CIK的体外增殖及体内外杀瘤活性的实验研究

目的:从人骨髓造血前体细胞体外培养扩增树突状细胞(dendritic cells,DCs),测定其表型及T细胞刺激活性.方法:采用Mini-MACS分离技术,从正常人骨髓、脐血分离CD34~ 造血干细胞,体外以重组hGM-CSF,hTNF-α,hIL-3诱导培养2周,流式细胞术检测扩增细胞的表面表型及细胞内IL-12的表达,体外同种混合淋巴细胞反应检测扩增DCs的T细胞刺激活性.结果:从正常人骨髓、脐血分离得到高纯度(>90%)的CD34~ 造血干细胞,经重组hGM-CSF,hTNF-α的共同诱导培养,扩增得到大量DCs,加人hIL-3可以进一步增加DCs产量;FACS检测表明,扩增的DCs表达HLA-DR,CD40,CD54,CD80,CD86分子,细胞内有hIL-12的P35,P40亚基的表达;与外周血单核细胞培养生成的DCs相比,由CD34~ 干细胞扩增的DCs具有更强的激发同种T细胞增殖的能力.结论:人CD34~ 干细胞体外经诱导培养,可以生成大量功能成熟的DCs,从而为进一步开展DCs的基础及临床研究打下了基础.     

人外周血树突状细胞的体外扩增及鉴定

树突状细胞(DC)作为抗原提呈细胞,在激发T细胞免疫应答及T细胞依赖性抗体生成中起重要作用,骨髓及外周血的CD34造血前体细胞在GM-CSF和TNF-α的作用下,可在体外分化发育,生成树突状细胞.本研究从正常人外周血分离获得单核细胞,加入100ng/ml hGM-CSF、500U/ml hIL-4,体外培养1周后,获得大量高纯度的树突状细胞,其高表达MHC-Ⅰ、MHC-Ⅱ类分子及共刺激分子B7-1和CD40,能强烈激发同种异体T淋巴细胞增殖,培养条件以应用自体血清或胎牛血清为最佳;单独应用hGM-CSF只能生成巨噬细胞;在培养末期加入TNFα可促进DC进一步成熟.本研究为DC的深入研究与临床应用奠定了基础.     

人脐血树突状细胞抗神经胶质瘤的体外免疫效应研究

目的　研究肿瘤抗原致敏的树突状细胞 (DC)对神经胶质瘤细胞的免疫杀伤效应。方法　应用免疫磁珠分选脐血 CD34细胞 ,经 SCF FL3 GM- CSF IL- 4 TNF- α的联合诱导培养 DC,采用相差显微镜观察树突状细胞的形态 ;流式细胞仪作 DC的表面标志检测 ;MTT比色法测定同种异型的混合淋巴细胞反应能力和诱导CTL 毒性的检测。结果　 (1)脐血 CD34细胞在体外经细胞因子联合刺激后呈典型的 DC形态 ;(2 )流式检测 CD4 0、CD80和 CD86等成熟 DC特异性表面标志呈高表达 ,分别与诱导前比较 ,差异均有显著性 ;(3) MTT法测得脐血来源的 DC较外周血 DC刺激同种异体 T淋巴细胞增殖能力弱 ;经肿瘤抗原负载的 DC体外诱导出较强 CTL 毒性。结论　脐血 CD34细胞经体外扩增诱导的 DC具有典型的 CTL 毒性 ,为临床应用脐血来源 DC抗神经胶质瘤的生物治疗提供了理论依据     

