PDF(Pubmed)
Abstract：
The human intestinal mucus layer protects against pathogenic microorganisms and harmful substances, whereas it also provides an important colonization niche for mutualistic microbes. The main functional components of mucus are heavily glycosylated proteins, called mucins. Mucins can be cleaved and utilized by intestinal microbes. The mechanisms between intestinal microbes and the regulation of mucin glycosylation are still poorly understood. In this study, in vitro mucus was produced by HT29-MTX-E12 cells under Semi-Wet interface with Mechanical Stimulation. Cells were exposed to pasteurized nonpathogenic bacteria Akkermansia muciniphila, Ruminococcus gnavus, and Bacteroides fragilis to evaluate influence on glycosylation patterns. Following an optimized protocol, O- and N-glycans were efficiently and reproducibly released, identified, and semiquantified using MALDI-TOF-MS and PGC-LC-MS/MS. Exposure of cells to bacteria demonstrated increased diversity of sialylated O-glycans and increased abundance of high mannose N-glycans in in vitro produced mucus. Furthermore, changes in glycan ratios were observed. It is speculated that bacterial components interact with the enzymatic processes in glycan production and that pasteurized bacteria influence glycosyltransferases or genes involved. These results highlight the influence of pasteurized bacteria on glycosylation patterns, stress the intrinsic relationship between glycosylation and microbiota, and show the potential of using in vitro produced mucus to study glycosylation behavior.
摘要：
人体肠道粘液层可防止病原微生物和有害物质，而它也为共生微生物提供了重要的定植生态位。粘液的主要功能成分是高度糖基化的蛋白质，叫做粘蛋白。粘蛋白可以被肠道微生物裂解和利用。肠道微生物和粘蛋白糖基化调节之间的机制仍然知之甚少。在这项研究中,体外粘液由HT29-MTX-E12细胞在半湿界面下通过机械刺激产生。将细胞暴露于巴氏灭菌的非致病性细菌Akkermansiamuciniphila，gnavusRuminococcus,和脆弱拟杆菌评估对糖基化模式的影响。遵循优化的协议，O-和N-聚糖被有效和可重复地释放，已识别,并使用MALDI-TOF-MS和PGC-LC-MS/MS进行半定量。细胞暴露于细菌表明唾液酸化的O-聚糖的多样性增加,并且在体外产生的粘液中高甘露糖N-聚糖的丰度增加。此外，观察到聚糖比率的变化。据推测,细菌组分与聚糖生产中的酶促过程相互作用,并且巴氏灭菌的细菌影响所涉及的糖基转移酶或基因。这些结果突出了巴氏杀菌细菌对糖基化模式的影响，强调糖基化和微生物群之间的内在关系，并显示了使用体外产生的粘液研究糖基化行为的潜力。
