Abstract: Objective　To investigate the alleviation of Nippostrongylus brasiliensis infection on dextran sulfate sodium salt (DSS)⁃induced ulcerative colitis in mice, and to explore the underlying mechanism. Methods　Thirty male C57BL/6J mice of the SPF grade, each weighing approximately 25 g, were randomly divided into three groups, including the blank control group (NC group), DSS modeling group (DSS group), and N. brasiliensis treatment group (Nb + DSS group), of 10 mice in each group. Mice in the DSS group were orally administered with 3.5% DSS daily since day 1 (D0) for 6 successive days, and given normal drinking water since D6, and animals in the Nb + DSS group were subcutaneously injected with the third⁃stage larvae of N. brasiliensis at a dose of 500 larvae per mice 5 days prior to D0, followed by oral administration with 3.5% DSS daily since D0 for 6 successive days and normal drinking water since D6, while mice in the NC group were given normal drinking water. Mouse body weight and stool were observed and the disease activity index (DAI) was scored in each group during the study period. All mice were sacrificed on D9. The mouse colon length was measured, and mouse colon specimens were subjected to hematoxylin⁃eosin (HE) staining and histopathological scoring. In addition, the mRNA and protein expression of interleukin (IL)⁃1β and IL⁃10 was quantified in mouse colon specimens using quantitative fluorescent real⁃time PCR (qPCR) assay and enzyme⁃linked immunosorbent assay (ELISA), and the mRNA and protein expression of mucosal repair⁃associated molecules zonula occludens⁃1 (ZO⁃1), mucin 2 (MUC2) and claudin⁃1 was detected in mouse colon specimens using qPCR assay and immunofluorescence assay. Results　The mice body weights, DAI scores and colon lengths were (26.26 ± 1.93), (22.39 ± 1.65), (25.00 ± 1.58) g (F = 8.06, P < 0.01); (1.89 ± 0.34), (0.47 ± 0.39), 0 points (F = 57.61, P < 0.000 1); and (42.50 ± 5.75), (56.20 ± 5.96) mm and (61.17 ± 7.88) mm (F = 13.72, P < 0.001) in the NC, DSS and Nb + DSS groups on D9, respectively, and elevated mouse body weight (P < 0.05), reduced DAI score (P < 0.000 1) and increased colon length (P < 0.01) were observed in the Nb + DSS group relative to the DSS group on D9. Pathological examinations showed that the colonic crypts were relatively intact and the inflammatory cell infiltration was lower in the mouse colon specimens in the Nb + DSS group than in DSS the group. There was a significant difference in the histopathological scores of mouse colon specimens among the NC group (0 point), the DSS group [(2.00 ± 1.22) points] and the Nb + DSS group [(0.20 ± 0.45) points] (F = 10.71, P < 0.01), respectively, and the histopathological score of mouse colon specimens was significantly higher in the DSS group than in the NC and Nb + DSS groups (both P values < 0.01). qPCR assay quantified that the relative IL⁃10 and IL⁃1β mRNA expression was 1.25 ± 0.08, 0.44 ± 0.14 and 1.30 ± 0.45 (F = 10.66, P < 0.01), and 0.22 ± 0.13, 1.14 ± 0.31 and 0.41 ± 0.19 (F = 16.89, P < 0.001) in mouse colon specimens in the NC, DSS and Nb + DSS groups, respectively, and higher IL⁃10 mRNA expression and lower IL⁃1β mRNA expression were found in mouse colon specimens in the Nb + DSS group than in the DSS group (both P values < 0.01). The relative MUC2, claudin⁃1 and ZO⁃1 mRNA expression was 0.87 ± 0.25, 0.34 ± 0.26 and 4.21 ± 0.55 (F = 121.60, P < 0.000 1), 1.05 ± 0.41, 0.16 ± 0.09 and 0.22 ± 0.11 (F = 14.00, P < 0.01), and 1.03 ± 0.10, 0.60 ± 0.11 and 1.64 ± 0.28 (F = 32.16, P < 0.000 1) in mouse colon specimens in the NC, DSS and Nb + DSS groups, respectively, and significantly higher MUC2 and ZO⁃1 mRNA expression was quantified in mouse colon specimens in the Nb + DSS group than in the DSS group (both P values < 0.05). The mean fluorescence intensities of ZO⁃1 and claudin⁃1 were 17.18 ± 2.08, 12.38 ± 1.21 and 18.06 ± 2.59 (F = 8.95, P < 0.01) and 13.50 ± 1.63, 9.66 ± 2.03 and 13.61 ± 0.97 (F = 6.96, P < 0.05) in mouse colon specimens in the NC, DSS and Nb + DSS groups, respectively, and the mean fluorescence intensities of ZO⁃1 and claudin⁃1 were significantly greater in mouse colon specimens in the Nb + DSS group than in the DSS group (both P values < 0.05).  Conclusion　N. brasiliensis infection may remarkably alleviate DSS⁃induced ulcerative colitis in mice through promoting expression of anti⁃inflammatory cytokines, inhibiting expression of pro⁃inflammatory cytokines and facilitating mucosal repair in colon tissues.

