摘要
目的探讨脂多糖(LPS)通过TLRs信号途径进一步促进人外周血单核细胞来源树突状细胞(DC)成熟的机理。方法从外周血分离单核细胞,加入rhGM-CSF及rhIL-4,在培养的第6d,实验组加入LPS刺激24h,对照组不加。分别提取细胞总RNA,行半定量RT-PCR,产物进行琼脂糖凝胶电泳分析。激光扫描共聚焦显微镜检测DC内NF-κB表达情况。结果单核细胞经GM-CSF及IL-4诱导7d后的DC表达较高水平TLR2、3、4。LPS刺激24h后,TLR2、3、4表达下降,TLR4几乎测不到。激光扫描共聚焦显微镜检测到,未刺激组DC胞浆染色阳性,胞核为阴性。LPS刺激组的细胞核染色为强阳性,胞浆染色为阳性。结论DC在受LPS刺激前后TLRs的表达有显著变化,NF-κB也从胞浆移位至核内。这标志着经LPS刺激后DC在功能上进一步成熟。为下一步研究DC的功能奠定了基础。
Objective To find out how lipopolysaccharide(LPS) induces dendritic cells(DC) to be more mature through Toll-like receptors(TLRs) signal pathway. Methods Monocytes were isolated from heparinized whole blood. 7 days later the monocytes were induced into dendritic cells by adding rhGM-CSF and rhIL-4. Total cellular RNA of unstimulated and stimulated DC were extracted by LPS 24 h later. Semi-quantitative RT-PCR and agarose gel electrophoresis were conducted to analyze the PCR products. The expression of NF-κB was detected by laser scanning confocal microscopy (LSCM). Results Expressions of TLR2,3,4 were significantly higher in monocytes-derived DC induced for 7 days, the plasm of DC was stained positive and the nucleus of DC was stained negative. After being stimulated by LPS for 24 h, the expressions of TLR2,3,4 decreased, and that of TLR4 was almost undetectable; the nucleus of DC was stained strong positive, the plasm of DC was stained positive. Conclusion The expressions of TLR2,3,4 on DC change significantly after being stimulated by LPS. And NF-κB shifts from the plasm to the nucleus. This signifies that LPS induces DC to be more mature in function.
出处
《四川大学学报（医学版）》
CAS
CSCD
北大核心
2005年第4期513-515,共3页
Journal of Sichuan University(Medical Sciences)
基金
高等学校博士学科点专项科研基金(编号:0040105102006)资助
