4-氨基水杨酸对大鼠实验性结肠炎中性粒细胞趋化、活化及凋亡的影响

Effects of 4-aminosalicylic acid on migration, activation and apoptosis of polymorphonuclear neutrophil in rats with experimental colitis

目的观察4-氨基水杨酸（4-ASA）对三硝基苯磺酸（TNBs）诱导的大鼠实验性结肠炎炎症损伤、肠组织一氧化氮合酶（iNOs）表达、血中性粒细胞（PMN）凋亡及血清白细胞介素（IL）-8水平等的影响，探讨4-ASA对炎症性肠病（IBD）的治疗作用及机制。方法40只SD大鼠分为正常对照组（n=10）和实验组（n=30），实验组建立大鼠结肠炎模型。建模第5天将实验组按照处理方式分为实验对照组（n=10，0．9％氯化钠1ml灌肠），5-ASA组[n=10，5-ASA液（200mg／kg）1m1灌肠]和4-ASA组[n=10，4-ASA液（200mg／kg）1ml灌肠]。连续治疗7d后处死动物，取病变段肠组织，行结肠大体损伤及结肠组织学损伤评分；生化法检测髓过氧化物酶活性；免疫组化法检测肠组织iNOS表达量；流式细胞术检测血PMN凋亡率；酶联免疫吸附法检测血清IL-8浓度。结果经4-ASA治疗7d后，4-ASA组大鼠体重明显较实验对照组增加（P〈0．01，t=14．09）；疾病活动指数评分、大体评分、组织学评分和MPO活性显著较实验对照组降低（t值分别=7．87、18．37、6．66和19．60，P值均〈0．01）。而5-ASA组与4-ASA组间上述各指标差异均无统计学意义（P值均〉0．05）。实验对照组肠组织iN0s表达率为（73．55±5．15）％，较正常对照组显著增加[（5．95±1．45）％，t=39．93，P〈0．01）]；5-ASA和4-ASA组大鼠肠组织iNOS表达率分别为（37．80±3．82）％和（42．27士3．52）％，均较实验对照组显著降低（t值分别=17．62和15．76，P值均d0．01）。实验对照组血清IL-8的平均浓度明显高于正常对照组（t=25．25，P〈0．01）；5-ASA和4-ASA组明显低于实验对照组（t值分别=12．31和11．57，P值均〈0．01）。实验对照组血PMN凋亡率明显低于正常对照组（t=11．48，P〈0．01）；5-ASA和4-ASA组凋亡率明显高于实验对照组（t值分别=7．51和10．47，P值均〈0．01）。结论4-ASA灌肠对实验性结肠炎大鼠具有显著的治疗作用，其治疗机制可能与降低PMN的趋化与激活、上调血PMN凋亡率、减少肠组织局部损伤因子有关。

Objective To investigate the effects of 4 aminosalieylic acid （4-ASA） on rats with 2,4,6-trinitrobenzenesulfonie acid-induced colitis in order to understand its mechanisms in treatment of inflammatory bowel disease （IBD）. Methods Thirty SD rats were given 2,4,6 trinitrobenzenesulfonic acid to induce colitis and were divided into model group, 5-ASA （200 mg/kg） treated group and 4- ASA （200 mg/kg） treated group with 10 each. Another 10 rats were severed as normal control. Seven days later,all animals were sacraficed for estimation of colonic tissues. The iNOS and serum level of interleukin （IL）-8 were detected by immunohistochemistry and enzyme linked immunosorbent assay （ELISA）,respectively. And the apoptosis of polymorphonuclear neutrophil （PMN） was examined by flow cytometry. Results In comparison with model group, the body weight was increased in rats treated with 4-ASA （t= 14.09, P〈0.01）, whereas the macroscopic and histological scores and MPO activity were decreased （t= 7.87,18.37,6.66 and 19.60, respectively, all P values 〈0.01）, which were similar to 5-ASA treated group （all P values 〉0. 05）. The expression of tissue iNOS was 73.55 ±5.15 % in model group, which was higher than that in control group [ （5.95 ± 1.45）% t= 39.93,P〈0.01）1, but was lower than that in 5-ASA treated group [（37.80± 3.82）%,t = 17.62, P〈0.01 ] and 4-ASA treated group [ （42.27±3.52）%, t = 15.76, P〈0.01]. The serum level of IL- 8 in model group was significantly higher than that in 5-ASA treated group and 4-ASA treated group （P%0.01）. The apoptosis of PMN in model group was lower than that in control group （t=11. 48, P%0.01）, but higher than that in 5-ASA treated group （t=7.51 ,P〈0. 01） and 4 ASA treated group （t= 10.47, P〈0.01）. Conclusions The efficacy of 4-ASA in treatment of IBD may be related to the mechanisms of reducing the migration and the activities of PMN, up-regulating PMN apoptosis and scavenging reactive oxygen radicals produced by PMN.