肠道菌群影响无菌小鼠对卵清蛋白的敏感性

Influence of intestinal microbiota on sensitivity in germ-free mice to ovalbumin

目的通过观察过敏和无过敏反应小鼠肠道菌群在无菌小鼠肠道内定植后对卵清蛋白(OVA)刺激的反应,探究肠道菌群在食物过敏中的作用。方法采集用卵清蛋白成功诱导出的过敏和无过敏反应SPF级小鼠的粪便,分别移植给无菌小鼠(各10只),为过敏组(FA组)和无过敏反应组(NR组)。粪菌移植后第3周,两组分别随机抽取7只再用OVA激发为实验组(FA-o组、NR-o组),余3只为生理盐水对照组(FA-c组、NR-c组)。用变性梯度凝胶电泳(DGGE)技术评价粪菌移植后肠道菌群在肠道内的定植情况,比较FA组和NR组肠道菌群的差别。观察两组再用OVA激发后的临床表现,HE染色观察小鼠空肠组织形态学改变。结果 (1)粪菌移植后第3周,与供体小鼠相比,FA组与NR组小鼠肠道菌群的DGGE条带多数一致,而FA组与NR组肠道菌群比较,DGGE条带有明显不同,聚类分析和主成分分析(principal component analysis,PCA)两组均能很好分离,测序结果也表明无过敏反应组中厚壁菌门的细菌占优势,过敏组中变形菌门的细菌占优势;(2)移植粪菌的无菌鼠再次用OVA激发时,FA-o组有临床表现,NR-o组无明显临床表现;HE染色可见FA-o组较NR-o组小鼠空肠绒毛上皮细胞局灶性坏死、脱落,炎性细胞浸润明显严重。结论肠道菌群不同使无菌小鼠对OVA的敏感性不同,说明肠道菌群的变化可能是食物过敏的始动因素,肠道菌群可传递宿主对食物过敏的敏感性。

Objective To explore the effects of intestinal microbiota on food allergy through observing the reaction of germ-free mice to ovalbumin（ OVA） stimulation after transplantation with fecal microbiota to allergic and non-allergic reaction SPF mice. Methods The intestinal microbiota of the allergic and nonallergic reaction SPF mice that were successfully induced by OVA were respectively transplanted into the germfree mice to establish the allergic reaction group（ FA group） and non-allergic reaction group（ NR group）,with 10 mice in each group. At the 3rd week after transplantation,7 mice in each group were randomly selected for OVA treatment and defined as FA-o group and NR-o group,and the rest 3 mice in each group were treated with sodium chloride physiological solution and defined as FA-c group and NR-c group. The colonization of the intestinal microbiota in the germ-free mice after transplantation was detected by denaturing gradient gel electrophoresis（ DGGE）,and the differences of microbiota composition between FA group and NR group were compared. The clinical symptoms of the 2 groups after OVA stimulation were observed,and then the morphological changes of intestinal tissue were examined by HE staining. Results At the 3rd week after fecal microbiota transplantation,the DGGE bands were almost the same in the FA group and NR group as their own donor＇s,but the DGGE bands of the FA group and NR group were significantly different. The 2groups showed a clear separation in cluster analysis and principal component analysis（ PCA）. The sequencing results also showed that Firmicutes was predominate in the NR group,whereas Proteobacteria was dominant bacteria in the FA group. After OVA stimulation,the FA-o group had more obvious clinical symptoms than the NR-o group. HE staining showed that the FA-o group had more serious pathological changes than the NR-o group,such as inflammatory cell infiltration,focal necrosis and shedding of the epithelial cells in the intestinal villi. Conclusion The intestinal microbiota can affect the sensitivity to OVA in the transplanted mice,and the sensitivity to food allergy can be transmitted via the intestinal microbiota.