Key words: ippostrongylus brasiliensis, Anti?inflammatory effect, Mucosal repair, Ulcerative colitis, Inflammatory bowel disease, Mouse

摘要： 目的　观察巴西日圆线虫（Nipponstrongylus brasiliensis，Nb）感染对葡聚糖硫酸钠盐（dextran sulphate sodium salt，DSS）诱导的小鼠溃疡性结肠炎的缓解作用，并初步探讨其作用机制。方法　将30只SPF级雄性C57BL/6J小鼠（体质量约25 g）随机分为3组，分别为空白对照组（NC组）、DSS造模组（DSS组）以及巴西日圆线虫干预组（Nb + DSS组），每组10只。DSS组小鼠于实验第1天（D0）开始每日口服3.5% DSS，至D6换为正常饮用水；在DSS组小鼠口服3.5% DSS 5 d前，Nb + DSS组小鼠按500条/鼠的剂量经皮下注射Nb Ⅲ期幼虫进行感染，并于D0开始每日口服3.5% DSS溶液，至D6换为正常饮用水；NC组小鼠给予正常饮用水。实验期间，观察各组小鼠体质量及粪便性状，并进行疾病活动指数（disease activity index，DAI）评分。各组小鼠均于D9剖杀，取结肠测量长度；收集小鼠结肠组织样本，采用苏木精⁃伊红染色观察组织病理学变化，进行病理学评分。采用实时荧光定量PCR（quantitative fluorescent real⁃time PCR，qPCR）法及酶联免疫吸附试验（enzyme⁃linked immunosorbent assay，ELISA）检测小鼠结肠组织白细胞介素（interleukin，IL）⁃1β、IL⁃10 mRNA和蛋白表达水平；采用qPCR和免疫荧光法检测小鼠结肠黏膜修复相关分子闭锁小带蛋白1（zonula occludens⁃1，ZO⁃1）、黏蛋白2（mucin⁃2，MUC2）、封闭蛋白1（claudin⁃1）mRNA和蛋白表达水平。结果　D9时，NC组、DSS组、Nb＋DSS组小鼠体质量分别为（26.26 ± 1.93）、（22.39 ± 1.65）、（25.00 ± 1.58） g（F = 8.06，P < 0.01），3组小鼠DAI评分分别为（1.89 ± 0.34）、（0.47 ± 0.39）、0分（F = 57.61，P < 0.000 1），结肠长度分别为（42.50 ± 5.75）、（56.20 ± 5.96）、（61.17 ± 7.88） mm（F = 13.72，P < 0.001）；两两比较结果显示，D9时，Nb + DSS组小鼠体质量较DSS组显著增加（P < 0.05）、DAI评分显著降低（P < 0.000 1）、结肠长度显著增加（P < 0.01）。病理学观察结果显示，与DSS组相比，Nb + DSS组小鼠结肠隐窝相对完整、炎性细胞浸润水平较低。NC组、DSS组、Nb + DSS组小鼠结肠组织病理学评分分别为0、（2.00 ± 1.22）、（0.20 ± 0.45）分，差异有统计学意义（F = 10.71，P < 0.01）；两两比较结果显示，DSS组小鼠结肠组织病理学评分高于NC组和Nb + DSS组（P均< 0.01）。NC组、DSS组、Nb + DSS组小鼠结肠组织IL⁃10 mRNA相对表达水平分别为1.25 ± 0.08、0.44 ± 0.14、1.30 ± 0.45（F = 10.66、P < 0.01），IL⁃1β mRNA相对表达水平分别为0.22 ± 0.13、1.14 ± 0.31、0.41 ± 0.19（F = 16.89，P < 0.001），差异均有统计学意义；与DSS组相比，Nb + DSS组小鼠结肠组织IL⁃10 mRNA相对表达水平显著升高（P < 0.01）、IL⁃1β mRNA表达水平显著下降（P < 0.01）。黏膜修复相关分子检测结果显示，NC组、DSS组、Nb + DSS组MUC2 mRNA相对表达水平分别为0.87 ± 0.25、0.34 ± 0.26、4.21 ± 0.55（F = 121.60，P < 0.000 1），claudin⁃1 mRNA相对表达水平分别为1.05 ± 0.41、0.16 ± 0.09、0.22 ± 0.11（F = 14.00，P < 0.01），ZO⁃1 mRNA相对表达水平分别为1.03 ± 0.10、0.60 ± 0.11、1.64 ± 0.28（F = 32.16，P < 0.000 1），差异均有统计学意义；相较于DSS组，Nb + DSS组小鼠结肠组织中MUC2、ZO⁃1 mRNA水平显著升高（P均< 0.05）。NC组、DSS组、Nb + DSS组小鼠结肠组织中ZO⁃1平均荧光强度分别为17.18 ± 2.08、12.38 ± 1.21、18.06 ± 2.59（F = 8.947，P < 0.01），claudin⁃1平均荧光强度分别为13.50 ± 1.63、9.66 ± 2.03、13.61 ± 0.97（F = 6.96，P < 0.05），差异均有统计学意义；相较于DSS组，Nb + DSS组小鼠结肠组织中ZO⁃1、claudin⁃1平均荧光强度显著升高（P均< 0.05）。结论　Nb感染可显著缓解DSS诱导的小鼠溃疡性结肠炎，其作用机制可能与感染促进小鼠结肠组织中抗炎细胞因子表达、抑制促炎细胞因子表达以及促进黏膜修复等有关。

关键词: 巴西日圆线虫, 抗炎作用, 黏膜修复, 溃疡性结肠炎, 炎症性肠病, 小鼠