四种不同来源的树突状细胞的性能比较研究

目的:探讨正常人外周血单按细胞、肿瘤患者外周血单按细胞、脐带血CD34^ 细胞、非淋巴细胞白血病细胞来源的树突状细胞性能差异。方法：分别利用正常人及肿瘤患者的外周血分离单个核细胞，黏附贴壁法收集单核细胞，在GM-CSF、IL-4及TNF-α细胞因子组合下诱导树突状细胞；脐带血细胞则利用MACS系统分离纯化CD34^ 细胞作为树突状细胞的前体细胞，培养体系为FL SCF GM-CSF IL-4 TNF-α；非淋巴细胞白血病细胞作为树突状细胞的前体细胞培养条件同CD34^ 细胞。流式细胞仅分析诱导细胞的表型，电镜分析细胞形态。结果：利用4种不同来源的前体细胞在不同细胞因子组合下可分别诱导出DC细胞，细胞CD1a，CD80，HLA-DR的表达较诱导分化前明显上调．细胞形态学表明4种细胞来源的DC均有较典型的DC细胞形态，并均可刺激同种异体T细胞的增生。结论：利用健康人或肿瘤患者外周血可成功诱导出树突状细胞，利用脐血CD34^ 细胞及部分非淋巴细胞白血病原代细胞也可体外诱生出树突状细胞，不同来源的树突状细胞在形态及初步功能上无明显区别。     

IFN-α联合GM-CSF诱导胃癌患者外周血单个核细胞分化为树突状细胞

目的： 探索干扰素-α（interferon-α,IFN-α）联合粒细胞-巨噬细胞集落刺激因子（granulocyte-macrophage colony-stimulating factor, GM-CSF）体外诱导胃癌患者外周血单个核细胞（peripheral blood mononuclear cell,PBMC）向树突状细胞（dendritic cell,DC）分化的可能性。 方法： 10 例胃癌患者PBMC分别用GM-CSF 100 ng/ml联合IFN-α 500 IU/ml（命名为IFN-α DC） 或GM-CSF 100 ng/ml联合50 ng/ml IL-4（命名为IL-4 DC）体外培养,然后用CD40L、LPS诱导DC成熟。Giemsa染色法观察IFN-α DC和IL-4 DC的形态,流式细胞术分析IFN-α DC和IL-4 DC表面CD1a、CD80、CD83、CD86和HLA-DR的表达情况,同种异体混合淋巴细胞反应（mixed lymphocyte reaction,MLR）检测不同的成熟DC刺激同种异体T淋巴细胞增殖的能力。 结果： IFN-α DC和IL-4 DC均呈现典型DC形态。IFN-α DC和IL-4 DC分别在诱导第3天和第5天时,细胞表面CD1a、CD80、CD83、CD86和HLA-DR表达达到较高水平,成熟IFN-α DC表面CD83\[（78.25±15.36）% vs （50.14±10.24）%,P<0.05\]和CD86\[（84.84±10.12）% vs （62.93±15.12）%,P<0.05\]的表达均高于成熟IL-4 DC。成熟IFN-α DC刺激异体T淋巴细胞增殖能力强于未成熟IFN-α DC（P<005）。在DC与T细胞数量比为1∶40和1∶20时,成熟IFN-α DC刺激同种异体T淋巴细胞增殖的能力明显强于成熟IL-4 DC\[（39.43±9.21）% vs （27.34±10.63）%,（60.31±7.86）% vs （48.63±6.25）%;均P<0.05\]。 结论： 相比常用的IL-4联合GM-CSF诱导方法,IFN-α联合GM-CSF可以在更短时间内将胃癌患者PBMC诱导成具有更强刺激同种异体T淋巴细胞增殖能力的DC细胞,这可能与其表面CD83和CD86表达增高有关。     

