TRAIL基因敲除对实验性结肠炎小鼠Th17细胞的影响

Impact of TRAIL deficiency on Th17 cells in the mice experimental colitis induced by dextran sodium sulphate

目的在实验性结肠炎小鼠中探讨TRAIL基因敲除(TRAIL^-/-)对Th17细胞数目及活性的影响。方法C57BL/6小鼠给予3.5%葡聚糖硫酸钠(DSS)自由饮用7 d诱导实验性结肠炎模型。小鼠随机分为4组,每组6只:野生型(WT)对照组、WT结肠炎组、TRAIL^-/-对照组和TRAIL-/-结肠炎组。从临床表现和组织病理学上对结肠炎组的严重程度进行评估。取外周血单个核细胞(PBMC)和肠系膜淋巴结(MLN),采用流式细胞术检测Th17细胞数目,并采用定量聚合酶链反应技术检测白细胞介素(IL)-17A和视黄酸相关孤儿受体(ROR)-γt的表达水平。结果WT结肠炎组PBMC和MLN中CD4+IL-17A+ Th17细胞比例(0.29±0.07比0.08±0.03,P<0.01;1.20±0.36比0.40±0.11,P<0.05),以及IL-17A和ROR-γt的mRNA表达水平均高于WT对照组(IL-17A:2.43±0.87比0.37±0.19,5.03±1.77比1.05±0.48,均P<0.05;ROR-γt:2.49±0.48比0.93±0.47,23.75±7.60比1.31±0.90,均P<0.05)。TRAIL^-/-结肠炎组较WT结肠炎组发生更为严重的结肠炎:疾病活动指数增高,结肠缩短及炎性细胞浸润增多。并且TRAIL^-/-结肠炎组的PBMC和MLN中CD4+IL-17A+ Th17细胞比例(0.57±0.22比0.29±0.07,P<0.001;2.92±0.98比1.20±0.36,P<0.000 1),以及IL-17A和ROR-γt的mRNA表达水平均高于WT结肠炎组(IL-17A:4.10±1.96比2.43±0.87,15.88±2.86比5.03±1.77,均P<0.05;ROR-γt:4.05±0.62比2.49±0.48,69.61±10.48比23.75±7.60,均P<0.05)。结论TRAIL缺乏会导致PBMC和MLN中Th17细胞数目和活性增高,并加重DSS诱导的实验性结肠炎。

Objective To investigate the impact of TNF-related apoptosis-inducing ligand (TRAIL) gene knock-out (TRAIL^-/-) on Th17 cells in the mice colitis induced by dextran sulphate sodium (DSS). Methods Mice were randomly assigned to 4 subgroups: wild type (WT), TRAIL^-/-, WT colitis and TRAIL^-/-colitis (n=6/group). Colitis was induced by oral administration of 3.5% DSS for 7 consecutive days. The severity of colitis in each mouse was scored both clinically and histopathologically. Flow cytometry was performed to assess Th17 cell population in peripheral blood mononuclear cells (PBMC) and mesenteric lymph nodes (MLN). The expression levels of Th17 cell markers interleukin (IL)-17A and retinoic acid-related orphan receptor (ROR)-γt in PBMC and MLN were also examined using a quantitative polymerase chain reaction method. Results Compared with WT group, WT colitis group displayed elevated CD4+IL-17A+ Th17 cells (0.29±0.07 vs 0.08±0.03, 1.20±0.36 vs 0.40±0.11, both P<0.05) and enhanced mRNA expression of IL-17A and ROR-γt in PBMC and MLN (IL-17A: 2.43±0.87 vs 0.37±0.19, 5.03±1.77 vs 1.05±0.48, both P<0.05;ROR-γt: 2.49±0.48 vs 0.93±0.47, 23.75±7.60 vs 1.31±0.90, both P<0.05). After the DSS administration, TRAIL^-/- group developed more severe colitis than WT group, mainly manifesting higher disease activity index, reduction of colon length and increased infiltration of inflammatory cells. In addition, TRAIL^-/- colitis group exhibited increased proportion of CD4+ IL-17A+ Th17 cells (0.57±0.22 vs 0.29±0.07, P<0.001;2.92±0.98 vs 1.20±0.36, P<0.000 1) as well as enhanced mRNA expression of IL-17A and ROR-γt in PBMC and MLN when compared with WT colitis group (IL-17A: 4.10±1.96 vs 2.43±0.87, 15.88±2.86 vs 5.03±1.77, both P<0.05;ROR-γt: 4.05±0.62 vs 2.49±0.48, 69.61±10.48 vs 23.75±7.60, both P<0.05). Conclusions TRAIL deficiency not only promots the number and activity of Th17 cells in PBMC and in MLN, but also aggravats DSS-induced colitis in mice.