Impaired intestinal barrier structure and function have been validated as an important pathogenic process in type 2 diabetes mellitus (T2DM). Gut dysbiosis is thought to be the critical factor in diabetic intestinal pathogenesis. As the most abundant commensal bacteria, Faecalibacterium prausnitzii (F. prausnitzii) play important roles in gut homeostasis. The microbial anti‐inflammatory molecule (MAM), an F. prausnitzii metabolite, has anti‐inflammatory potential in inflammatory bowel disease (IBD). Thus, we aimed to explore the function and mechanism of MAM on the diabetic intestinal epithelium.
16S high‐throughput sequencing was used to analyze the gut microbiota of db/db mice (T2DM mouse model). We transfected a FLAG‐tagged MAM plasmid into human colonic cells to explore the protein‐protein interactions and observe cell monolayer permeability. For in vivo experiments, db/db mice were supplemented with recombinant His‐tagged MAM protein from E. coli BL21 (DE3).
The abundance of F. prausnitzii was downregulated in the gut microbiota of db/db mice. Immunoprecipitation (IP) and mass spectroscopy (MS) analyses revealed that MAM potentially interacts with proteins in the tight junction pathway, including zona occludens 1 (ZO‐1). FLAG‐tagged MAM plasmid transfection stabilized the cell permeability and increased ZO‐1 expression in NCM460, Caco2, and HT‐29 cells. The db/db mice supplemented with recombinant His‐tagged MAM protein showed restored intestinal barrier function and elevated ZO‐1 expression.
Highlights
Diabetic conditions induce compositional dysbiosis of the gut microbiota and impair gut barrier integrity.
Diabetic conditions markedly downregulate the abundance of Faecalibacterium prausnitzii in the gut.
Under diabetic conditions, microbial anti‐inflammatory molecules from Faecalibacterium prausnitzii restore the gut barrier and ZO‐1 expression possibly through the tight junction pathway.
肠道屏障结构和功能异常是 2 型糖尿病的重要病理改变之一。 而肠道菌群失调被认为是诱发糖尿病肠道病变的关键因素。 普拉梭菌作为肠道中最为常见的共生菌, 在维持肠道内环境稳态中扮演着重要的角色。前沿研究发现, 源于普拉梭菌的活性产物:微生物抗炎分子(MAM)对炎症性肠病具有炎症抑制潜能。因此, 我们在本研究中探索 MAM 在糖尿病肠道上皮中的功能及机制。
我们运用 16S 高通量测序分析 db/db 小鼠(2 型糖尿病小鼠模型)的肠道菌群。其次, 通过合成 FLAG 标记的 MAM 质粒, 并进行结肠上皮细胞转染, 以探索蛋白‐蛋白互作机制并观测细胞间通透性改变情况。在体内实验中, 我们构建起 E.coli BL21(DE3)细菌蛋白表达系统用于合成 His 标签重组 MAM 蛋白。运用所合成提纯的重组 MAM 蛋白对 db/db 小鼠进行肠道干预, 观察体内干预后肠道上皮屏障结构和功能改善情况。