成人外周血来源树突状细胞体外诱导培养动态监测

目的:分析体外诱导培养树突状细胞(dendritic cells,DCs)表面分子及成熟度的动态变化;方法:利用流式细胞仪(fluorescence activated cell sorter,FACS)检测分析DCs表面成熟标志分子CD83和T细胞辅助分子CD58、CD54、CD40、CD80、CD86、HLA-DR、HLA-ABC表达水平;结果:体外诱导培养5 d细胞即高表达DCs标志分子CD11c,细胞成熟度与细胞表面T细胞辅助分子的表达水平随培养天数的增加有一定提高,但显著低于大肠杆菌LPS刺激的DCs;结论:诱导培养6 d DCs表型为CD83 low、CD58low、CD54 low、CD40 low、CD80 low、CD86 low、HLA-DRhigh、HLA-ABChigh,为不成熟DCs.     

黄氏多糖对小鼠骨髓来源的树突状细胞表面分子MHC Ⅱ类分子表达的影响

目的:黄氏多糖对小鼠骨髓来源的树突状细胞表面分子MHCⅡ类分子表达的影响。方法:利用黄氏多糖作用小鼠骨髓来源的树突状细胞,利用流式细胞仪检测树突状细胞的表面MHCⅡ类分子表达的情况。结果:黄氏多糖作用后的小鼠骨髓来源的树突状细胞表面分子MHCⅡ类分子表达升高。结论:黄氏多糖能够提高小鼠骨髓来源的树突状细胞表面分子MHCⅡ类分子的表达。     

小鼠骨髓源性树突状细胞体外诱导培养方法的比较研究

目的探讨最佳体外诱导培养小鼠成熟树突状细胞（dendritic cells,DC）的方法。方法分离、纯化6周龄C57BL／6小鼠骨髓单核细胞,以含10％胎牛血清、20ng／ml重组小鼠粒细胞-巨噬细胞集落刺激因子（GM—CSF）和10ng／ml重组小鼠白细胞介素-4（IL-4）的RPMI-1640培养基培养7d,然后将细胞分成对照未刺激组、肿瘤坏死因子-α（TNF-α）刺激组和TNF-α＋脂多糖（lipopolysaccharides,LPS）刺激组。继续培养48h后,观察各组细胞形态,检测IL-12、IL-6浓度及细胞表面标志CD11c、CD80、CD86和MHC II。结果培养9d后,两刺激组培养的细胞经相差显微镜观察有DC生长。TNF—α刺激组细胞培养上清液中IL-6、IL-12含量显著高于对照组（P〈0．01）,但显著低于TNF—α＋LPS刺激组（P〈0．05）。3组均高表达CD11c,各组间无显著差异;而CD80、CD86和MHC II表达阳性率TNF-α刺激组显著高于对照组（P〈0．01）,TNF-α＋LPS刺激组显著高于单纯TNF—α刺激组（P〈0．05）。结论联合使用TNF-α与LPS刺激可使DC成熟度提高,分泌IL-6、IL-12增加。     

双歧杆菌完整肽聚糖对脐血来源树突状细胞分化成熟的影响

目的研究两歧双歧杆菌完整肽聚糖（WPG）对脐血来源树突状细胞（DC）形态及分泌细胞因子的影响,了解双歧杆菌WPG对DC分化、成熟及免疫调节功能的作用;并为益生菌及其生物活性成分的进一步开发提供依据。方法分离正常孕妇脐血单个核细胞诱导生成未成熟树突状细胞（Dendritic cells,DCs）,实验组在培养的第7天分别加入两歧双歧杆菌WPG（5μg／ml）、两歧双歧杆菌全菌（100μg/ml）,阳性对照组加入脂多糖（LPS）,阴性对照组仅加入培养基。倒置显微镜在培养各期形态学观察,流式细胞术检测表面标志物CD83及CD1a的表达,NLR检测DCs刺激同种异体T淋巴细胞能力,ELISA法测定DCs培养上清中IL-12p70、IL-10的分泌。结果脐血单核细胞在双歧杆菌WPG与GM-CSF、IL-4协同诱导作用下,能成为形态上具有典型树突状突起的DCs;诱导后的CB-MDDCs刺激同种异体T细胞的增殖能力及分泌IL-12：p70、IL-10的水平显著高于阴性对照组（P〈0．01）且细胞表面标志物CD83及CD1a的表达增加。结论（1）双歧杆菌WPG能影响CB-MDDCs的成熟状态。（2）双歧杆菌WPG对CB-MDDCs成熟程度及分泌细胞因子水平的影响强于双歧杆菌全菌,说明WPG是双歧杆菌主要免疫活性成分。     

