Nature揭示是什么防止你的肠道菌入侵肠道
2016/04/12
在4月6日Nature中发表的研究表明LY6 /PLAUR家族的一个成员—— Lypd8蛋白,可以在小鼠中防止鞭毛微生物入侵结肠上皮细胞。


结肠上皮细胞内外都由厚的粘液层覆盖。内粘液层有自由的共生微生物,这有助于维持肠道的稳定态。但为什么共生的微生物不会侵入肠道造成炎症还是很让人困惑的。在小肠,预防细菌入侵上皮细胞的重要分子,如潘氏细胞衍生的抗微生物肽和Regenerating islet-derived 3 (RegIII)家族蛋白已经确定。虽然在大肠内有粘液层提供对大量菌群的物理障碍,但将细菌和结肠上皮细胞分开的机制在本研究刊登之前还未完全阐明。

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Lypd8蛋白特异性表达在大肠上皮

在4月6日Nature中发表的研究表明LY6/PLAUR家族的一个成员—— Lypd8蛋白,可以在小鼠中防止鞭毛微生物入侵结肠上皮细胞。研究人员在大肠的公共数据库里寻找特定表达的基因,发现Lypd8选择性地表达在小鼠肠胃道的细胞,特别是盲肠和大肠。Lypd8在无菌小鼠和无特定病原小鼠中表达水平是相似的。这提示了Lypd8的表达不受微生物依赖诱导的炎症信号诱导。他们观察到高水平的Lypd8 mRNA表达在大肠肠腺最上层的表皮细胞中。

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Lypd8蛋白和鞭毛细菌的关系

没有Lypd8时,细菌出现在内部粘液层,很多有鞭毛的细菌入侵上皮细胞。在Lypd8−/ −的小鼠中肠道细菌贯穿内粘膜层,进一步进入大肠隐窝。用通用引物扩增野生型和Lypd8 −/ −小鼠肠道内容物和细菌16S rRNA,细菌组成并没有怎么改变。为了做比较,Lypd8 −/ −小鼠结肠组织的细菌DNA也被扩增检测。入侵Lypd8−/ −小鼠结肠上皮细胞的细菌种属被特异性引物所鉴定。结果表明在Lypd8 −/ −小鼠结肠组织中,特别是变形杆菌Proteus、幽门螺杆菌Helicobacter 和大肠杆菌 Escherichia增多。这些都是革兰氏阴性杆菌具多鞭毛,侵入内粘液层。

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实验证明Lypd8蛋白和鞭毛菌结合限制细菌运动

重组的Lypd8和野生型小鼠的粪便样品共培养揭示了Lypd8抑制细菌侵入上皮层的机制。通过流式细胞仪和荧光显微镜的观察发现Lypd8优先结合大肠杆菌和幽门螺杆菌等鞭毛细菌。Lypd8和体外纯培养的Proteus mirabilis结合,但不和无鞭毛的细菌结合。Lypd8没有显示出对P. mirabilis的杀菌活性。对Lypd8和共培养的 P. mirabilis进行免疫金扫描电镜观察显示Lypd8特异性结合细菌鞭毛。

Lypd8 −/ −小鼠对葡聚糖硫酸钠诱导的肠道炎症反应高度灵敏。革兰阴性抗生素消除鞭毛细菌可以恢复粘液层内的细菌自由状态,改善葡聚糖硫酸钠引起的肠炎。

将体外培养的P.mirabilis或者大肠杆菌加到有重组Lypd8的半固体培养基中发现:Lypd8和鞭毛结合抑制鞭毛细菌的运动。因此Lypd8介导肠细菌和结肠上皮细胞的分离并维持肠内平衡,Lypd8可能也有预防肠道感染的作用。

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  • Lypd8 promotes the segregation of flagellated microbiota and colonic epithelia

    Colonic epithelial cells are covered by thick inner and outer mucus layers. The inner mucus layer is free of commensal microbiota, which contributes to the maintenance of gut homeostasis. In the small intestine, molecules critical for prevention of bacterial invasion into epithelia such as Paneth-cell-derived anti-microbial peptides and regenerating islet-derived 3 (RegIII) family proteins have been identified. Although there are mucus layers providing physical barriers against the large number of microbiota present in the large intestine, the mechanisms that separate bacteria and colonic epithelia are not fully elucidated. Here we show that Ly6/PLAUR domain containing 8 (Lypd8) protein prevents flagellated microbiota invading the colonic epithelia in mice. Lypd8, selectively expressed in epithelial cells at the uppermost layer of the large intestinal gland, was secreted into the lumen and bound flagellated bacteria including Proteus mirabilis. In the absence of Lypd8, bacteria were present in the inner mucus layer and many flagellated bacteria invaded epithelia. Lypd8−/− mice were highly sensitive to intestinal inflammation induced by dextran sulfate sodium (DSS). Antibiotic elimination of Gram-negative flagellated bacteria restored the bacterial-free state of the inner mucus layer and ameliorated DSS-induced intestinal inflammation in Lypd8−/− mice. Lypd8 bound to flagella and suppressed motility of flagellated bacteria. Thus, Lypd8 mediates segregation of intestinal bacteria and epithelial cells in the colon to preserve intestinal homeostasis.

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