Attenuation of dextran sodium sulphate induced colitis in matrix metalloproteinase-9 deficient mice

INTRODUCTIONMatrix metalloproteinases(MMPs)comprise a group of zinc and calcium-dependent endopeptidases that exhibit differential proteolytic activity against extracellular matrix(ECM)proteins.Based on substrate specificity,MMPs have been classically div…     

Development of colonic neoplasia in p53 deficient mice with experimental colitis induced by dextran sulphate sodium

BACKGROUND: Several animal models for human ulcerative colitis (UC) associated neoplasia have been reported. However, most neoplasias developed in these models have morphological and genetic characteristics different from UC associated neoplasia. AIMS: To establish a new colitis associated neoplasia model in p53 deficient mice by treatment with dextran sulphate sodium (DSS). METHODS: DSS colitis was induced in homozygous p53 deficient mice (p53(-/-)-DSS), heterozygous p53 deficient mice (p53(+/-)-DSS) and wild-type mice (p53+/+-DSS) by treatment with 4% DSS. Numbers of developed neoplasias were compared among the experimental groups, and macroscopic and microscopic features of the neoplasias were analysed. Furthermore, K-ras mutation and beta-catenin expression were assessed. RESULTS: p53(-/-)-DSS mice showed 100% incidence of neoplasias whereas the incidences in p53(+/-)-DSS and p53+/+-DSS mice were 46.2% and 13.3%, respectively. No neoplasias were observed in the control groups. The mean numbers of total neoplasias per mouse were 5.0 (p53(-/-)-DSS), 0.62 (p53(+/-)-DSS), and 0.2 (p53+/+-DSS). The number of neoplasias per mouse in the p53(-/-)-DSS group was significantly higher than that in the other DSS groups. The incidences of superficial type neoplasias were 91.7% in p53(-/-)-DSS mice, 75.0% in p53(+/-)-DSS mice, and 33.3% in p53+/+-DSS mice. The K-ras mutation was not detected in any of the neoplasias tested. Translocation of beta-catenin from the cell membrane to the cytoplasm or nucleus was observed in 19 of 23 (82.6%) neoplasias. CONCLUSIONS: The p53(-/-)-DSS mice is an excellent animal model of UC associated neoplasia because the morphological features and molecular genetics are similar to those of UC associated neoplasia. Therefore, this model will contribute to the analysis of tumorigenesis related to human UC associated neoplasia and the development of chemopreventive agents.     

Role of interleukin 15 in colitis induced by dextran sulphate sodium in mice

BACKGROUND AND AIMS: Interleukin (IL)-15 is a member of the IL-2 family, stimulating dendritic cells, natural killer (NK) cells, NK T cells and memory CD8+ T cells. IL-15 levels were elevated in the intestinal mucosa of inflammatory bowel diseases. Here we investigated the involvement of IL-15 in the pathogenesis of acute and chronic dextran sulphate sodium (DSS) induced colitis. METHODS: IL-15 knockout (KO) mice and control C57BL/6 mice were used to induce colitis with DSS in their drinking water. Survival rate, clinical activity of diseases, extent of tissue damage, leucocyte population, and cytokine production of lamina propria (LP) cells of the large intestines were assessed. RESULTS: IL-15 KO mice exhibited resistance to DSS induced acute colitis, as reflected by lower lethality, weight loss, clinical scores, and histological scores compared with those in control mice (p<0.05). The proportions of CD44(high) CD8+ T cells and NK cells in LP cells and levels of interferon (IFN)-gamma, tumour necrosis factor (TNF)-alpha, and IL-12p40 in culture supernatants of LP cells were reduced in IL-15 KO mice (p<0.05). In vivo depletion of CD8+ T cells and NK cells decreased levels of IFN-gamma, TNF-alpha, and IL-12p40 in culture supernatants of LP cells in C57BL/6 mice (p<0.01). In chronic colitis, weight loss and clinical scores were improved and levels of IFN-gamma, TNF-alpha, and IL-12p40 in culture supernatants of LP cells were also reduced in IL-15 KO mice (p<0.05). CONCLUSIONS: IL-15 plays an important role in the pathogenesis of both acute and chronic colitis induced by DSS in mice.     

