健康人单核细胞来源树突状细胞Toll样受体的表达模式分析  

Expression of Toll-like receptor s of human dendritic cells

作　　者：韩亚萍[1] 李军[1] 孔练花[1] 刘源[1] 陈念[1] 万玉峰[1] 黄祖瑚[1] 

机构地区：[1]南京医科大学第一附属医院感染科,南京210029

出　　处：《临床检验杂志》2007年第6期427-429,共3页Chinese Journal of Clinical Laboratory Science

基　　金：江苏省医学重点学科工程资助(135-07);江苏省卫生厅医学科技发展基金(H200311);江苏省"333工程"培养资金资助项目(2002)。

摘　　要：目的体外诱导健康人外周血单核细胞来源的树突状细胞(dendritic cell,DC),动态定量分析DC表达Toll样受体(TLRs)的种类,探讨TLRs的表达水平与DC成熟的相关性。方法用羟乙基淀粉(HES)分离健康人外周血单个核细胞(PB-MC),经重组人粒细胞-巨噬细胞集落刺激因子(rhGM-CSF)、重组人白细胞介素4(rhIL-4)诱导培养DC,于培养5d加或不加肿瘤坏死因子-α(TNF-α)和干扰素-α(IFN-α),诱导其为成熟与未成熟两种类型的细胞。分别于3、5、7d用特异性单克隆抗体(单抗)标记未成熟DC(immature DC,imDC)和7d成熟DC(mature DC,mDC),在流式细胞仪(FACS)上检测,用CellQuest软件分析细胞表面共刺激分子(B7-1、B7-2)和MHCⅡ类分子以及细胞内、外TLR3、TLR4、TLR7、TLR8、TLR9的表达百分率。结果健康人TLR3在imDC细胞膜表面的表达高于细胞内,并随着培养时间的延长有所下降;TLR4则主要表达在imDC细胞膜外,并伴随着干预性成熟而下降;TLR7在imDC细胞内的表达高于细胞表面,并随着imDC生长时间延长而逐渐增加,DC成熟时表达下降;TLR8和TLR9细胞内外均有表达,但细胞内稍高于细胞外,随DC成熟而表达增加。结论健康人单核细胞来源的DC表面TLRs的表达具有多样性,TLRs在DC的不同分化阶段表达模式不同,这种差异可能是DC通过TLRs识别病原体相关分子模式(pathogen associated molecular pattern,PAMPs)的基础,从而有效地调节或控制固有免疫与获得性免疫。Objective To dynamically, quantitatively analyze the varieties of Toll-like receptors （TLRs） of dentritic cell （ DC ） derived from peripheral blood mononuclear cells （PMBC） of healthy people in vitro, and to investigate the relationship between the protein ex- pression level of TLRs and maturity of DC. Methods PMBC was separated by HES. Mature and immature DCs were induced with adding or not adding tumor necrosis factor-or （TNF-α） and interferon-or （IFN-α） correspondingly on the fifth day after being cultured with the recombinant human granulocyte-macrophage colony-stimulating factor （rhGM-CSF） and recombinant human interleukin （rhIL）-4. The immture dendritic cell （imDC） on the 3rd, 5th and 7th day, and the mature dendritic cell （mDC） on the 7th day were labeled by specific monoclonal antibody respectively and tested by flow cytometry. The percentage of costimulatory molecules B7-1, B7- 2 and MHC Ⅱ on cell surface and TLR3, TLR4, TLR7, TLR8 and TLR9 intra- and extra-membrane were then analyzed. Results In healthy people the expression of the TLR3 in membrane surface of imDC was higher than that inside cells and decreased with the prolonged culture time. TLR4 was mainly expressed on extra-membrane of imDC and decreased during mature with intervention. The ex- pression of TLR7 inside imDC was higher than that on membrane surface and increased with imDC aging but decreased when DC matured. Both TLR8 and TLR9 were expressed inside and outside cells with a little higher expression inside cells compared with that outside cells, and increased when DC matured. Conclusion The expression of TLRs on DC derived from PMBC of healthy people showed different pattern in various stages of DC differentiation. The varieties may be the basis of recognizing pathogen associated molecular pattern which generally resides on surface of various pathogens, The various TLRs of DC effectively adjust and control the inherent and acquired immunity.

关 键 词：TOLL样受体 树突状细胞 抗原提呈细胞 固有免疫 

分 类 号：R446.6[医药卫生—诊断学]