阿萨希毛孢子菌对人外周血单核源树突状细胞表型和功能的影响

目的通过体外研究热灭活阿萨希毛孢子菌(Trichosporon asahii,T.asahii)孢子对健康人外周血单核源树突状细胞(dendritic cells,DCs)的表型和功能的影响,探讨DCs在T.asahii感染中可能发挥的作用。方法体外诱导培养正常人外周血DCs,与不同浓度的热灭活T.asahii孢子进行孵育,DCs与T.asahii浓度比分别为1∶1、1∶5和1∶10,RPMI-1640及脂多糖(lipopolysaccharide,LPS)刺激组分别作为空白对照及阳性对照,在显微镜下观察各组DCs形态的变化;瑞氏-姬姆萨染色观察实验组DCs对T.asahii的吞噬情况并计算吞噬率;流式细胞术检测各组DCs表面共刺激分子表达情况。结果诱导孵育过程中DCs形态发生明显改变;24 h时实验组部分DCs内可观察到不止一个被吞噬的T.asahii孢子,吞噬率组间差异有统计学意义(P0.05);与空白对照组相比,实验组DCs形态发生明显改变,且表面共刺激分子CD80、CD86、CD83表达水平明显升高(P0.05),但升高水平低于LPS刺激组,差异有统计学意义(P0.05)。结论人外周血DCs吞噬热灭活的T.asahii孢子后,形态发生改变,表面分子表达水平升高,DCs进一步成熟。     

甘草甜素对树突状细胞表型及生物学功能的影响

培养小鼠髓系DC2．4细胞,加入LPS（阳性对照组）或甘草甜素,用扫描电镜观察DC的超微结构、流式细胞仪检测DC表面分子MHCII、CD86及CD40的表达、4-氨基安替比林（4-AAP）比色检测DC内酸性磷酸酶活性、ELISA方法检测DC培养上清中IL—12的浓度,体外刺激淋巴细胞增殖实验检测DC对同种异体T淋巴细胞的刺激能力。结果表明,与对照组相比,甘草甜素刺激后,DC表面树突状突起增多,表面分子MHCⅡ、CD86及CD40表达增加,酸性磷酸酶活性下降,培养上清中IL-12浓度升高,刺激同种异体T淋巴细胞的能力也明显增强。结果表明,甘草甜素能够促进小鼠髓系DC2.4表型及功能的成熟。     

树突状细胞与马尔尼菲青霉酵母体外相互作用的研究

本文探讨了树突状细胞(DCs)在抗马尔尼菲青霉感染免疫中的作用。用细胞因子 rhGM-CSF 和rhIL-4诱导人外周血单核细胞分化为树突状细胞, 观察DCs的形态, 并用流式细胞仪进行DCs的表型测定, ELISA方法检测培养上清液IL-12p70的浓度, 混合淋巴细胞反应检测DCs刺激T淋巴细胞的增殖能力, 实时荧光定量PCR检测趋化因子受体CCR7、CXCR4的mRNA的表达。倒置显微镜下可见诱导获得的DCs细胞形态不规则, 表面伸展出大量树突。与马尔尼菲青霉酵母共同培养24 h后DCs的胞内含有大量的酵母细胞; 细胞表型CD86、CD83、HLA-DR和CD40的表达明显增高; 刺激T淋巴细胞增殖的能力增强; 趋化因子受体CCR7和CXCR4的mRNA表达量增加且能够产生IL-12p70但产生的量低于LPS刺激组。DCs能吞噬加热灭活的马尔尼菲青霉酵母, 并趋于成熟, 抗原呈递能力增加, 但是产生IL-12p70的量较低, 可能造成宿主抗马尔尼菲青霉酵母的细胞免疫功能的不足。     

生理层流剪切力对树突状细胞免疫表型的影响

从力学生物学的角度探索生理层流剪切力(shear stress,SS)对树突状细胞(dendritic cells,DCs)的形态、细胞骨架和免疫表型分子表达水平的影响。采用常规方法从CD14~+单核细胞诱导获得未成熟DCs(immature DCs,im DCs)和成熟DCs(mature DCs,m DCs),旋转锥板装置给DCs加载10 dyn/cm~2的剪切力,观察DCs形态和细胞骨架的变化,流式细胞术和实时荧光定量PCR技术检测DCs表面分子CD80、CD83和CD86在蛋白和基因水平上的表达情况。在流体剪切力的作用下,DCs的细胞直径变小(p0.05),细胞骨架(F-actin)发生了明显的重组。DCs的免疫表型分子CD80、CD83和CD86在蛋白和基因水平上的表达均受到影响。因此,DCs能够对生理层流剪切力作出应答,生理层流剪切力可能是DCs免疫功能的一个负调控因子,支持"力学免疫学(mechanoimmunology)"和"免疫力学生物学(immunomechanobiology)"的学术观点,这对于深入理解DCs的免疫调节功能来说具有重要意义。     