Orally administered RDP58 reduces the severity of dextran sodium sulphate induced colitis

Oral administration of the novel anti-inflammatory peptide RDP58 markedly reduced the severity of dextran sulphate sodium (DSS) colitis as determined by clinical and quantitative histological criteria. The architecture of the colonic epithelium in DSS treated mice receiving RDP58 remained relatively normal compared with that of control DSS treated animals. 5-Bromo-2'-deoxyuridine (BrdU) labelling studies showed a pronounced inhibition of colonic epithelial cell proliferation during DSS treatment, which was partially reversed by RDP58 therapy. Remarkably, RDP58 almost completely prevented colonic epithelial cell death induced by DSS treatment. RDP58 therapy also inhibited the accumulation of neutrophils in the colon of DSS treated mice and effectively down regulated tumour necrosis factor (TNF) expression. Preservation of the intestinal mucosa by RDP58 may thus derive from its influence on TNF expression as well as additional anti-inflammatory properties. These findings indicate that RDP58 represents a new, orally available agent potentially useful in the treatment of inflammatory bowel disease.     

T辅助细胞在右旋葡聚糖硫酸钠诱导的小鼠结肠炎中的作用研究

目的　探讨Th1/Th2细胞在右旋葡聚糖硫酸钠 (DSS)诱导小鼠结肠炎中的作用。方法　利用细胞内细胞因子流式细胞检测法 ,测定DSS诱导急、慢性结肠炎小鼠肠黏膜和脾脏单核细胞 (SMC)中Th1/Th2比例。结果　①结肠炎急性期 ,小鼠肠黏膜固有层单核细胞 (LPMC)中Th1细胞为 (8.90± 1.2 3) % ,较对照组升高 (P <0 .0 5 ) ;Th2细胞占 (3.2 5± 1.2 5 ) % ,与对照组差异无显著性 (P >0 .0 5 ) ;Th1/Th2比例为 3.0 9± 1.18,较对照组升高 (P <0 .0 5 )。②在慢性期 ,肠黏膜LPMC中Th1、Th2细胞分别占 (5 .5 2± 1.2 8) %和 (10 .0 8± 1.75 ) % ,均较对照组升高 (P <0 .0 5 ) ;Th1/Th2比例为 0 .5 2± 0 .2 1,较对照组降低 (P <0 .0 5 )。脾脏SMC中Th1、Th2百分比、Th1/Th2比例 ,各期无明显差异。结论　DSS结肠炎是一种以肠黏膜免疫功能紊乱为主的炎症性病变。Th1优势反应可能与急性期损伤有关 ,而Th2优势反应可能在炎症的慢性化过程中发挥重要作用。     

葡聚糖硫酸钠诱导小鼠实验性结肠炎的临床和组织病理学特征研究

目的研究葡聚糖硫酸钠(DSS)诱导小鼠实验性结肠炎的临床和病理特征,以便于指导选择合适的溃疡性结肠炎(ulcerative colitis,UC)小鼠模型。方法予3%DSS溶液喂饲野生型C57BL/6成年小鼠诱导急性结肠炎模型,分别于第0天、第3天、第5天、第7天和第10天麻醉处死小鼠,取结肠观察不同时期结肠病理学特征,造模过程中连续观察并记录小鼠体质量、大便和死亡情况。结果小鼠造模第3天开始出现粪便潜血,第5天开始出现血便,第10天开始出现死亡小鼠,死亡率为30%。DSS诱导小鼠急性结肠炎模型疾病活动指数第3天后与饮用纯净水的小鼠比较明显增高(P0.05)。小鼠结肠炎造模第3天黏膜上皮细胞开始逐渐丧失,第7~10天最为严重;隐窝结构的紊乱从第3天开始发生,第7天出现固有层塌陷;炎性细胞浸润从第3天开始数量逐渐增加,第10天最为严重。DSS诱导小鼠急性结肠炎模型结肠组织病理评分第5天后与饮用纯净水的小鼠比较明显增高(P0.05)。结论 3%DSS诱导野生型C57BL/6成年小鼠急性结肠炎模型可用于UC的实验研究,第3天出现明显的炎症表现,第7天模型较理想,小鼠死亡少。     

