Surface expression of toll-like receptor 9 is upregulated on intestinal epithelial cells in response to pathogenic bacterial DNA
Surface expression of toll-like receptor 9 is upregulated on intestinal epithelial cells in response to pathogenic bacterial DNA
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DOI：
10.1128/iai.01662-06
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发表时间：
2007-05-01
影响因子：
3.1
通讯作者：
Madsen, Karen L.
中科院分区：
文献类型：
--
作者：
Ewaschuk, Julia B.;Backer, Jody L.;Madsen, Karen L.
结肠上皮细胞不断暴露于肠腔中高水平的细菌 DNA，必须识别病原体并做出适当反应，同时保持对非致病性共生菌株的耐受性。细菌 DNA 被 Toll 样受体 9 (TLR9) 识别。本研究的目的是研究基础条件下结肠上皮细胞中 TLR9 的表达和定位以及对细菌 DNA 的反应。 HT-29 细胞暴露于来自各种共生微生物和病原微生物菌株的 DNA。通过实时逆转录 PCR 测定 TLR9 mRNA 表达，通过酶联免疫吸附测定测定白介素 8 (IL-8) 分泌。在 HT-29 细胞中通过流式细胞术以及在 HT-29 细胞和小鼠结肠组织中通过免疫荧光测定 TLR9 的定位。免疫荧光和流式细胞术分析表明，基础条件下HT-29细胞内和表面均存在TLR9表达。将细胞暴露于来自沙门氏菌和大肠杆菌致病菌株的 DNA 会导致 TLR9 mRNA 表达显着增加。肠道沙门氏菌血清型都柏林 DNA 增加了表面 TLR9 蛋白和 IL-8 的分泌。对于短双歧杆菌的反应，TLR9 的 mRNA 水平或定位没有变化。氯喹不会阻断肠沙门氏菌都柏林 DNA 血清型 IL-8 的分泌。 TLR9 在野生型小鼠的结肠顶端表面表达，但在无菌小鼠中不表达。这些结果表明肠上皮细胞识别病原细菌DNA并通过增加TLR9的表面定位和表达来做出反应，这表明对病原DNA的上皮炎症反应至少部分是由TLR9表达增加介导的。
Colonic epithelial cells are constantly exposed to high levels of bacterial DNA in the intestinal lumen and must recognize and respond appropriately to pathogens, while they maintain a tolerance to nonpathogenic commensal bacterial strains. Bacterial DNA is recognized by Toll-like receptor 9 (TLR9). The aim of this study was to investigate TLR9 expression and localization in colonic epithelial cells under basal conditions and in response to bacterial DNA. HT-29 cells were exposed to DNA from various strains of commensal and pathogenic microbes. TLR9 mRNA expression was determined by real-time reverse transcription-PCR, and interleukin-8 (IL-8) secretion was measured by an enzyme-linked immunosorbent assay. Localization of TLR9 was determined by flow cytometry in HT-29 cells and by immunofluorescence in HT-29 cells and mouse colonic tissue. Immunofluorescence and flow cytometric analyses demonstrated that there was intracellular and surface expression of TLR9 in HT-29 cells under basal conditions. Exposure of cells to DNA from pathogenic strains of Salmonella and Escherichia coli resulted in a significant increase in TLR9 mRNA expression. Salmonella enterica serovar Dublin DNA increased surface TLR9 protein and IL-8 secretion. There was no change in mRNA levels or localization of TLR9 in response to Bifidobacterium breve. Chloroquine did not block IL-8 secretion in response to S. enterica serovar Dublin DNA. TLR9 was expressed on the colonic apical surface in wild-type mice but not in germfree mice. These results demonstrate that intestinal epithelial cells recognize pathogenic bacterial DNA and respond by increasing surface localization and expression of TLR9, suggesting that the epithelial inflammatory response to pathogenic DNA is mediated at least in part by increased TLR9 expression.