粒-巨噬细胞集落刺激因子剂量对小鼠骨髓源性树突状细胞成熟状态的影响

 目的　观察不同剂量粒-巨噬细胞集落刺激因子（GM-CSF）对小鼠树突状细胞（DC）形态、纯度、成熟程度的影响,为不同用途的DC疫苗制备选择合适的GM-CSF质量浓度提供理论依据。方法　用低剂量（5 ng／ml）、常规剂量（20 ng／ml）、高剂量（50 ng／ml）GM-CSF联合重组小鼠白细胞介素-4（mIL-4）诱导小鼠骨髓细胞体外分化为DC,流式细胞术测定CD11c、CD80和CD86表达率。结果　低剂量GM-CSF组诱导的DC不具有典型的DC形态,低表达CD11c、CD80和CD86。常规剂量GM-CSF组诱导的DC高表达CD11c、CD80和CD86,具有更强的刺激同种异体淋巴细胞增殖能力。高剂量GM-CSF组CD11c、CD80和CD86表达率最高,并具有最强的刺激同种异体淋巴细胞增殖的能力。结论　GM-CSF剂量影响诱导分化的DC成熟程度。低剂量的GM-CSF诱导的DC为不成熟的DC,常规剂量和高剂量GM-CSF诱导分化的DC为细胞表型和功能成熟的DC。高剂量GM-CSF诱导的DC细胞表型和功能最成熟。     

脐血来源树突状细胞体外诱导抗卵巢癌免疫特异性

[目的]研究脐血来源树突状细胞(DC)体外诱导特异性抗卵巢癌细胞的免疫效应.[方法]①从脐血中分离单个核细胞(MNCs)后,获得单核细胞(Mo).粒单集落刺激因子(GM-CSF)和白介素4(IL-4)诱导分化,培养7天后应用流式细胞仪进行细胞表型分析.②诱导单核细胞分化的第3天加入人卵巢癌细胞株3AO的冻融抗原,共培养4天后获得负载肿瘤抗原的成熟DC;将致敏DC与从脐血中分离的同种异体T淋巴细胞共培养3天,获得细胞毒T淋巴细胞(CTL);四甲基偶氮唑蓝(MTT)法检测CTL及上清对人卵巢癌细胞株3AO、人胚肾细胞株293T(对照细胞)、人肝癌细胞株HCCC-9810的细胞毒作用.[结果]①脐血来源单核细胞(Mo)在GM-CSF和IL-4作用下,7天后可分化生成成熟的DC,高表达DC特异性抗原CDla、CD80(B7-1)、CD86(B7-2)、HLA-DR、CD83.②DC可负载并递呈肿瘤抗原,激活同种异体T淋巴细胞,诱导肿瘤特异性CTL产生.不同浓度CTL及上清对卵巢癌细胞3AO有特异性杀伤、抑制作用(P＜0.05).[结论]脐血中单核细胞可体外分化扩增为成熟的功能性DC,并诱导出特异性杀伤卵巢癌细胞的免疫效应.     

人白细胞介素17(IL-17)的克隆及其在大肠杆菌中的表达

树突状细胞(DC)是一类能特异性启动T淋巴细胞介导的免疫反应的抗原提呈细胞,与其它抗原提呈细胞不同的是,树突状细胞能激活初始型T淋巴细胞(naive T cells).近年来,人们试图通过刺激血液中的前体细胞以提高DC的量.最近,有学者通过GM-CSF、IL-4及MCM的培养单个核细胞中获得了DC.本研究对此法加以改良并从恒河猴血中获得了形态、表型及功能均与人成熟DC非常类似的DC.去除了T细胞的单个核细胞在含有1%人血清及GM-CSF和IL-4的培养基中培养,在第7天,50%的培养基换成单核细胞的条件培养基MCM.此培养基是获得成熟的具有免疫刺激作用的DC所必需的,这样可从2Oml血中获得0.5-1.0×10~6DC.对这些DC与在同样培养基中培养产生的巨噬细胞的比较表明,这些DC刺激同种异体T细胞的能力较巨噬细胞明显增强,而且这些DC的形态和表型很典型且能表达高水平的p55及与细胞内CD68.对比之下,贴壁的巨噬细胞表达非常低水平的p55及高水平的弥漫性CD68,用此方法制备的恒河猴     