Reduced susceptibility of mice overexpressing transforming growth factor α to dextran sodium sulphate induced colitis

Background—Transforminggrowth factor α (TGF-α) knockout mice have increased susceptibilityto dextran sodium sulphate (DSS) induced colitis.
Aim—To substantiatethe findings that TGF-α is a key mediator of colonic mucosalprotection and/or repair mechanisms by evaluating the susceptibility ofmice overexpressing TGF-α to DSS induced colitis.
Methods—TGF-αoverexpression was induced in transgenic mice by ZnSO₄administration in drinking water (TG+). Three groups were used ascontrols: one transgenic group without ZnSO₄ administration (TG−), and two non-transgenic littermate groups receivingZnSO₄ (Non-TG+) or only water (Non-TG−). Acute colitiswas induced in all groups by administration of DSS (5%, w/v) indrinking water for six days ad libitum.
Results—About 35-39%of the entire colonic mucosa was destroyed in Non-TG−, Non-TG+, andTG− animals compared with 9% in TG+ mice. The crypt damage score was18.7 (0.9), 18.2 (1.0), 18.9(0.8), and 6.8 (1.5) (means (SEM)) inNon-TG−, Non-TG+, TG−, and TG+ mice respectively. Mucin andbromodeoxyuridine staining were markedly enhanced in colons of TG+ micecompared with controls, indicating increased mucosal protection and regeneration.
Conclusions—Thesignificantly reduced susceptibility of mice overexpressing TGF-α toDSS further substantiates that endogenous TGF-α is a pivotal mediatorof protection and/or healing mechanisms in the colon.

Keywords:transforming growth factor α; epidermal growthfactor; dextran sodium sulphate; colitis; inflammatory bowel disease; transgenic mice

     

Vanilloid receptor-1 containing primary sensory neurones mediate dextran sulphate sodium induced colitis in rats

BACKGROUND AND AIMS: The role of sensory neurones in colitis was studied by chemical denervation of primary sensory neurones as well as antagonism of the vanilloid receptor-1 (VR-1) in rats prior to administration of dextran sulphate sodium (DSS) to induce colitis. METHODS: Neonatal rats were chemically denervated by subcutaneous administration of capsaicin; controls received capsaicin vehicle only. When animals reached maturity, colitis was induced by administration of 5% DSS in drinking water for seven days. Additionally, normal adult rats were treated with a VR-1 antagonist capsazepine (CPZ) or vehicle twice daily via an enema from day 0 to day 6 of the DSS regimen. Control rats were treated with an enema infusion of vehicle and 5% DSS, or without either an enema infusion or DSS in drinking water. For both groups of rats, severity of inflammation was quantitated by disease activity index (DAI), myeloperoxidase (MPO) activity, and histological examination. RESULTS: DSS induced active colitis in all control rats with resultant epithelial ulceration, crypt shortening, and neutrophil infiltration. Both neonatal capsaicinised rats and normal adult rats treated with CPZ enemas exhibited significantly lower levels of DAI, MPO, and histological damage compared with vehicle treated rats (p< 0.05). CONCLUSIONS: Neonatal capsaicinisation and local administration of CPZ prevents intestinal inflammation in a well established model of colitis indicating that primary sensory neurones possessing VR-1 receptors are required in the propagation of colonic inflammation.     

