Human intestinal microvascular endothelial cells express toll-like receptor 5: A binding partner for bacterial flagellin
Human intestinal microvascular endothelial cells express toll-like receptor 5: A binding partner for bacterial flagellin
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DOI：
10.4049/jimmunol.172.8.5056
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发表时间：
2004-04-15
影响因子：
4.4
通讯作者：
Kucharzik, T
中科院分区：
文献类型：
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作者：
Maaser, C;Heidemann, J;Kucharzik, T
细菌鞭毛蛋白最近被确定为Toll样受体5(TLR5)的配体。已知的人类特异表达TLR5的部位包括巨噬细胞以及胃和肠上皮。由于沙门氏菌感染肠上皮细胞导致鞭毛蛋白主动转运到微血管附近的上皮下室，我们假设人肠内皮细胞功能性地表达TLR5，从而使转位的鞭毛蛋白结合后能够产生积极的炎症反应。采用RT-PCR、免疫印迹分析和免疫荧光法检测TLR5在人巨、微血管内皮细胞中的表达。免疫组织化学方法检测TLR5在体内的内皮表达。用流式细胞仪定量检测内皮细胞对ICAM-1表达的调节，用内皮迁移试验评价白细胞在体外的迁移。利用体外共培养系统研究了活体沙门氏菌感染时内皮细胞与上皮细胞的相互作用。我们发现沙门氏菌感染的肠上皮细胞可诱导共培养的人内皮细胞表达ICAM-1。来自人皮肤(人真皮微血管内皮细胞1)和人结肠(人肠道微血管内皮细胞)的巨(HUVEC)和微血管内皮细胞(HUVEC)均高表达TLR5mRNA和蛋白。这些发现与体内正常人结肠微血管中TLR5的强烈免疫反应是平行的。此外，人真皮微血管内皮细胞与临床分离的大肠埃希菌和沙门氏菌的鞭毛蛋白孵育后，ICAM-1的表达显著上调，并促进了白细胞跨内皮细胞的迁移。这些结果表明，内皮表达的TLR5可能在针对细菌AGS的先天免疫反应中发挥以前未被认识到的作用。
Bacterial flagellin has recently been identified as a ligand for Toll-like receptor 5 (TLR5). Human sites known to specifically express TLR5 include macrophages and gastric and intestinal epithelium. Because infection of intestinal epithelial cells with Salmonella leads to an active transport of flagellin to the subepithelial compartment in proximity to microvessels, we hypothesized that human intestinal endothelial cells functionally express TLR5, thus enabling an active inflammatory response upon binding of translocated flagellin. Endothelial expression of TLR5 in human macro- and microvascular endothelial cells was examined by RT-PCR, immunoblot analysis, and immunofluorescence. Endothelial expression of TLR5 in vivo was verified by immunohistochemistry. Endothelial modulation of ICAM-1 expression was quantitated using flow cytometry, and leukocyte transmigration in vitro was assessed by an endothelial transmigration assay. Epithelial-endothelial cellular interactions upon infection with viable Salmonella were investigated using a coculture system in vitro. We found that Salmonella-infected intestinal epithelial cells induce endothelial ICAM-1 expression in cocultured human endothelial cells. Both macro- (HUVEC) and microvascular endothelial cells derived from human skin (human dermal microvascular endothelial cell 1) and human colon (human intestinal microvascular endothelial cells) were found to express high constitutive amounts of TLR5 mRNA and protein. These findings were paralleled by strong immunoreactivity for TLR5 of normal human colonic microvessels in vivo. Furthermore, incubation of human dermal microvascular endothelial cells with flagellin from clinical isolates of Escherichia and Salmonella strains led to a marked up-regulation of ICAM-1, as well as to an enhanced leukocyte transendothelial cell migration. These results suggest that endothelially expressed TLR5 might play a previously unrecognized role in the innate immune response toward bacterial Ags.