中华消化杂志
中華消化雜誌
중화소화잡지
Chinese Journal of Digestion
2010年
2期
81-86
,共6页
炎症性肠病%血吸虫属%紧密连接蛋白%模型%动物
炎癥性腸病%血吸蟲屬%緊密連接蛋白%模型%動物
염증성장병%혈흡충속%긴밀련접단백%모형%동물
Inflammatory bowel diseases%Schistosorna%Tight junctions protein%Modles,animal
目的 研究灭活血吸虫卵对2,4,6一三硝基苯磺酸(tinitrobenzene sulfonic acid,TNBS)诱导小鼠结肠炎肠黏膜紧密连接蛋白ZO-1和Occludin基因及蛋白表达影响及其机制.方法 清洁级BALB/C雌性小鼠50只分成对照组(10只)、TNBS+0.9%氯化钠溶液组(20只)和TNBS+血吸虫卵组(20只).TNBS+血吸虫卵组在造模前第3、14天分别给予腹腔注射冰冻灭活血吸虫卵10 000个(1 ml冰0.9%氯化钠溶液混悬液);TNBS+0.9%氯化钠溶液组给予相同体积的冰0.9%氯化钠溶液腹腔注射.后两组予TNBS溶液灌肠(100 mg/kg)建立结肠炎模型,建模后第7天处死存活小鼠,观察各组小鼠结肠的大体形态和HE染色光镜下病理特征;荧光实时定量PCR法测定结肠组织的Occludin和ZO-1基因表达;Western印迹法检测蛋白表达;免疫组化法测定结肠组织紧密连接蛋白表达分布.结果 TNBS+血吸虫卵组小鼠死亡率较TNBS+0.9%氯化钠溶液组明显下降(15%比30%).TNBS+0.9%氯化钠溶液组组织学评分为(4.21±0.40)分,较TNBS+血吸虫卵组和对照组高[(1.74±0.10)和(1.06±0.20)分,P<0.05].TNBS+0.9%氯化钠溶液组ZO-1和Occludin mRNA表达量较对照组显著下降(P<0.01),而TNBS+血吸虫卵组较TNBS+0.9%氯化钠溶液组显著增加(P<0.05).TNBs+0.9%氯化钠溶液组ZO-1蛋白相对灰度值较正常对照组降低50.3%(P<0.05),而TNBS+血吸虫卵组较TNBS+0.9%氯化钠溶液增加41.1%(P<0.05);TNBS+0.9%氯化钠溶液组Occludin相对灰度值较对照组下降48.7%(P<0.05),而血吸虫卵组较TNBS+0.9%氯化钠溶液组增加23.6%(P<0.05).ZO-1、Occludin蛋白染色强度TNBS+0.9%氯化钠溶液组分布均较对照组间增强(P<0.01),而TNBS+血吸虫卵组染色强度信号分布较TNBS+0.9%氯化钠溶液组显著增加(P<0.05).结论 灭活血吸虫卵能在细胞水平加强紧密连接蛋白ZO-1、Occludin聚集及表达,通过稳定紧密连接蛋白,增加肠道黏膜屏障功能,显著改善实验性结肠炎的肠道炎症反应.
目的 研究滅活血吸蟲卵對2,4,6一三硝基苯磺痠(tinitrobenzene sulfonic acid,TNBS)誘導小鼠結腸炎腸黏膜緊密連接蛋白ZO-1和Occludin基因及蛋白錶達影響及其機製.方法 清潔級BALB/C雌性小鼠50隻分成對照組(10隻)、TNBS+0.9%氯化鈉溶液組(20隻)和TNBS+血吸蟲卵組(20隻).TNBS+血吸蟲卵組在造模前第3、14天分彆給予腹腔註射冰凍滅活血吸蟲卵10 000箇(1 ml冰0.9%氯化鈉溶液混懸液);TNBS+0.9%氯化鈉溶液組給予相同體積的冰0.9%氯化鈉溶液腹腔註射.後兩組予TNBS溶液灌腸(100 mg/kg)建立結腸炎模型,建模後第7天處死存活小鼠,觀察各組小鼠結腸的大體形態和HE染色光鏡下病理特徵;熒光實時定量PCR法測定結腸組織的Occludin和ZO-1基因錶達;Western印跡法檢測蛋白錶達;免疫組化法測定結腸組織緊密連接蛋白錶達分佈.結果 TNBS+血吸蟲卵組小鼠死亡率較TNBS+0.9%氯化鈉溶液組明顯下降(15%比30%).TNBS+0.9%氯化鈉溶液組組織學評分為(4.21±0.40)分,較TNBS+血吸蟲卵組和對照組高[(1.74±0.10)和(1.06±0.20)分,P<0.05].TNBS+0.9%氯化鈉溶液組ZO-1和Occludin mRNA錶達量較對照組顯著下降(P<0.01),而TNBS+血吸蟲卵組較TNBS+0.9%氯化鈉溶液組顯著增加(P<0.05).TNBs+0.9%氯化鈉溶液組ZO-1蛋白相對灰度值較正常對照組降低50.3%(P<0.05),而TNBS+血吸蟲卵組較TNBS+0.9%氯化鈉溶液增加41.1%(P<0.05);TNBS+0.9%氯化鈉溶液組Occludin相對灰度值較對照組下降48.7%(P<0.05),而血吸蟲卵組較TNBS+0.9%氯化鈉溶液組增加23.6%(P<0.05).ZO-1、Occludin蛋白染色彊度TNBS+0.9%氯化鈉溶液組分佈均較對照組間增彊(P<0.01),而TNBS+血吸蟲卵組染色彊度信號分佈較TNBS+0.9%氯化鈉溶液組顯著增加(P<0.05).結論 滅活血吸蟲卵能在細胞水平加彊緊密連接蛋白ZO-1、Occludin聚集及錶達,通過穩定緊密連接蛋白,增加腸道黏膜屏障功能,顯著改善實驗性結腸炎的腸道炎癥反應.
