摘要目的 在肿瘤坏死因子相关凋亡诱导配体基因敲除(TRAIL-/-)小鼠中诱导实验性结肠炎模型,分析肿瘤坏死因子相关凋亡诱导配体( TRAIL)缺乏对小鼠结肠炎症和肠道菌群组分的影响.方法 选取C57BL/6 品系的TRAIL-/-小鼠和野生型小鼠各24 只,分为TRAIL-/-对照组(8 只)和TRAIL-/-结肠炎组(16只),以及野生型对照组(8只)和野生型结肠炎组(16只).两组结肠炎小鼠均连续7 d口服3.5%葡聚糖硫酸钠饮用水,诱导实验性结肠炎模型,从临床表现和组织病理学方面评估结肠炎严重程度.收集小鼠结肠组织标本,采用高通量16S rDNA测序技术检测结肠菌群组分,再应用USEARCH软件和R语言软件分析TRAIL-/-对照组、TRAIL-/-结肠炎组、野生型对照组和野生型结肠炎组小鼠结肠菌群组分的差异.组间比较采用独立样本t检验和秩和检验.结果 造模开始后,野生型结肠炎组小鼠和 TRAIL-/-结肠炎组小鼠的结肠炎疾病活动指数( DAI)都随时间推移逐渐增高, TRAIL-/-结肠炎组小鼠比野生型结肠炎组小鼠更早出现体质量下降、腹泻和便血.在造模后第7天, TRAIL-/-结肠炎组小鼠体质量下降(28.98 ±2.84)%,野生型结肠炎组小鼠体质量下降(17.87 ± 3.70)%,两组间比较差异有统计学意义(t＝9.53,P?0.01);TRAIL-/-结肠炎组小鼠的结肠长度短于野生型结肠炎组小鼠[(4.63 ±0.28) cm比(6.02 ±0.41) cm],差异有统计学意义(t＝11.20,P?0.01);TRAIL-/-结肠炎组小鼠的DAI高于野生型结肠炎组小鼠(3.00 ±0.00比2.32 ±0.05),差异有统计学意义(t＝54.40,P?0.01);TRAIL-/-结肠炎组小鼠的组织学评分高于野生型结肠炎组小鼠[(6.19 ± 0.25)分比(3.87 ±0.22)分],差异有统计学意义(t＝27.87,P?0.01).与野生型结肠炎组小鼠相比, TRAIL-/-结肠炎组小鼠的结肠黏膜上皮受损、隐窝结构破坏和炎症细胞浸润更加明显.与野生型结肠炎组小鼠相比,TRAIL-/-结肠炎组小鼠的肠道菌群具有更高的α多样性:在科分类水平,TRAIL-/-结肠炎组小鼠肠道菌群中的脱铁杆菌科、疣微菌科、理研菌科、F16和Paraprevotellaceae的相对丰度均明显增高,肠球菌科相对丰度明显降低[(19.839 ±19.991)%比(7.224 ±11.241)%、(3.564 ±2.543)%比(2.861 ±3.821)%、(0.123 ±0.066)%比(0.068 ±0.049)%、(0.032 ±0.033)%比(0.006 ±0.011)%、(0.153 ±0.098)%比(0.062 ±0.054)%、(0.013 ±0.027)%比(0.054 ±0.121)%,U＝51、69、53、35、49、69,P分别?0.01、0.05];在属分类水平,TRAIL-/-结肠炎组小鼠肠道菌群中的颤螺旋菌属、黏菌属和噬细胞菌属的相对丰度均明显增高,而肠球菌属相对丰度明显降低[(2.363 ±2.147)%比(1.813 ± 2.847)%、(19.839 ±19.991)%比(7.223 ±11.241)%、(0.104 ±0.153)%比( 0.046 ±0.069)%、(0.076 ±0.049)%比(0.135 ±0.074)%,U＝70、51、66、65,P分别?0.05、0.01].结论 TRAIL缺乏能加重葡聚糖硫酸钠诱导的结肠炎,并引起小鼠结肠菌群的α多样性增高.