白细胞介素18基因修饰的树突状细胞的表型分析及其免疫刺激活性 *

目的:研究IL-18基因修饰后的树突状细胞(DC)的表型和功能的变化.方法:以含IL-18基因的重组腺病毒体外转染小鼠骨髓来源的DC,RLIDS检测IL-18分泌水平,以RT-PCR检测IL-18mRNA表达,以FACS法分析其DC表型变化,采用TdR检测其MLR的变化.结果:IL-18基因修饰的DC有效分泌IL-18,可检测到IL-18mnRNA的表达,分泌上清具有诱导T细胞产生IFN-γ的作用,IL-18基因修饰的DC同种异体MLR明显增强,且仅能被IL-18抗体部分阻断,表型分析表明,IL-18基因修饰后的Ia,B7.2,ICAM-1太阳性的DC的比例明显增加.结论:IL-18基因修饰的DC可有效表达有功能的IL-18,部分表面分子表达上调,刺激同种异体T淋巴细胞增殖能力的提高,提示其抗原提星功能增强.     

负载腮腺腺样囊性癌抗原的树突状细胞诱导的抗肿瘤效应

目的:探讨腺样囊性癌肿瘤抗原负载的树突状细胞通过淋巴细胞介导的免疫反应,体外杀伤腺样囊性癌细胞的细胞毒性效应.方法:外周血单核细胞在GM-CSF + IL-4 的诱导下体外培养,用肿瘤细胞抗原冲击后,流式细胞仪检测树突状细胞抗原负载前后CD1a、CD83表达量的变化.MTT比色法检测同种异体的混合淋巴细胞反应和诱导细胞毒淋巴细胞CTL杀伤肿瘤细胞.结果:凋亡肿瘤抗原刺激后,CD83 表达增加﹙P<0.01﹚,而CD1a表达量下调﹙P<0.05).负荷肿瘤抗原树突状细胞体外诱导出明显的细胞不良反应,并刺激同种T淋巴细胞增殖.结论:GM-CSF + IL-4 诱导的单核细胞来源的树突状细胞,能在体外摄取肿瘤抗原而进一步成熟,通过激活淋巴细胞杀伤癌细胞.     

rhIL-2对人骨髓CFU-GM与CFU-E和BFU-E的影响

目的:为了探索rhIL-2对人骨髓造血干祖细胞体外增殖的影响。方法:本文采用甲基纤维素体外半固体培养法。结果:证明了50～1000U/ml rhIL-2单用时对人骨髓造血祖细胞的CFU-GM、CFU-E及BFU-E无直接刺激增生作用,当与GM-CSF联合应用时,100～400U/ml rhIL-2其CFU-GM集落形成率明显高于单用GM-CSF(100U/ml)的对照组(P<0.05),当与EPO(2U/mi)联合应用时其CFU-E,及BFU-E集落形成率也明显高于单用EPO对照组(P<0.05),而1000U/ml rhIL-2其三种集落形成数目呈下降趋势,可能有抑制作用。结论:以上结果说明100～400U/ml rhIL-2可应用于协同GM-CSF及EPO体外扩增骨髓干/祖细胞来进行骨髓移植。     

小鼠骨髓树突状细胞对胃癌细胞杀伤作用的研究

目的研究小鼠骨髓树突状细胞的制备及其对胃癌细胞的杀伤作用。方法添加细胞因子培养扩增小鼠骨髓源树突状细胞;流式细胞仪检测DC表面标志物CD11c、CD86的表达;MTT法测定DC对胃癌细胞的杀伤作用。结果培养第7天出现扩增之骨髓源DC,加入TNF-α24 h后,成熟骨髓源DC形成。每只615小鼠平均能分离出3×10^7个骨髓细胞,一只小鼠的骨髓单个核细胞DC的产量约为7×10^6个。加入TNF-α48 h后,DC表面标志物CD86阳性表达率较加入TNF-α前增高;CD11c阳性表达率加入TNF–α前后无差别。成熟DC与615小鼠胃癌细胞共孵育24 h、48 h,效靶比分别为10∶1、20∶1、30∶1时,DC对胃癌细胞的杀伤活性随效靶比的增大而增强。结论提取纯化小鼠骨髓细胞,经细胞因子GM-CSF、IL-4、TNF-α的刺激诱导可产生成熟DC,成熟DC对胃癌细胞有较强的杀伤作用,其杀伤活性随效靶比的增大而增强,随共孵育时间的延长杀伤活性有增强的倾向。     

