INSERM U913, Place Alexis Ricordeau, Nantes Cedex, France.
Gut. 2011 Apr;60(4):473-84. doi: 10.1136/gut.2010.229237. Epub 2010 Dec 7.
Enteric glial cells (EGCs) are important regulators of intestinal epithelial barrier (IEB) functions. EGC-derived S-nitrosoglutathione (GSNO) has been shown to regulate IEB permeability. Whether EGCs and GSNO protect the IEB during infectious insult by pathogens such as Shigella flexneri is not known.
S flexneri effects were characterised using in vitro coculture models of Caco-2 cells and EGCs (or GSNO), ex vivo human colonic mucosa, and in vivo ligated rabbit intestinal loops. The effect of EGCs on S flexneri-induced changes in the invasion area and the inflammatory response were analysed by combining immunohistochemical, ELISA and PCR methods. Expression of small G-proteins was analysed by western blot. Expression of ZO-1 and localisation of bacteria were analysed by fluorescence microscopy.
EGCs significantly reduced barrier lesions and inflammatory response induced by S flexneri in Caco-2 monolayers. The EGC-mediated effects were reproduced by GSNO, but not by reduced glutathione, and pharmacological inhibition of pathways involved in GSNO synthesis reduced EGC protecting effects. Furthermore, expression of Cdc42 and phospho-PAK in Caco-2 monolayers was significantly reduced in the presence of EGCs or GSNO. In addition, changes in ZO-1 expression and distribution induced by S flexneri were prevented by EGCs and GSNO. Finally, GSNO reduced S flexneri-induced lesions of the IEB in human mucosal colonic explants and in a rabbit model of shigellosis.
These results highlight a major protective function of EGCs and GSNO in the IEB against S flexneri attack. Consequently, this study lays the scientific basis for using GSNO to reduce barrier susceptibility to infectious or inflammatory challenge.
肠胶质细胞（EGCs）是调节肠道上皮屏障（IEB）功能的重要调节因子。已证实 EGC 衍生的 S-亚硝基谷胱甘肽（GSNO）可调节 IEB 的通透性。然而，EGC 和 GSNO 是否在诸如福氏志贺菌等病原体感染时保护 IEB 尚不清楚。
采用 Caco-2 细胞和 EGC（或 GSNO）体外共培养模型、离体人结肠黏膜和体内结扎兔肠袢，研究福氏志贺菌的作用。通过结合免疫组织化学、ELISA 和 PCR 方法分析 EGC 对福氏志贺菌诱导的侵袭面积和炎症反应变化的影响。通过 Western blot 分析小 G 蛋白的表达。通过荧光显微镜分析 ZO-1 的表达和细菌的定位。
EGC 显著减轻了 Caco-2 单层中由福氏志贺菌引起的屏障损伤和炎症反应。GSNO 可再现 EGC 介导的作用，但还原型谷胱甘肽则不能，而 GSNO 合成途径的药理学抑制降低了 EGC 的保护作用。此外，在存在 EGC 或 GSNO 的情况下，Caco-2 单层中 Cdc42 和磷酸化 PAK 的表达明显减少。此外，EGC 和 GSNO 可预防福氏志贺菌引起的 ZO-1 表达和分布变化。最后，GSNO 减轻了人黏膜结肠标本和兔志贺菌病模型中 IEB 由福氏志贺菌引起的损伤。
这些结果突出了 EGC 和 GSNO 在 IEB 抵抗福氏志贺菌攻击中的主要保护功能。因此，本研究为利用 GSNO 降低屏障对感染或炎症挑战的易感性奠定了科学基础。