双歧杆菌对葡聚糖硫酸钠(DSS)诱导的小鼠结肠炎肠道黏膜的保护作用及其对NF-κB的影响

目的随着对炎症性肠病(inflammatory bowel disease,IBD)发病机制的深入研究,肠道微生态与免疫系统的密切关系日益受到重视。本研究旨在评价双歧杆菌单一活菌制剂对葡聚糖硫酸钠(dextran sulfate sodium,DSS)诱导的小鼠结肠炎肠道黏膜的保护作用及其可能作用机制。方法 BALB/c小鼠饮用含7%DSS的饮用水10d,诱导小鼠亚急性结肠炎的模型,模拟人类溃疡性结肠炎。空白对照组(n=6)饮用未添加DSS的饮用水。饮用DSS的小鼠(n=37)随机分为5组,分别给予不同的药物治疗:①提前给予双歧杆菌治疗组(Pre-BIF):在小鼠暴露于DSS前10d及暴露过程中给予双歧杆菌治疗;②强的松治疗组(PRED):在小鼠暴露于DSS同时给予强的松治疗;③双歧杆菌治疗组(BIF):小鼠暴露于DSS的同时给予双歧杆菌治疗;④强的松+双歧杆菌联合治疗组(PRED+BIF):小鼠暴露于DSS的同时给予强的松和双歧杆菌联合治疗;⑤生理盐水对照组(NS):小鼠暴露于DSS的同时给予生理盐水作为对照。不同药物皆以溶液形式灌胃给药。从4方面评价各组的处理反应:①一般情况:包括体重、结肠长度、大体评分;②组织病理评分;③化学比色法检测病变组织髓过氧化物酶(MPO)活性;④免疫组织化学方法检测活化的NF-κB。结果 Pre-BIF治疗组和BIF治疗组的大体评分、组织病理评分、MPO活性与NS对照组相比明显改善(P0.01;P0.05)。Pre-BIF治疗组、BIF治疗组和PRED+BIF治疗组的NF-κB活性评分显著低于NS对照组(P均0.01)。结论双歧杆菌对小鼠DSS结肠炎肠道黏膜具有保护作用,提前应用保护效果更好。双歧杆菌对肠道黏膜的保护作用与抑制NF-κB的活化有关。     

硝酸酯在小鼠急性实验性结肠炎中的治疗作用

目的 观察硝酸酯对小鼠急性实验性结肠炎的疗效.方法 将40只BALB/c小鼠均分为模型组和治疗组,模型组予4％葡聚糖硫酸钠(DSS)溶液,治疗组予4％DSS溶液和1.5 g/L硝酸酯溶液,均连续饮用7d.对小鼠疾病活动指数(DAI)进行评分.取小鼠结肠组织进行苏木精-伊红染色和髓过氧化物酶(MPO)免疫组织化学染色并进行观察.分别用MPO和一氧化氮检测试剂盒检测结肠组织MPO和一氧化氮活性.统计学处理采用t检验.结果 第6和第7天治疗组和模型组的DAI差异有统计学意义(t=5.12和6.72,P=0.012和0.008).第7天时模型组组织学评分(2.5±0.5)高于治疗组(1.9±0.4),差异有统计学意义(t=3.82,P＜0.01).与模型组相比,治疗组小鼠结肠组织病理损伤明显减轻,中性粒细胞浸润减少.第7天时模型组MPO活性、NO2-浓度、NO3-浓度分别为(2.8±0.6) U/g、(10.4±4.3) mmol/g、(100.3±50.1)mmol/g,治疗组则分别为(1.5±0.3)U/g、(17.5±7.0)mmol/g、(190.7±85.3) mmol/g,差异均有统计学意义(t=11.23、3.81、4.50,P均＜0.01).结论 硝酸酯可减轻DSS诱导的小鼠急性实验性结肠炎.     

Contrasting action of IL-12 and IL-18 in the development of dextran sodium sulphate colitis in mice