abstractsObjective To investigate the influence of tumor necrosis factor-related apoptosis-inducing ligant (TRAIL) deficiency on mice colitis and the gut microbiota composition by inclding the expermental colitis model in tumor necrosis factor-related apoptosis-inducing ligand gene knockout ( TRAIL-/-) mice. Methods C57BL/6 TRAIL-/-mice and wild type (WT) mice were selected and assigned into TRAIL-/-control group (eight mice), TRAIL-/-colitis group (16 mice), WT control group (eight mice) and WT colitis group (16 mice).The mice of two colitis groups were oral administrated with 3.5% dextran sulphate sodium (DSS) in drinking water for seven consecutive days to induce experimental colitis model .The severity of colitis was evaluated by clinical appearance and histopathological examination .The colonic tissue samples of mice were collected and microbiota profile was analyzed by 16S rDNA sequencing method.USEARCH software and R language were used to analyze the difference of gut microbiota among TRAIL-/-control group, TRAIL-/-colitis group, WT control group and WT colitis group .T test and Mann-Whitney U test were used for statistical analysis . Results After modeling, the disease activity index (DAI) of WT colitis mice and TRAIL-/-colitis mice both gradually increased over time .Furthermore, compared with colitis mice, TRAIL-/-colitis mice developed body weight loss, diarrhea and hemafecia earlier .On the seventh day after modeling , the percentage of body weight loss of TRAIL-/-colitis mice and WT colitis mice was (28.98 ±2.84)%and (17.87 ±3.70)%, respectively; and the difference was statistically significant (t＝9.53, P?0.01).The length of colon of TRAIL-/-colitis mice was shorter than that of WT colitis mice ((4.63 ±0.28) cm vs.(6.02 ±0.41) cm), and the difference was statistically significant (t＝11.20, P?0.01).The DAI of TRAIL-/-colitis mice was higher than that of WT colitis mice (3.00 ±0.00 vs.2.32 ±0.05), and the difference was statistically significant (t ＝54.40, P? 0.01).The histological score of TRAIL-/-colitis mice was higher than that of WT colitis mice (6.19 ±0.25 vs. 3.87 ±0.22), and the difference was statistically significant (t ＝27.87, P?0.01).Under the microscope, colonic mucosal epithelial injury , crypt structure destruction and inflammatory cell infiltration were more obvious in TRAIL-/-colitis mice than in WT colitis mice.The alpha diversity of colonic flora was more significant in TRAIL-/-colitis group compared with that of WT colitis group .At the family level, the relative richness of Deferribacteraceae, Ruminococcaceae, Rikenellaceae, F16 and Paraprevotellaceae significantly increased in TRAIL-/-colitis group, but the relative richness of Enterococcaceae obviously reduced ((19.839 ±19.991)%vs. (7.224 ±11.241)%, (3.564 ±2.543)% vs.(2.861 ±3.821)%, (0.123 ±0.066)% vs.(0.068 ± 0.049)%, (0.032 ±0.033)% vs.(0.006 ±0.011)%, (0.153 ±0.098)% vs.(0.062 ±0.054)% and (0.013 ±0.027)%vs.(0.054 ±0.121)%, respectively; U＝51, 69, 53, 35, 49 and 69, respectively; P? 0.01 and 0.05, respectively).In addition, at the genus level the relative richness of Oscillospira, Mucispirillum and Cytophaga in TRAIL-/-colitis group remarkably elevated , and the relative richness of Enterococcus significantly decreased ((2.363 ±2.147)% vs.(1.813 ±2.847)%, (19.839 ±19.991)% vs.(7.223 ± 11.241)%, (0.104 ±0.153)%vs.(0.046 ±0.069)% and (0.076 ±0.049)% vs.(0.135 ±0.074)%, respectively; U＝70, 51, 66 and 65, respectively; P ?0.05 and 0.01, respectively).Conclusion TRAIL deficiency aggravate DSS-induced colitis, and increase the alpha diversity of colonic microbiota in colitis mice .