis is not fully understood. To further investigate this matter, we examined the pathological features of MIF transgenic mice with dextran sulphate sodium (DSS)-induced colitis. We generated transgenic mice carrying a murine MIF cDNA driven by a cytomegalovirus enhancer and a beta-actin/beta-globin promoter. Mice were orally administered 1-4% DSS in drinking water for 7 days. Clinical disease activity, survival and histological features were evaluated. The level of myeloperoxidase (MPO) activity in the colon tissue was measured to assess neutrophil infiltration. The level of corticosterone in the serum was measured by enzyme linked-immunosorbent assay (ELISA). MIF mRNA and protein were markedly up-regulated in the colon and serum obtained from MIF transgenic mice. The severity of the colitis induced by 1% DSS treatment was markedly higher in MIF transgenic mice than in wild-type mice. We also found that MPO activity was significantly higher in MIF transgenic mice than wild-type mice in response to DSS stimulation. Interestingly, the corticosterone level remained unchanged in MIF transgenic mice. MIF enhances DSS-induced colitis, in part via neutrophil accumulation and inhibition of glucocorticoid bioactivity.     

Non-lymphoid and lymphoid cells in acute, chronic and relapsing experimental colitis.

In rodents, intracolonic administration of ethanol 30% induces an acute colitis, while administration of 2,4,6-trinitrobenzene sulphonic acid (TNBS) in ethanol induces a longer lasting colitis. In the acute and chronic stages of experimental colitis, lymphoid and non-lymphoid cells were studied in the colon by immunohistochemistry. During the acute inflammation a high damage score of the colon was observed, which was related to an increase in the number of macrophages and granulocytes. Also a change in distributional patterns of macrophage subpopulations was found. The chronic stage of TNBS-ethanol-induced colitis was characterized by an increase in the number of lymphocytes, especially T cells. These data suggest that macrophages and granulocytes are important in the acute phase of experimental colitis, while lymphocytes play a pivotal role in the chronic stage. As most inflammatory bowel disease (IBD) patients have relapses during the chronic disease, we attempted to induce a relapse during experimental colitis by giving a second i.p. or s.c. dose of TNBS. This resulted in increased damage scores of the colon, new areas of ulceration and a further increase in macrophage numbers. No effect on the number of granulocytes was seen. These results indicate that it is possible to mimic relapses in experimental colitis by a second administration of TNBS, and suggest that the rats had been sensitized by the first dose of TNBS, given into the colon.     

显微镜结肠炎39例临床病理学分析

目的：探讨显微镜结肠炎(microscopic colitis,MC)的临床病理学特点、诊断及鉴别诊断。方法：对39例MC的临床特点、组织形态学、免疫组织化学及Masson染色结果进行分析,并复习相关文献。结果：39例MC中男性18例,女性21例,年龄25~74岁,中位年龄50岁。所有患者均有慢性水样腹泻症状,结肠镜检查正常,其中淋巴细胞性结肠炎(lymphocytic colitis,LC)19例,胶原性结肠炎(collagenous colitis,CC)20例。显微镜下表层上皮细胞间淋巴细胞数(intraepithelial lymphocytes,IEL)为(21.5±8.6),表层上皮下胶原带宽带为(14.3±4.0)μm,与对照组相比差异均有显著性(P<0.01)。结论：MC是老年患者慢性水样腹泻较常见的一种病因,诊断依赖于临床病史、结肠镜及病理组织学检查。     

Dextran sulfate sodium (DSS) induced experimental colitis in immunodeficient mice: Effects in CD4+-cell depleted,athymic and NK-cell depleted SCID mice

Administration of dextran sulfate to mice, given in the drinking water results in acute or subacute colonic inflammation, depending on the administration protocol. This colonic inflammation exhibits ulceration, healing and repair, and a therapeutic response that makes it valuable for the study of mechanisms that could act in the pathogenesis of human ulcerative colitis, a disease thought to have an immunologically dependent pathogenesis. To investigate if immunological mechanisms were involved in the induction of colonic inflammation in this model, mice with different degrees of immunodeficiency were used. It was shown that dextran sulfate induced colitis could be induced in Balb/c mice depleted of CD4⁺ helper T cells by treatment with monoclonal antibodies preceded by adult thymectomy. The depletion of CD4⁺ was verified by flow cytometric analysis. Furthermore, the colonic inflammation could equally be induced in athymic CD-1 nu/nu mice lacking thymusderived T cells, in T and B-cell deficient SCID mice, and also in SCID mice depleted of NK cells by treatment with anti-asialo GM1 antibodies. The NK-cell depletion was verified by measuring spleen NK-cell activity. The resulting colonic inflammation in all these types of deficient mice was qualitatively comparable, as shown by clinical and histological appearance. These results indicate that the presence of functional T, B and NK cells is not crucial for the induction of dextran sulfate colitis in mice.accepted by M. J. Parnham