健康人外周血树突状细胞的体外诱导及其功能分析

 目的　分析以健康人AB血清与rhCD40L体外诱导健康人外周血树突状细胞（DC）的功能。方法　对健康人外周血单个核细胞进行体外培养,在以健康人AB血清为基础的培养体系中加入粒细胞巨噬细胞集落刺激因子（GM-CSF）、 重组人白细胞介素（rhIL）-4、rhCD40L等细胞因子,诱导单个核细胞分化形成DC,采用倒置显微镜及瑞特-吉姆萨染色观察,流式细胞术行DC表型鉴定,四甲基偶氮唑蓝比色（MTT）法进行混合淋巴细胞反应（MLR）,检测其抗原刺激能力,酶联免疫吸附（ELISA） 法检测DC 培养上清IL-12的分泌。结果　培养7 d后的细胞具有典型的DC 形态,并上调表达DC 特征性表面分子CD83及共刺激分子CD40、CD80、CD86,第0、1 、3、5、7天,5个时间点间CD83、CD40、CD80、CD86、CD14表达差异有统计学意义（F值分别为50.253、243.769、248.181、191.267、226.339,均P＜0.05）。培养后的DC 可较强地刺激同种自体淋巴细胞增殖,GM-CSF加rhIL-4、rhCD40L组较GM-CSF加rhIL-4 组刺激反应能力强。培养的DC 自培养第5天始即有IL-12 分泌,未加CD40L组IL-12 p40分泌量为（42.92±1.54）pg／ml,加CD40L组为（136.18±5.27） pg／ml;培养第7天,IL-12 p40分泌明显增多,两组分别为（60.09±2.27） pg／ml及（322.30±30.60） pg／ml,差异有统计学意义（t＝-44.941、-22.611,均P＜0.05）。结论　健康人外周血单个核细胞可在以健康人AB血清与rhCD40L为主的培养体系中诱导成DC。     

In vitro immunization of patient T cells with autologous bone marrow antigen presenting cells pulsed with tumor lysates

Presentation of cell-associated antigen to T cells is a critical event in the initiation of an anti-tumor immune response but it appears to often be deficient or limiting. Here we report an experimental system for stimulation of human T lymphocytes using autologous antigen presenting cells (APCs) and autologous tumor cells. Two types of APCs were prepared from human bone marrow: MC and DC. MC were produced by using GM-CSF and SCF. DC were obtained with the same cytokines plus IL-4. DC and MC were generated in parallel from the same patients and their phenotypes and capacities to prime T lymphocytes were analyzed and compared. MC were CD14+, CD1a-, CD33+ and HLA-DR+. Two populations of DC were defined: immature DC were uniformly CD1a-; mature DC expressed CD1a, CD80, CD86, HLA-DR, CD54 and CD58 but lacked surface CD14. Stimulation of autologous T lymphocytes was studied by measuring their proliferation and cytotoxic function. In more than 80% of our experiments the proliferation of autologous T lymphocytes cocultured with APC pulsed or not with tumor cell lysates was higher than that of T cells cultured alone. DC were more effective than MC in stimulating proliferation of lymphocytes. The capacity of a patient's autologous bone marrow-derived APC to stimulate T cells when exposed to autologous tumor cell lysates suggest that such antigen-exposed APC may be useful in specific anti-tumor immunotherapy protocols.     