树突状细胞与马尔尼菲青霉酵母体外相互作用的研究

本文探讨了树突状细胞(DCs)在抗马尔尼菲青霉感染免疫中的作用.用细胞因子rhGM-CSF和rhIL-4诱导人外周血单核细胞分化为树突状细胞,观察DCs的形态,并用流式细胞仪进行DCs的表型测定,ELISA方法检测培养上清液IL-12p70的浓度,混合淋巴细胞反应检测DCs刺激T淋巴细胞的增殖能力.实时荧光定量PCR检测趋化因子受体CCR7、CXCR4的mRNA 的表达.倒置显微镜下可见诱导获得的DCs细胞形态不规则,表面伸展出大量树突.与马尔尼菲青霉酵母共同培养24 h后DCs的胞内含有大量的酵母细胞;细胞表型CD86、CD83、HLA-DR和CD40的表达明显增高;刺激T淋巴细胞增殖的能力增强;趋化因子受体CCR7和CXCR4的mRNA表达量增加且能够产生IL-12p70但产生的量低于LPS刺激组.DCs能吞噬加热灭活的马尔尼菲青霉酵母,并趋于成熟,抗原呈递能力增加,但是产生IL-12p70的量较低,可能造成宿主抗马尔尼菲青霉酵母的细胞免疫功能的不足.     

小鼠骨髓源树突状细胞体外诱导培养及初步鉴定

用重组小鼠粒细胞-巨噬细胞集落刺激因子（rmGM-CSF）和重组小鼠白细胞介素4（rmIL-4）体外诱导小鼠骨髓细胞分化为树突状细胞,进行形态学变化观察,分析细胞表面分子,刺激T细胞增殖,探讨小鼠骨髓源树突状细胞（BMDC）体外诱导培养并进行初步鉴定。体外培养9d后BMDC可达80%以上,光镜下可见典型的树突状细胞形态。清楚表达成熟期主要表面标志物,可显著刺激同种异体混合淋巴细胞增殖。获得了较高纯度的BMDC,避免了使用传统磁珠分离方法所带来的成本高,操作复杂,产出率低的弊端,为研究BMDC功能以及运用开展下游实验提供材料。     

Butyrate inhibits functional differentiation of human monocyte-derived dendritic cells

Dendritic cells (DCs) play a key role in immune function through antigen presentation by MHC and CD1, as well as cytokine production that shapes the immune response. Here we report that butyrate, a histone deacetylase inhibitor, inhibits the functional differentiation of human monocyte-derived DCs. Mature DCs were generated from monocytes in the presence of granulocyte macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4), followed by 2 day LPS stimulation. Butyrate treatment throughout the culture period inhibited the expression of CD1 molecules, but not on CD83, CD86, and MHC molecules. The suppression was exerted at protein and mRNA levels. Butyrate-treated immature DCs also showed decreased expression of CD1 molecules. Moreover the butyrate-treated immature DCs showed lower production of IL-12 p40 and IL-6 in response to lipopolysaccharides and induced less Th1 cells in allogenic mixed lymphocyte reactions. Our results imply that histone acetylation is involved in regulating immune responses through regulating functional differentiation of DC. Thus HDAC may be one of the targets for controlling the immune response.     

Analysis of maturation of murine dendritic cells (DCs) induced by purified Ganoderma lucidum polysaccharides (GLPs)

To investigate and analyze induction of phenotypic and functional maturation of murine DCs by GLP. Both phenotypic and functional activities were assessed with use of conventional scanning electronic microscopy (SEM) for the morphology of the DCs, transmitted electron microscopy (TEM) for intracellular lysosomes inside the DC, cellular immunohistochemistry for phagocytosis by the DCs, flow cytometry (FCM) for the changes in key surface molecules, bio-assay for the activity of acid phosphatases (ACP), and ELISA for the production of pro-inflammatory cytokine IL-12. It was found that GLP induced phenotypic maturation, as evidenced by increased expression of CD86, CD40, and MHC II. Functional experiments showed the down-regulation of ACP inside the DCs, which occurs when phagocytosis of DCs decreased, and antigen presentation increased with maturation. Finally, GLP increased the production of IL-12.These data reveals that GLP promotes effective activation of murine DCs. This adjuvant-like activity may have therapeutic applications in clinical settings that require a boosting of the immune response. Therefore concluded that GLP can exert positive induction to murine DCs at the used concentration.     

