目的 观察小鼠骨髓来源的未成熟树突状细胞(imDC)对2,4,6-三硝基苯磺酸(TNBS)诱导出的克罗恩病小鼠模型的治疗作用,探讨其缓解小鼠克罗恩病的作用机制.方法 40只小鼠随机分为4组,每组10只,分为对照组、TNBS组、imDC组和成熟树突状细胞(mDC)组,除对照组予以乙醇灌肠外,其余3组都予以乙醇-TNBS混合溶液灌肠造模,在灌肠前后各1d经尾静脉注射磷酸盐缓冲液(PBS)或者细胞进行干预,对照组和TNBS组尾静脉注射PBS,imDC组尾静脉注射imDC,mDC组尾静脉注射mDC进行干预.每天观察小鼠的一般状况,于造模第7天后处死小鼠,分别行疾病活动指数(DAI)、结肠病理组织学(TDI)和结肠大体形态损伤指数(CMDI)评分,用酶联免疫吸附试验(ELISA)法分析小鼠结肠组织白细胞介素-10 (IL-10)、转化生子因子-β(TGF-β)的表达,Western blot分析小鼠结肠组织叉头状/翅状螺旋蛋白3(Foxp3)蛋白的表达.结果 TNBS组的DAI、CMDI、TDI的评分别为(5.420 ±0.326)、(4.923±0.226)、(7.200±0.851)分,较对照组明显升高,差异有统计学意义(P<0.01),结肠组织抗炎因子IL-10和TGF-β分别为(6.103±1.140)、(21.164 ±3.252) ng/L,调节性T细胞(Treg)转录因子Foxp3的相对表达量为0.439 2±0.045 9,较对照组明显降低,差异有统计学意义(P<0.01);imDC能显著降低小鼠DAI、CMDI、TDI评分,下调抗炎因子IL-10和TGF-β和转录因子Foxp3的水平;mDC则显著升高小鼠DAI、CMDI、TDI评分,降低抗炎因子IL-10和TGF-β和转录因子Foxp3蛋白的水平,差异均有统计学意义(P<0.05).结论 本实验成功地制作出了小鼠克罗恩病模型,并证明mDC能明显缓解小鼠克罗恩病模型的进程,而诱导Treg的产生可能是其缓解炎症的机制之一.
Objective To investigate the effect of immature dentritic cell(imDC) on the Crohn's mice models induced by 2,4,6-trinitrobenzene sulfonic acid (TNBS).Methods The mouse were randomly divided into 4 groups:normal group,TNBS group,immature dentritic cell (imDC)group,mature dentritic cell (mDC) group.The other three gourps were administered 50% ethanol/TNBS vain anus except control group,Each group was injected saline or cells via tail vein before and after administration of anus enema.Control group and TNBS group were injected saline.imDC group was injected imDC.mDC-group was injected mDC.Then we observe the general situation of murine.After one week,the disease activity index (DAI),tissue damage index (TDI) and colon macroscopic damage index (CMDI) were evaluated,the colon level of interleukin-10 (IL-10),transforming growth factor-β (TGF-β) determined by enzyme-linked immunosorbent assay (ELISA).The level of colon forkhead/winged helix protein 3 (Foxp3) protein was determined by Western blotting.Results The scores of DAI,CMDI,TDI in TNBS group were 5.420 ± 0.326,4.923 ± 0.226,7.200 ± 0.851,incresed significantly compared with control group (P<0.01).And the colon tissue level of IL-10,TGF-β protein were (6.103 ± 1.140),(21.164 ±3.252) ng/L,the relatively expression of protein was 0.439 2 ±0.045 9,decresed significantly compared with crotrol group (P < 0.01).ImDC can significantly decrease the scores of DAI,CMDI,TDI and increse the colon tissue level of IL-10,TGF-β,Foxp3 protein (P <0.05).But the mDC increased the scores of DAI,CMDI,TDI and decrese the colon tissue level of IL-10,TGF-β,Foxp3 protein (P < 0.05).Conclusion We succsefully induce Crohn' s mice models by TNBS.And obseve the imDC can significantly inhabit intestinal inflammation in Crohn' s mice models and the mechanism of immune tolerance induced by imDC might be inhibit T lymphocytes responsiveness by regulatory T cells.