Dendritic cells and tumor specific immunity

Dendritic cells (DCs) are the most efficient antigen presenting cells (APCs) that initiate and modulate our internal immune responses in stimulating both B cells to produce various antibodies and T cells to control cell-mediated immunity. Such DCs can be classified into three groups based on their origin. One is the myeloid DCs originating from CD34+ stem cells that are further differentiated into CD14+ CD1a- phagocytotic, glass-adherent macrophages with the help of M-CSF, or into CD14- CD1a+, Birbeck granule containing LAG-1+ Langerhans cells by GM-CSF, TNF-alpha and TGF-beta 1 stimulation. The latter Langerhans cells appear to differentiate into DC1 as strong stimulators of T cells displaying large amounts of MHC-peptide complexes and co-stimulatory molecules, such as B7-1 and B7-2, after capturing antigens and migrating to a regional lymphoid organ. The second group is the lymphoid DCs originating from CD4+CD11c- cells, which differentiate into DC2 when cultured with IL-3. Third is the follicular dendritic cells (FDC) observed in lymphofollicules that capture foreign antigens with their Fc-receptor or complement-receptors and keep the antigens inside the follicules. DC1s secrete IL-12, which turns CD4 T cells into Th1 cells to induce cellular immunity, whereas DC2s favor production of Th2 cells to organize humoral immunity. Therefore, DCs appear to control our internal self-defense system. These unique features of DCs enable us to manipulate Th1 and Th2 activation selectively, and thus antigen-pulsed DCs are currently thought of as excellent tools to induce specific T cell immunity towards virus-infected cells or tumor cells.     

大鼠骨髓间充质干细胞分泌多种造血相关因子的研究

目的：初步探讨大鼠骨髓间充质干细胞（MSC）分离培养的方法及检测其所表达的多种细胞因子。方法：采用全骨髓贴壁法和单克隆培养法分离培养大鼠骨髓MSC,间接免疫荧光法鉴定其生物学特性,RT-PCR方法检测多种细胞因子的表达。结果：成功分离出大鼠骨髓MSC,CD44表达率为92.3%,CD45表达率为6.9%;RT-PCR分析示有干细胞因子（SCF）、血小板生成素（TPO）、粒细胞巨噬细胞集落刺激因子（GM-CSF）、转化生长因子（TGF-β1）的表达,不表达白介素-3（IL-3）。结论：成功从大鼠骨髓中分离培养出MSC,并分泌多种造血相关因子,这可能揭示了其促造血功能的部分分子基础。     

The Comparison of Biologic Characteristics between Mice Embryonic Stem Cells and Bone Marrow Derived Dendritic Cells

OBJECTIVE This research was to induce dendritic cells (DCs)from mice embryonic stem cells and bone marrow mononuclear cells in vitro, and then compare the biologic characteristics of them.METHODS Embryonic stem cells (ESCs) suspending cultured in petri dishes were induced to generate embryonic bodies (EBs).Fourteen-day well-developed EBs were transferred to histological culture with the same medium and supplemented 25 ng/ml GM-CSF and 25 ng/ml IL-3. In the next 2 weeks, there were numerous immature DCs outgrown. Meantime, mononuclear cells isolated from mice bone marrow were induced to derive dendritic cells by supplementing 25 ng/ml GM-CSF and 25 ng/ml IL-4, and then the morphology, phenotype and function of both dendritic cells from different origins were examined.RESULTS Growing mature through exposure to lipopolysaccharide (LPS), both ESC-DCs and BM-DCs exhibited dramatic veils of cytoplasm and extensive dendrites on their surfaces, highly expressed CD11c, MHC-Ⅱ and CD86 with strong capacity to stimulate primary T cell responses in mixed leukocyte reaction (MLR).CONCLUSION ESC-DC has the same biologic characteristics as BM-DC, and it provides a new, reliable source for the functional research of DC and next produce corresponding anti-tumor vaccine.     

