新生儿粪便中分离的粪肠球菌(Enterococcus faecalis)RZ18提高肠道黏膜屏障完整性

Promoting Intestinal Mucosal Barrier Integrity with Enterococcus faecalis RZ18 Isolated from Neonatal Feces

本实验旨在研究新生儿粪便样品中乳酸菌对肠道黏膜屏障完整性的促进作用.从健康新生儿粪便中分离出15株乳酸菌.对这些乳酸菌进行耐酸、耐胆盐和细胞黏附能力的实验,分析其定植能力.再通过荧光素酶报告基因分析实验、实时荧光定量PCR(Real-time PCR)、酶联免疫吸附测定检测各菌发酵上清液对HT-29细胞中黏蛋白2(mucin2,MUC2)表达的影响,结果显示粪肠球菌(Enterococcus faecalis)RZ18发酵上清液对MUC2启动子活性、转录和翻译水平均有增强作用.并通过Real-time PCR、免疫印迹实验检测E.faecalis RZ18发酵上清液对肠道黏膜屏障中闭锁蛋白、封闭蛋白1、闭锁小带蛋白1表达的影响,发现该菌可以促进这些紧密连接蛋白的表达.最后用牛津杯法研究发现该菌对藤黄微球菌、金黄色葡萄球菌、单增李斯特菌和大肠杆菌等多种致病菌生长具有抑制作用.上述结果说明,新生儿肠道早期定植的E.faecalis可能对新生儿肠道黏膜屏障完整性的发育具有促进作用,有望应用于炎症性肠病的预防与治疗.

This study was to investigate the effect of lactic acid bacteria on intestinal mucosal barrier integrity in neonatal feces.Fifteen strains of lactic acid bacteria were isolated from healthy neonatal feces.The intestinal colonization ability of the lactic acid bacteria such as their resistance to acid or bile salts and their adhesive ability to intestinal cells were evaluated.The effect of fermentation broth supernatant on the level of mucin 2 in HT-29 cells were tested with the luciferase report gene analysis experiment,Real-time PCR and enzyme linked immunosorbent assay.The results showed that the fermentation supernatant of Enterococcus faecalis RZ18 increased the promoter activity of MUC2 gene and upregulated the transcription and translation level of MUC2.The fermentation broth supernatant of RZ18 also enhanced the expression of tight junction proteins including occludin,claudin1 and zonula occluden-1,confirmed by Real-time PCR and Western blot test.The Oxford cup method showed that RZ18 can inhibit the growth of a variety of pathogenic bacteria,including Micrococcus luteus,Staphylococcus aureus,Listeria monocytogenes and Escherichia coli.The results of the research indicate that Enterococcus faecalis colonized in the early stage of neonatal intestinal tract could promote the development of neonatal intestinal mucosal barrier,which can be expected to be applied to the prevention and treatment of inflammatory bowel disease.