人外周血树突状细胞的超微结构及其细胞化学特征

The cytologic ultrastructure and cytochemicalfeaturesof human dendritic cellsfrom peripheral blood

目的 观察体外富集的人外周血树突状细胞（ｄｅｎｄｒｉｔｉｃ ｃｅｌｌｓ，ＤＣ） 的光镜形态学，细胞超微结构及细胞化学特征。方法 取健康人新鲜外周抗凝血，采用淋巴细胞分离液分离单个核细胞进行培养，经细胞因子（ｒｈＧＭＣＳＦ，ｒｈＩＬ４ ，ｒｈＴＮＦα） 联合刺激培养９～１２ 天，富集含ＤＣ的细胞群，光镜形态学分辨ＤＣ形态及含量，用抗人血ＤＣ表面特异单克隆抗体（ｍＡｂ Ⅻ） 确认ＤＣ的细胞化学特征。结果 体外培养富集的人外周血ＤＣ群的ＤＣ含量达７０ ％ ～８０ ％ 。透射电镜观察表明，细胞因子诱导的人外周血ＤＣ群可见到两类ＤＣ亚群：一类可能来自ＤＣ前体细胞，其细胞表面有大量不规则树枝状突起（ 粗细不等） ，细胞核不规则，核仁小，细胞化学染色［髓过氧化物酶（ＰＯＸ） 、非特异性酯酶（ＮＳＥ）］ 均为阴性；另一类是由单核细胞分化来的ＤＣ，细胞形态不规则，细胞表面粗糙，有大量皱褶及较粗大树枝状突起，胞核常呈现肾形或马蹄形，核内异染色质边集。结论 细胞因子诱导的人外周血ＤＣ群存在多相性。

Objective To observe the cytomorphological,cellular ultrastructure and cytochemicalfeatures ofthe dendritic cell(DC) enriched populationsin vitro. Methods Fresh peripheralbloodfrom healthy volunteers was collectedand mononuclearcells(PBMC) wereisolatedonlymphocyteseparatingsolution.The PBMCwere cul turedin mediumscontainingrhGM CSF,rhIL 4 and rhTNF α9～12 daysto generate DC enriched populations,the morphologicalfeatures and numbersof DCwereidentified and counted respectively by light microscope (LM) and confirmed byimmunofluorescence assay(IFA) ,theultrastructurecharacteristicsofthe DCpopulations wereobserved bytransmission electron microscope,andthe cytochemicalfeaturesofthem were alsoexamined.Results Theper centage of DCs was70 % ～80% inthe DC enriched populations asconfirmed byIFAusing mABⅫ.Transmission electron microscopyrevealedthatthe DCpopulations containedtwosubsets of DC,one wasconsideredas precursor cell derived DC(PC DC) ,thecellsurfaceofwhich had manyirregulardendrites with airregularnucleus,smallnu cleoli,numerous mitochondria,butscantylysosomes,ribosome,rough endoplasmic reticulum and free polysomes. Cytochemicalstainingshowedthe DCs were both peroxidase(POX) and non specific esterase(NSE) negative.The other wasindentifiedas monocyte derived DC(MO DC),thefeaturesofwhichincluded:irregularshape,manyruf fles and someroughlameson cellsurfaces,nephritic orhorseshoe nuclei with athick rim ofheterochromatin,nu merous mitochondria,much morelysosomes,ribosomes,rough endoplasmic reticulum and Golgiosome.The cyto chemicalreactionsindicatedthat mostofthem had POX positivegranules,and were NSEslightly positive.Conclu sion Cytokinesin vitroinduced DCsfrom human peripheral blood are heterogeneous.