Clark Robin T, Hope Andrew, Lopez-Fraga Marta, Schiller Neal, Lo David D
Division of Biomedical Sciences, University of California, Riverside, Riverside, California 92521, USA.
Clin Vaccine Immunol. 2009 Mar;16(3):397-407. doi: 10.1128/CVI.00210-08. Epub 2009 Jan 7.
Bacterial pathogens use virulence strategies to invade epithelial barriers, but active processes of epithelial cells may also contribute to the endocytosis of microbial particles. To focus on the latter, we studied the uptake of fixed and fluorescently labeled bacterial particles in intestinal and bronchoepithelial cell cultures and found it to be enhanced in Caco-2BBe and NCI-H292 cells after treatment with tumor necrosis factor alpha and an agonist antibody against the lymphotoxin beta receptor. Confocal fluorescence microscopy, flow cytometry, and transmission electron microscopy revealed that Staphylococcus aureus and Yersinia enterocolitica were readily endocytosed, although there was scant uptake of Shigella sonnei, Salmonella enterica serovar Typhimurium, and Klebsiella pneumoniae particles. Endocytosed Staphylococcus was often associated with cytoplasmic claudin-4 vesicles; this was not found for Yersinia, suggesting that cytokine treatment upregulated two distinct endocytosis pathways. Interestingly, when Staphylococcus and Yersinia were coincubated with epithelial monolayers, the cells were unlikely to take up Yersinia unless they had also endocytosed large numbers of Staphylococcus particles, although the two bacteria were apparently processed in distinct compartments. Cytokine treatment induced an upregulation and redistribution of beta1 integrin to the apical surface of NCI-H292 cells; consistent with this effect, treatment with anti-beta1 integrin antibody blocked uptake of both Yersinia and Staphylococcus in NCI-H292 and Caco-2BBe cells. Our results suggest that capture of bacterial particles by mucosal epithelial cells is selective and that different endocytic mechanisms are enhanced by proinflammatory cytokines.
细菌病原体利用毒力策略侵入上皮屏障，但上皮细胞的主动过程也可能有助于微生物颗粒的内吞作用。为了聚焦于后者，我们研究了在肠道和支气管上皮细胞培养物中固定并荧光标记的细菌颗粒的摄取情况，发现用肿瘤坏死因子α和抗淋巴毒素β受体激动剂抗体处理后，Caco-2BBe和NCI-H292细胞中的摄取增强。共聚焦荧光显微镜、流式细胞术和透射电子显微镜显示，金黄色葡萄球菌和小肠结肠炎耶尔森菌很容易被内吞，尽管宋内志贺菌、鼠伤寒沙门氏菌和肺炎克雷伯菌颗粒的摄取很少。内吞的金黄色葡萄球菌常与细胞质中的紧密连接蛋白4囊泡相关；而耶尔森菌则未发现这种情况，这表明细胞因子处理上调了两种不同的内吞途径。有趣的是，当金黄色葡萄球菌和耶尔森菌与上皮单层细胞共同孵育时，除非细胞也内吞了大量金黄色葡萄球菌颗粒，否则细胞不太可能摄取耶尔森菌，尽管这两种细菌显然在不同的区室中被处理。细胞因子处理诱导β1整合素上调并重新分布到NCI-H292细胞的顶端表面；与此效应一致，用抗β1整合素抗体处理可阻断NCI-H292和Caco-2BBe细胞中耶尔森菌和金黄色葡萄球菌的摄取。我们的结果表明，黏膜上皮细胞对细菌颗粒的捕获具有选择性，并且促炎细胞因子可增强不同的内吞机制。
