具有抗炎和促干细胞活性双效应评价功能的新型结肠炎小鼠模型的构建及应用

Construction and application of a novel mouse model of colitis with the function evaluating effects of anti-inflammatory and pro-stemness

目的建立具有抗炎和促肠道干细胞活性双效应评价功能的新型结肠炎小鼠模型。方法将免疫功能缺陷的Il10^(-/-)小鼠和具有干细胞示踪功能的Lgr5-EGFP小鼠进行杂交,得到Il10^(-/-);Lgr5-EGFP小鼠。在SPF环境中,用3%的硫酸葡聚糖钠(dextran sulfate sodium salt,DSS)诱导该基因型小鼠形成慢性结肠炎,给予美沙拉嗪(50或150 mg·kg^(-1)·d^(-1))作为阳性药物治疗组。治疗期间对小鼠进行疾病活动指数评分并记录结果。7周后,对小鼠血清中的炎症因子进行ELISA检测,对结肠组织的炎症因子表达进行RT-qPCR检测,对结肠组织切片进行免疫荧光观察,比较Lgr5-EGFP+细胞的数量。结果美沙拉嗪作为轻、中度溃疡性结肠炎(ulcerative colitis,UC)的一线用药,对此模型有一定的治疗效果:显著改善了小鼠慢性结肠炎的症状;抑制了IFN-γ、IL-17A、TNF-α和IL-12/23 p40等炎症因子的血清水平及其在结肠组织的表达;修复了Lgr5+干细胞群的丢失。结论具有炎症免疫和肠道干细胞活性双靶向效应评价功能的新型结肠炎小鼠模型构建成功,适用于筛选新型抗结肠炎药物。

Objective To establish a novel colitis model with the function evaluating effects of antiinflammatory and pro-stemness.Methods Il10^(-/-);Lgr5-EGFP mice were obtained by crossing immunodeficient Il10^(-/-)mice with Lgr5-EGFP mice with function of tracing ISCs.Mice of this genotype were induced with 3%dextran sulfate sodium salt(DSS)to develop chronic colitis in SPF facilities and orally given mesalazine for 50 or 150 mg·kg^(-1)·day^(-1) as positive drug.Disease activity index was evaluated and recorded.After 8 weeks,the ELISA was performed to detect the level of inflammatory cytokines in mouse serum,and RT-qPCR was performed for inflammatory cytokines expression in colon tissue.Immunofluorescence was observed on colon tissue sections to compare the number of Lgr5-EGFP+cells between different groups.Results The results showed that mesalazine,as the first-line drug for mild to moderate ulcerative colitis(UC),had some degree of therapeutic effects in this model,mesalazine significantly alleviated the chronic colitis;inhibited the serum levels and colonic expression of inflammatory cytokines such as IFN-γ,IL-17A,TNF-αand IL-12/23 p40;and restored the loss of Lgr5+ISCs.Conclusion A novel mouse model of colitis with the function evaluating effects of anti-inflammatory and pro-stemness was successfully constructed,which is applicable for the new drug discovery for colitis.