All-trans-retinoic acid effects the growth, differentiation and apoptosis of normal human myeloid progenitors derived from purified CD34+ bone marrow cells.

We have previously shown that all-trans retinoic acid (ATRA) increases the number of CFU-GM colonies grown from unseparated human bone marrow cells with crude sources of colony stimulating factors. In this study, we further characterized the effect of ATRA on the growth of CFU-GM stimulated by individual cytokines from multiple samples of CD34+ enriched or purified human bone marrow cells. The number of IL-3- or GM-CSF-induced CFU-GM with 3 x 10(-7) M ATRA was 3.25+/-1.13, and 2.17+/-0.8-fold greater respectively, compared to controls without ATRA, while G-CSF had no effect and the ratio of colony-induced with or without ATRA was 1.06+/-0.17 (P = 0.00012). No colonies grew with ATRA + IL-6 or ATRA without a cytokine. Maximum enhancing effect on IL-3-induced CFU-GM occurred when ATRA was added on day 2, gradually diminished when delaying ATRA, and in cultures of day 9 or older adding ATRA had no effect. In 14 days liquid cultures of purified CD34+ cells with IL-3, ATRA increased the number of myeloid differentiated cells to 91-95%, compared to 37-70% with IL-3 alone. In addition, the number of apoptotic cells using the annexin V method increased after 14 days from 5.1% with IL-3 to 17.1% with IL-3 + ATRA and by the TUNEL in situ method from 10-26% to 60-95%, respectively. This study demonstrates that ATRA consistently enhances the growth of myeloid progenitors from CD34+ cells. This effect is dependent on the stimulating cytokine, suggesting the myeloid cells responding to ATRA are the less mature CFU-GMs that are targets of IL-3 and GM-CSF and not the G-CSF-responding mature progenitors. The growth stimulation by ATRA and IL-3 is also associated with granulocyte differentiation and increased apoptosis. These studies further suggest a potential role of pharmacological doses of ATRA on the development of normal human hematopoietic cells.     

IL-4 prevents the blockade of dendritic cell differentiation induced by tumor cells

Malignant cells may escape from the immune response in vivo because of a defective differentiation of professional antigen-presenting cells (APCs), i.e., dendritic cells (DCs). We recently reported that tumor cells release interleukin (IL)-6 and macrophage colony stimulating factor (M-CSF), which inhibit the differentiation of CD34+ cells into DCs and promote their commitment toward monocytic lineage with a poor APC function. The results presented here show that both IL-4 and IL-13 reverse the inhibitory effects of renal cell carcinoma conditioned media (RCC CM) or IL-6+M-CSF on the phenotypic and functional differentiation of CD34+ into DCs. IL-4 was found to act through a rapid blockade of the expression of M-CSF and the IL-6 receptor-transducing chain (gp130), along with a decrease of the secondary production of M-CSF, thereby preventing the loss of granulocyte macrophage colony stimulating factor (GM-CSF) receptor alpha chain expression on differentiating CD34+ cells. Consistent with these observations, the differentiation of DCs from monocytes cultured with GM-CSF and IL-4 was also impaired by RCC CM, but the minimal inhibitory concentrations of RCC CM were 10-fold higher than for CD34+ cells. In these conditions, monocytes cultured with GM-CSF and IL-4 also exhibited profound phenotypic changes (CD14+ D32+ CD86+ HLA-DR+ CD115(low) CD23(low) CD1a-) and a poor APC function. These alterations were overcome in a dose-dependent manner by IL-4 (5-500 IU/ml), although not beyond a 40% final concentration of RCC CM. The capacity of RCC CM to block DC differentiation from monocytes strongly correlated with IL-6 and M-CSF concentrations in medium. Taken together, these results demonstrate that IL-4 and IL-13 reverse the inhibitory effect of tumor cells on DC differentiation.