BACKGROUND: Interleukin (IL)-12 and IL-18 are major interferon (IFN)-gamma-inducing factors that collaborate with each other. The present study was conducted to determine the distinct roles of IL-12 and IL-18 in the development of dextran sulphate sodium (DSS) colitis in mice. METHODS: Colitis was induced in IL-12p35(-/-), IL-18(-/-), IL-18 receptor(-/-) and control mice with DSS. Clinical and histopathological analysis was conducted using survival rate, weight loss score, diarrhoea score, bloody stool score and histological score. In addition, cytokine production by lamina propria mononuclear cells (LPMCs) was examined using the specific enzyme-linked immunoassay. RESULTS: IL-12p35(-/-) mice developed only a mild disease associated with no lethality and few histopathological abnormalities. In contrast, IL-18(-/-) and IL-18R(-/-) mice developed more severe colitis associated with high lethality and more histopathological abnormalities compared with control mice. LPMCs from DSS-fed IL-18(-/-) mice produced significantly higher amounts of IFN-gamma, while LPMCs from DSS-fed IL-12(-/-) mice produced lower amounts of IFN-gamma and tumour necrosis factor (TNF)-alpha compared with control mice. CONCLUSION: These results suggest that IL-18 might function with manners different from IL-12 at some pathological conditions in the development of colitis.     

双歧杆菌对小鼠急性实验性结肠炎作用的研究

目的 研究双歧杆菌在葡聚糖硫酸钠(DSS)诱导的急性溃疡性结肠炎小鼠模型中的作用.方法 将80只BALB/C小鼠均分为8组,每组10只.除正常对照组、单纯0501菌株组、单纯c122菌株组外,其他5组动物均给予5%DSS造模7 d.在造模开始前2 d,阴性对照组用0.9%氯化钠液灌肠、阳性对照组用柳氮磺胺吡啶(SASP)20 mg/ml灌肠、DSS+0501菌株组用1×109 CFU/ml 0501菌液灌肠、DSS+c122菌株组用1×109 CFU/ml c122液菌灌肠.9 d后处死动物取结肠标本,行H-E染色,观察镜下结肠病理变化,并用免疫组化染色、RT-PCR技术分析结肠黏膜中白细胞介素(IL)-10 mRNA及蛋白表达.结果 DSS造模过程中给予双歧杆菌灌肠小鼠(0501菌株组和c122菌株组)的结肠炎性反应程度较模型组加重,结肠黏膜IL-10表达减少.而单纯给予双歧杆菌灌肠小鼠(单纯0501菌株组和单纯c122菌株组)未见结肠炎表现.结论 双歧杆菌的某些菌株在结肠黏膜屏障功能受损情况下,可加重溃疡性结肠炎小鼠的结肠黏膜损伤.     

Occurrence of dysplasia and adenocarcinoma after experimental chronic ulcerative colitis in hamsters induced by dextran sulphate sodium.

In this study, long term dextran sulphate sodium administration was studied to ascertain whether colorectal carcinoma could be produced in patients with long standing ulcerative colitis. Simultaneously, changes in the intestinal microflora were analysed. Low grade to high grade dysplasia was seen in three of the five hamsters treated with 1% dextran sulphate sodium solution for 100 days, while no dysplasia was detected in the eight animals concomitantly treated with metronidazole, an antianerobic microbial agent, which prevents colonic ulceration. In these two groups, none of the animals developed colorectal cancer over 100 day period. In a group treated for 180 days, seven of the eight animals had dysplasia, and one had two adenomas. Furthermore, four of the eight animals had adenocarcinoma in the transverse colon; they were protruding well differentiated adenocarcinoma in one and non-protruding lesions infiltrating into the musclaris propria in three. The three non-protruding infiltrating adenocarcinomas were classified to be well differentiated adenocarcinoma in one and mucinous adenocarcinoma in two, resembling the type of cancer which complicates ulcerative colitis in man.     

Blueberry husks,rye bran and multi-strain probiotics affect the severity of colitis induced by dextran sulphate sodium