목적 연구멸활혈흡충란대2,4,6일삼초기분광산(tinitrobenzene sulfonic acid,TNBS)유도소서결장염장점막긴밀련접단백ZO-1화Occludin기인급단백표체영향급기궤제.방법 청길급BALB/C자성소서50지분성대조조(10지)、TNBS+0.9%록화납용액조(20지)화TNBS+혈흡충란조(20지).TNBS+혈흡충란조재조모전제3、14천분별급여복강주사빙동멸활혈흡충란10 000개(1 ml빙0.9%록화납용액혼현액);TNBS+0.9%록화납용액조급여상동체적적빙0.9%록화납용액복강주사.후량조여TNBS용액관장(100 mg/kg)건립결장염모형,건모후제7천처사존활소서,관찰각조소서결장적대체형태화HE염색광경하병리특정;형광실시정량PCR법측정결장조직적Occludin화ZO-1기인표체;Western인적법검측단백표체;면역조화법측정결장조직긴밀련접단백표체분포.결과 TNBS+혈흡충란조소서사망솔교TNBS+0.9%록화납용액조명현하강(15%비30%).TNBS+0.9%록화납용액조조직학평분위(4.21±0.40)분,교TNBS+혈흡충란조화대조조고[(1.74±0.10)화(1.06±0.20)분,P<0.05].TNBS+0.9%록화납용액조ZO-1화Occludin mRNA표체량교대조조현저하강(P<0.01),이TNBS+혈흡충란조교TNBS+0.9%록화납용액조현저증가(P<0.05).TNBs+0.9%록화납용액조ZO-1단백상대회도치교정상대조조강저50.3%(P<0.05),이TNBS+혈흡충란조교TNBS+0.9%록화납용액증가41.1%(P<0.05);TNBS+0.9%록화납용액조Occludin상대회도치교대조조하강48.7%(P<0.05),이혈흡충란조교TNBS+0.9%록화납용액조증가23.6%(P<0.05).ZO-1、Occludin단백염색강도TNBS+0.9%록화납용액조분포균교대조조간증강(P<0.01),이TNBS+혈흡충란조염색강도신호분포교TNBS+0.9%록화납용액조현저증가(P<0.05).결론 멸활혈흡충란능재세포수평가강긴밀련접단백ZO-1、Occludin취집급표체,통과은정긴밀련접단백,증가장도점막병장공능,현저개선실험성결장염적장도염증반응.
Objective To observe the possible effect of inactivated schistosome ova on the expression of intestinal epithelial tight junctions ZO-1 and Occludin gene in mouse colitis induced by tinitrobenzene sulfonic acid (TNBS) and its mechanism. Methods Fifty mice were divided into control group (group A, n= 10),TNBS plus normal saline(NS) group(group B, n= 20) and TNBS plus inactivated schistosome ova group(group C, n= 20). Group C was exposed to 10 000 freeze-killed schistosome ova by intraperitoneal injection at day 14 and day 3 before colitis induction. Meanwhile,group B was exposed to 1 ml NS by intraperitoneal injection. The mice in group B and C were challenged with 3 mg TNBS to induce colitis. All mice were killed 7-day after colitis induction and assessed with following variables including mortality, pathological change with HE staining of colon.The transcription levels of ZO-1 and Oceludin in colon tissues were examined using Real-time PCR.The expression and distribution of ZO-1 and Occludin proteins were detected by Western blotting and immunohistochemistry. Results In comparison with group B inactivated schistosome ova most effectively reduced the mortality (30% vs 15 %) and histopathologic severity of TNBS-induced colitis (4.21±0.40 vs 1.74±0.10). The transcription levels of ZO-1 and Occludin in group B were decreased compared with those in group A and group C (P<0.01). When compared with group B,group C showed a significant elevation of the alteration of ZO-1, Occludin proteins expression and localization. Conclusion The results clearly show that schistosome ova treatment reduced the severity of experimental colitis through the regulation of tight junction proteins.
