摘要
目的利用多种细胞因子及肿瘤细胞裂解物刺激诱导树突状细胞(dendritic cell,DC)成熟,并利用基因芯片技术对其信号传导基因表达谱进行研究。方法外周血单个核细胞在体外用细胞因子GM-CSF(100 ng/m l),IL-4(1 000 U/m l)诱导下获取DC,以乳腺癌MCF-7细胞反复冻融的裂解物冲击DC,通过形态学和FCM检测来确定DC。再用基因芯片技术检测其细胞因子及信号传导相关基因表达的变化。结果DC在细胞因子的诱导下,形态上伸出许多伪足样突起,在摄取抗原后可见内吞样小泡。FCM检测呈现成熟DC的标志:CD40、CD83、CD80、CD86、HLA-DR等高表达。芯片扫描发现16种信号传导因子发生5倍以上的改变,其中包括钙调磷酸酶结合蛋白1、TGF-β3受体、FKBP-雷帕霉素相关蛋白、小分子细胞因子A(Cys-Cys)、人T细胞特异蛋白(RANTES)等。结论通过体外细胞因子诱导及抗原冲击的成熟DC不但在形态和表型上存在明显差异,而且在功能和信号传导方面也存在显著变化。
Objective To investigate the maturation of dendritic cells （DCs） and the expression profile of its signal transduction genes by DNA microarray. Methods DCs derived fi＇om peripheral blood monocytes （PBMCs） were isolated and cultured in vitro with rhGM-CSF （100 ng/ml） and rhIL-4 （1000U/ml）. Then they were pulsed with lysates from breast cancer cells MCF-7 that had been lysed by 5 freeze-thaw cycles. The mature DCs were identified by morphological examination and flow cytometry （FCM）. Finally, the DCs＇ expression profile of cytokine and signal transduction genes were examined by the DNA microarray under different circumstances. Results On the 7th day the mature DCs sent out many pseudopods. Some cndosomal vesicles could be seen after tumor antigen pulsing. FCM showed that the surface marker of the mature DCs were up-regulated as compared with immature DCs, ineluding CD40, CD83, CD80, CD86, HLA-DR. DNA microarray demonstrated that 16 genes, including calcineurin binding protein 1 , human transforming growth fac- tor-beta type Ⅲ receptor, FKBP-rapamycin associated protein （FRAP） , small inducible cytokine subfamily A （Cys-Cys）, C5a anaphylatoxin receptor ere experienced change by more than 5 folds. Conclusion The cytokines-driven DCs in vitro not only experience changes in morphology and phenotype, but also go through changes in function and signal transduction.
出处
《医学分子生物学杂志》
CAS
CSCD
2006年第2期96-100,共5页
Journal of Medical Molecular Biology
基金
天津市科委重大攻关项目(No.023111511-8)~~
关键词
树突状细胞
细胞因子
信号传导
基因芯片
dendritic cells
cytokine
signal transduction genes
DNA microarray