Objective. The enteric microbiota is a pivotal factor in the development of intestinal inflammation in humans but probiotics, dietary fibres and phytochemicals can have anti-inflammatory effects. The aim of this study was to evaluate the therapeutic effect of multi-strain probiotics and two conceivable prebiotics in an experimental colitis model. Material and methods. Sprague-Dawley rats were fed a fibre-free diet alone or in combination with Lactobacillus crispatus DSM 16743, L. gasseri DSM 16737 and Bifidobacterium infantis DSM 15158 and/or rye bran and blueberry husks. Colitis was induced by 5% dextran sulphate sodium (DSS) given by oro-gastric tube. Colitis severity, inflammatory markers, gut-load of lactobacilli and Enterobacteriaceae, bacterial translocation and formation of carboxylic acids (CAs) were analysed. Results. The disease activity index (DAI) was lower in all treatment groups. Viable counts of Enterobacteriaceae were reduced and correlated positively with colitis severity, while DAI was negatively correlated with several CAs, e.g. butyric acid. The addition of probiotics to blueberry husks lowered the level of caecal acetic acid and increased that of propionic acid, while rye bran in combination with probiotics increased caecal CA levels and decreased distal colonic levels. Blueberry husks with probiotics reduced the incidence of bacterial translocation to the liver, colonic levels of myeloperoxidase, malondialdehyde and serum interleukin-12. Acetic and butyric acids in colonic content correlated negatively to malondialdehyde. Conclusions. A combination of probiotics and blueberry husks or rye bran enhanced the anti-inflammatory effects compared with probiotics or dietary fibres alone. These combinations can be used as a preventive or therapeutic approach to dietary amelioration of intestinal inflammation.     

Attenuation of acid induced oesophagitis in VR-1 deficient mice

BACKGROUND AND AIMS: Activation of the vanilloid receptor subtype 1 (VR-1) results in release of proinflammatory peptides which initiate an inflammatory cascade known as neurogenic inflammation. We investigated its role in an acute model of surgically induced oesophagitis. METHODS: Oesophagitis was induced by pyloric ligation in wild-type and VR-1 deficient mice. A subset of animals were administered the VR-1 antagonist capsazepine, famotidine, or omeprazole one hour before surgery. Five hours after surgery, myeloperoxidase activity (MPO), histological damage scores, intragastric pH, and immunocytochemical analysis of substance P (SP) receptor endocytosis were determined. RESULTS: Oesophagitis induced knockout mice exhibited significantly lower levels of MPO activity, histological damage scores, and SP receptor endocytosis than wild-type mice. Inflammatory parameters were significantly reduced by acid inhibition and capsazepine in wild-type mice. CONCLUSIONS: We conclude that acute acid induced oesophagitis is reduced in animals lacking VR-1. This suggests that acid induced oesophagitis may act through VR-1 and that inhibition of the receptor may reduce inflammation.     

Intestinal blood flow in murine colitis induced with dextran sulfate sodium

The aim of this study was to assess whether colitis induced by dextran sulfate sodium (DSS; 10% in tap water for 7 days) in BALB/c mice is associated with changes in intestinal blood flow. After anaesthesia, systemic hemodynamic variable and regional blood flows and resistances in various organs were measured in both control and DSS-treated mice. Mean arterial blood pressure was significantly lower in DSS-treated mice than in controls (56 ± 4 vs 66 ± 3 mm Hg; P < 0.05), but no differences were found in regional blood flows to or vascular resistances in the lungs, liver, stomach, small intestine (upper, middle, and lower part), cecum, mesentery + pancreas, spleen, kidneys, brain, and skin. However, compared to the control mice, blood flows in the middle (0.88 ± 0.13 vs 0.55 ± 0.09 ml/min/g; P < 0.05) and distal (0.69 ± 0.11 vs 0.29 ± 0.05 ml/min/g; P < 0.05) colon were significantly higher, and vascular resistances in the proximal (0.87 ± 0.21 vs 1.36 ± 0.21 mm Hg min/ml/100 g; P < 0.05), middle (0.60 ± 0.10 vs 1.46 ± 0.35 mm Hg min/ml 100 g; P < 0.05) as well as distal (0.90 ± 0.25 vs 2.67 ± 0.49 mm Hg min/ml/100 g; P < 0.05) colon were significantly lower in mice with experimental colitis. Interestingly, there was a gradient in the intestinal blood flow in control mice from the upper small intestine (2.79 ± 0.72 ml/min/g) down to the distal colon (0.29 ± 0.05 ml/min/g); such a gradient was also present in the colitis mice. It is concluded that DSS-induced colitis in mice is associated with microcirculatory disturbances in the colon, mainly in its middle and distal parts.