粪菌移植干预治疗溃疡性结肠炎的免疫学机制研究

背景溃疡性结肠炎被世界卫生组织列为现代难治疾病之一。目前西医对其治疗尚存在诸多不足之处，研究显示粪菌移植（FMT）对其有一定的疗效，但机制尚不明确。目的采用FMT治疗小鼠溃疡性结肠炎模型，验证FMT的疗效和可能的作用机制。方法2019年12月至2020年4月，采用随机数字表法将60只小鼠分为正常对照组（Control组）、溃疡性结肠炎模型组（Model组）、溃疡性结肠炎模型+粪菌移植治疗组（Model+FMT组）和溃疡性结肠炎模型+5-氨基水杨酸（5-ASA）治疗组（Model+5-ASA组），各15只。Control组不作任何干预处理；Model组制备小鼠溃疡性结肠炎模型；Model+FMT组于造模成功后，给予制备的粪菌液0.2 ml/次灌肠；Model+5-ASA组于造模成功后，给予0.019 5 g/ml 5-ASA灌肠。通过透射电镜观察肠组织超微结构变化，流式细胞检测血液辅助性T细胞（Th）-17、Th-1、Th-2、Treg细胞含量变化，酶联免疫吸附试验（ELISA）检测血清干扰素γ（IFN-γ）、白介素（IL）-2、IL-17、IL-4、IL-10、转化生长因子β（TGF-β）水平变化。结果肠组织透射电镜超微结构显示Model组造模成功；Model+FMT组与Model+5-ASA组微绒毛较为致密，形态正常，杯状细胞数目较多，线粒体轻微肿胀，粗面内质网病变不明显。Model+FMT组Th17细胞含量高于Control组、低于Model组，Model+5-ASA组Th17细胞含量低于Control组、Model组、Model+FMT组；Model+FMT组、Model+5-ASA组Th1细胞含量均分别低于Control组、Model组；Model+FMT组Th2细胞含量低于Control组、高于Model组，Model+5-ASA组Th2细胞含量低于Control组、高于Model组和Model+FMT组；Model+FMT组、Model+5-ASA组Treg细胞含量均分别低于Control组、高于Model组（P<0.05）。Model+5-ASA组IFN-γ细胞含量低于Model组；Model+FMT组、Model+5-ASA组IL-2细胞含量低于Model组；Model+FMT组、Model+5-ASA组IL-17细胞含量均分别低于Control组和Model组，Model+5-ASA组IL-17细胞含量低于Model+FMT组；Model+FMT组IL-4细胞含量低于Control组、高于Model组，Model+5-ASA组IL-4细胞含量高于Model组；Model+FMT组IL-10细胞含量高于Control组和Model组，Model+5-ASA组IL-10细胞含量高于Model组；Model+FMT组、Model+5-ASA组TGF-β细胞含量均分别低于Control组、高于Model组（P<0.05）。结论FMT可改善小鼠溃疡性结肠炎症状，推测其可能是通过调节Th1/Th2细胞平衡、Th17/Treg细胞比例达到治疗的目的。

Immunological Mechanism of Faecal Bacterial Transplantation in the Intervention of Ulcerative Colitis

Background Ulcerative colitis is listed as one of the modern refractory diseases by the World Health Organization.At present,Western medicine still has many shortcomings in its treatment,and studies have shown that fecal bacterial transplantation(FMT)has a certain effect on it,but the mechanism is not clear.Objective FMT was used to treat mouse ulcerative colitis model to verify the efficacy and possible mechanism of FMT.Methods From December 2019 to April 2020,60 mice were divided into normal control group(Control group),ulcerative colitis model group(Model group),ulcerative colitis model+fecal bacteria transplantation treatment group(Model+FMT group)and ulcerative colitis model+5-aminosalicylic acid(5-ASA)treatment group(Model+5-ASA group)by random number table method,each of 15 mice.Control group did not make any intervention;Model group prepared mouse ulcerative colitis model;after successful modeling,the Model+FMT group was given 0.2 ml of fecal bacteria solution per enema;after successful modeling,the Model+5-ASA group was given 0.0195 g/ml 5-ASA enema.The ultrastructural changes of intestinal tissue through transmission electron microscope,flow cytometric detection of blood helper T cell(Th)-17,Th-1,Th-2,Treg cell content changes,and enzyme-linked immunosorbent assay(ELISA)were observed to detect serum interferonγ(IFN-γ),interleukin(IL)-2,IL-17,IL-4,IL-10,transforming growth factorβ(TGF-β)level changes.Results Intestinal tissue transmission electron microscopy ultrastructure showed that the Model group was successfully modeled;the microvilli in the Model+FMT group and Model+5-ASA group were denser,with normal morphology,more goblet cells,slight swelling of mitochondria,and insignificant rough endoplasmic reticulum lesions.The Th17 cell content of the Model+FMT group was higher than that of the Control group and lower than that of the Model group;the Th17 cell content of the Model+5-ASA group was lower than that of Control group,Model group and Model+FMT group.The Th1 cell content of Model+FMT group and Model+5-ASA group were lower than those of Control group and Model group,respectively;Th2 cell content of Model+FMT group was lower than that of Control group and higher than that of Model group,and Th2 cell content of Model+5-ASA group was lower than that of Control group and higher than that of Model group and Model+FMT group.Treg cell content in Model+FMT group and Model+5-ASA group were lower than that of Control group and higher than that of Model group(P<0.05).IFN-γcell content in Model+5-ASA group was lower than that of Model group.IL-2 cell content in Model+FMT group and Model+5-ASA group was lower than that of Model group;the IL-17 cell content of Model+FMT group and Model+5-ASA group were lower than those of Control group and Model group,respectively.The IL-17 cell content of Model+5-ASA group was lower than that of Model+FMT group;the IL-4 cell content of Model+FMT group was lower than that of Control group and higher than that of Model group.The IL-4 cell content in the Model+5-ASA group was higher than that in the Model group;the IL-10 cell content in the Model+FMT group was higher than that in the Control group and the Model group,and the IL-10 cell content in the Model+5-ASA group was higher than that in the Model group;the content of TGF-βcells in the Model+FMT group and Model+5-ASA group were lower than those in the Control group and higher than those in the Model group(P<0.05).Conclusion FMT can improve the symptoms of ulcerative colitis in mice.It is speculated that it may be achieved by adjusting the balance of Th1/Th2 cells and the ratio of Th17/Treg cells to achieve the purpose of treatment.