Kayal S, Lilienbaum A, Poyart C, Memet S, Israel A, Berche P
Laboratoire de Microbiologie, INSERM U-411, Faculté de Médecine Necker, Paris, France.
Mol Microbiol. 1999 Mar;31(6):1709-22. doi: 10.1046/j.1365-2958.1999.01305.x.
The facultative intracellular bacterium Listeria monocytogenes is an invasive pathogen that crosses the vascular endothelium and disseminates to the placenta and the central nervous system. Its interaction with endothelial cells is crucial for the pathogenesis of listeriosis. By infecting in vitro human umbilical vein endothelial cells (HUVEC) with L. monocytogenes, we found that wild-type bacteria induced the expression of the adhesion molecules (ICAM-1 and E-selectin), chemokine secretion (IL-8 and monocyte chemotactic protein-1) and NF-kappa B nuclear translocation. The activation of HUVEC required viable bacteria and was abolished in prfA-deficient mutants of L. monocytogenes, suggesting that virulence genes are associated with endothelial cell activation. Using a genetic approach with mutants of virulence genes, we found that listeriolysin O (LLO)-deficient mutants inactivated in the hly gene did not induce HUVEC activation, as opposed to mutants inactivated in the other virulence genes. Adhesion molecule expression, chemokine secretion and NF-kappa B activation were fully restored by a strain of Listeria innocua transformed with the hly gene encoding LLO. The relevance in vivo of endothelial cell activation for listerial pathogenesis was investigated in transgenic mice carrying an NF-kappa B-responsive lacZ reporter gene. NF-kappa B activation was visualized by a strong lacZ expression in endothelial cells of capillaries of mice infected with a virulent haemolytic strain, but was not seen in those infected with a non-haemolytic isogenic mutant. Direct evidence that LLO is involved in NF-kappa B activation in transgenic mice was provided by injecting intravenously purified LLO, thus inducing stimulation of NF-kappa B in endothelial cells of blood capillaries. Our results demonstrate that functional listeriolysin O secreted by bacteria contributes as a potent inflammatory stimulus to inducing endothelial cell activation during the infectious process.
兼性胞内菌单核细胞增生李斯特菌是一种侵袭性病原菌，可穿越血管内皮并扩散至胎盘和中枢神经系统。其与内皮细胞的相互作用对于李斯特菌病的发病机制至关重要。通过用单核细胞增生李斯特菌体外感染人脐静脉内皮细胞（HUVEC），我们发现野生型细菌可诱导黏附分子（ICAM - 1和E - 选择素）的表达、趋化因子分泌（IL - 8和单核细胞趋化蛋白 - 1）以及NF - κB核转位。HUVEC的激活需要活菌，并且在单核细胞增生李斯特菌的prfA缺陷型突变体中被消除，这表明毒力基因与内皮细胞激活有关。使用毒力基因突变体的遗传学方法，我们发现hly基因失活的李斯特菌溶血素O（LLO）缺陷型突变体不会诱导HUVEC激活，这与其他毒力基因突变体失活的情况相反。用编码LLO的hly基因转化的无害李斯特菌菌株可完全恢复黏附分子表达、趋化因子分泌和NF - κB激活。在携带NF - κB反应性lacZ报告基因的转基因小鼠中研究了内皮细胞激活在李斯特菌发病机制中的体内相关性。在感染强毒溶血菌株的小鼠毛细血管内皮细胞中，通过强烈的lacZ表达可观察到NF - κB激活，但在感染非溶血同基因突变体的小鼠中未观察到。通过静脉注射纯化的LLO，从而诱导毛细血管内皮细胞中的NF - κB刺激，为LLO参与转基因小鼠中的NF - κB激活提供了直接证据。我们的结果表明，细菌分泌的功能性李斯特菌溶血素O作为一种强大的炎症刺激物，在感染过程中有助于诱导内皮细胞激活。
