【摘要】 目的:分析慢性应激对大鼠结肠黏膜组织学、超微结构及杯状细胞黏蛋白MUC2表达的影响。方法:实验于2004-04/2005-04在武汉大学人民医院和宜昌市中心人民医院完成。选择健康成年Wistar大鼠36只,随机分为应激组、对照组和氨基胍组,每组12只。应激组和氨基胍组每天置于束缚笼内2h,氨基胍组同时给予氨基胍150mg/(kg·d)腹腔注射进行干预,对照组自由活动。持续14d后处死动物,病理组织切片分别观察各组结肠黏膜组织学及上皮细胞超微结构的变化,用免疫组化方法观察结肠杯状细胞内黏蛋白MUC2的蛋白表达情况,并用硝酸还原酶法测定结肠黏膜组织一氧化氮、分泌型一氧化氮合酶的含量。结果:36只大鼠在实验中无死亡,全部进入结果分析。①慢性应激大鼠结肠黏膜炎性细胞数目增加,应激组中性粒细胞、单核细胞明显多于对照组[(71.33±9.84),(30.58±3.82)/mm2,P<0.01];[(49.58±6.21),(26.33±4.56)/mm2,P<0.01];使用氨基胍组结肠黏膜中性粒细胞和单核细胞明显低于应激组,差异具有显著性[(32.92±4.48),(71.33±9.84)/mm2,P<0.01];[(28.50±4.66),(49.58±6.21)/mm2,P<0.01]。②应激组杯状上皮细胞见较多黏液分泌后残留的囊泡,胞浆黏蛋白MUC2表达的平均吸光度值低于对照组[(0.22±0.03),(0.26±0.12),P<0.01],也低于氨基胍组[(0.22±0.03),(0.25±0.02),P<0.01],而氨基胍组杯状细胞内黏蛋白丰富。③应激组结肠组织一氧化氮含量明显高于对照组[(41.72±4.70),(31.76±4.58)μmol/g,P<0.01];也高于氨基胍组[(41.72±4.70),(32.20±6.86)μmol/g,P<0.01];氨基胍组与对照组比较差异无显著性(P>0.05)。分泌型一氧化氮合酶的活性应激组也明显高于对照组[(6.47±0.95),(3.81±0.64)nkat/g,P<0.01],也高于氨基胍组[(6.47±0.95),(3.50±0.54)nkat/g,P<0.01];氨基胍组与对照组比较差异无显著性。④应激组大鼠结肠上皮细胞线粒体肿胀、嵴减少、消失,线粒体空泡变性,部分区域溶酶体增多;细胞间连接松弛,紧密连接间隙增大,上皮细胞表面微绒毛减少、稀疏、脱落。对照组大鼠结肠上皮细胞内细胞器完整,细胞间连接正常,细胞表面微绒毛完整。氨基胍组仅见轻微的细胞间连接的变化。结论:慢性应激损伤肠黏膜屏障,引起结肠黏膜炎性细胞浸润和上皮细胞超微结构的变化,并影响黏液分泌细胞的功能。这种形态学的变化与慢性应激所导致的结肠组织分泌型一氧化氮合酶活化、一氧化氮含量增加有关。氨基胍对慢性应激诱导的结肠黏膜损伤起保护作用。更多还原

【Abstract】 AIM: To analyze the effect of chronic stress on the histology of colonic mucosa, ultrastructural change and the expression of MUC2 protein in goblet cells of rats.METHODS: The experiment was completed in the laboratory of Renmin Hospital of Wuhan University and Yichang Central People’s Hospital from April 2004 to April 2005. Thirty-six healthy Wistar rats were randomly divided into control group (n=12), stress group (n=12), aminoguanidine group (n=12). Rats in the stress group and aminoguanidine group were submitted to immobilization stress for 2 hours per day. At the same time, those in the aminoguanidine group were intraperitoneally injected with aminoguanidine (150 mg/kg per day), while those in the control group moved freely. All the rats were killed after 14 days. Pathological tissue sections were made to observe the changes of histology in colonic mucosa and the ultrastructure in colon epithelial cells. The expression of MUC2 protein in goblet cells was determined by immuohistochemistry. The concentration of nitric oxide and inducible nitric oxide synthase in colonic mucosa were measured by the nitrate reductase method. RESULTS: All the 36 rats were involved in the analysis of results. ① The number of inflammatory cells in colonic mucosa was increased in rats underwent chronic stress, the numbers of neutrophils and monocytes in the aminoguanidine group were obviously more than those in the control group (71.33±9.84), (30.58±3.82)/mm2, P < 0.01; (49.58±6.21), (26.33±4.56)/mm2, P < 0.01 . The numbers of neutrophils and monocytes in colonic mucosa in the aminoguanidine group were significantly lower than those in the aminoguanidine group (32.92±4.48), (71.33±9.84)/mm2, P < 0.01; (28.50±4.66), (49.58±6.21)/mm2, P < 0.01 . ② There were many remainder vesicles after grume excretion of colonic goblet epithelial cells in the stress group, the expression of MUC2 protein in the aminoguanidine group was lower than that in the control group (0.22±0.03), (0.26±0.12), P < 0.01 , also lower than that in the aminoguanidine group (0.22±0.03), (0.25±0.02), P < 0.01 , but the mucin in the aminoguanidine group was abundant. ③ The content of nitric oxide in colonic mucosa in the aminoguanidine group was obviously higher than that in the control group (41.72±4.70), (31.76±4.58) μmol/g, P < 0.01 , also higher than that in the aminoguanidine group (41.72±4.70), (32.20±6.86) μmol/g, P < 0.01 , there was no significant difference between the aminoguanidine group and control group (P > 0.05). The activity of inducible nitric oxide synthase in the aminoguanidine group was obviously higher than that in the control group (6.47±0.95), (3.81±0.64) nkat/g, P < 0.01 , also higher than that in the aminoguanidine group (6.47±0.95), (3.50±0.54) nkat/g, P < 0.01 , and there was no significant difference between the aminoguanidine group and control group (P > 0.05). ④ There were some ultrastructural damages in the aminoguanidine group. The mitochondria swelled up, vacuolation was seen in some regions. The lean in mitochondria was diminuted. The tight junction was loose. In the control group, the organells in mitochondria were integrity, the bond between epithelial cell was tight, microvillus on cell surface was integrity. There were slight changes of mitochondria and tight junction in the amiguanidine group. CONCLUSION: Chronic stress can bring damages to colonic mucosa barrier, it can induce inflammatory cells of colonic mucosa increase and the ultrastructural change of colonic epithelial cell, it also affects the function of grume excretion cell. The changes may be related to the activation of inducible nitric oxide synthase and increase of nitric oxide content. Aminoguanidine has a protective effect on stress-induced damages in colonic mucosa.更多还原