17beta-estradiol alters the activity of conventional and IFN-producing killer dendritic cells

Estrogens increase aspects of innate immunity and contribute to sex differences in the prevalence of autoimmune diseases and in response to infection. The goal of the present study was to assess whether exposure to 17beta-estradiol (E2) affects the development and function of bone marrow-derived dendritic cells and to determine whether similar changes are observed in CD11c(+) splenocytes exposed to E2 in vivo. E2 facilitated the differentiation of BM precursor cells into functional CD11c(+)CD11b(+)MHC class II(+) dendritic cells (DCs) with increased expression of the costimulatory molecules CD40 and CD86. Exposure of bone marrow-derived dendritic cells to E2 also enhanced production of IL-12 in response to the TLR ligands, CpG and LPS. In contrast, CD11c(+) cells isolated from the spleens of female C57BL/6 mice that were intact, ovariectomized, or ovariectomized with E2 replacement exhibited no differences in the number or activity of CD11c(+)CD11b(+)MHC class II(+) DCs. The presence of E2 in vivo, however, increased the number of CD11c(+)CD49b(+)NK1.1(low) cells and reduced numbers of CD11c(+)CD49b(+)NK1.1(high) cells, a surface phenotype for IFN-producing killer DCs (IKDCs). Ultrastructural analysis demonstrated that CD11c(+)NK1.1(+) populations were comprised of cells that had the appearance of both DCs and IKDCs. CD11c(+) splenocytes isolated from animals with supplemental E2 produced more IFN-gamma in response to IL-12 and IL-18. These data illustrate that E2 has differential effects on the development and function of DCs and IKDCs and provide evidence that E2 may strengthen innate immunity by enhancing IFN-gamma production by CD11c(+) cells.     

Dynamics of dendritic cell phenotype and interactions with CD4+ T cells in airway inflammation and tolerance

An emerging concept is that different types of dendritic cells (DCs) initiate different immune outcomes, such as tolerance vs inflammation. In this study, we have characterized the DCs from the lung draining lymph nodes of mice immunized for allergic airway inflammation or tolerance and examined their interactions with CD4(+) T cells. The DC population derived from tolerized mice was predominantly CD11c(+), B220(+), Gr-1(+), CD11b(-), and MHC class II(low), which resembled plasmacytoid-type DCs whereas DCs from the inflammatory condition were largely CD11c(+), B220(-), Gr-1(-), CD11b(+), and MHC class II(high) resembling myeloid-type DCs. The DCs from the tolerogenic condition were poor inducers of T cell proliferation. DCs from both conditions induced T cell IL-4 production but the T cells cultured with tolerogenic DCs were unresponsive to IL-4 as indicated by inhibition of STAT6 activation and expression of growth factor-independent 1, which has been recently shown to be important for STAT6-activated Th2 cell expansion. Our data suggest that airway tolerance vs inflammation is determined by the DC phenotype in lung draining lymph nodes.     

IL-13 regulates the immune response to inhaled antigens

The large inhibitory effect of IL-13 blockers on the asthma phenotype prompted us to ask whether IL-13 would play a role in regulating the allergic immune response in addition to its documented effects on structural pulmonary cells. Because IL-13 does not interact with murine T or B cells, but with monocytes, macrophages, and dendritic cells (DCs), we examined the role of IL-13 in the activation of pulmonary macrophages and DCs and in the priming of an immune response to a harmless, inhaled Ag. We found that a majority of cells called "alveolar or interstitial macrophages" express CD11c at high levels (CD11c(high)) and are a mixture of at least two cell types as follows: 1) cells of a mixed phenotype expressing DC and macrophage markers (CD11c, CD205, and F4/80) but little MHC class II (MHC II); and 2) DC-like cells expressing CD11c, CD205, MHC II, and costimulatory molecules. Endogenous IL-13 was necessary to induce and sustain the increase in MHC II and CD40 expression by pulmonary CD11c(high) cells, demonstrated by giving an IL-13 inhibitor as a measure of prevention or reversal to allergen-primed and -challenged mice. Conversely, IL-13 given by inhalation to naive mice increased the expression of MHC II and costimulatory molecules by CD11c(high) cells in an IL-4Ralpha-dependent manner. We found that exogenous IL-13 exaggerated the immune and inflammatory responses to an inhaled, harmless Ag, whereas endogenous IL-13 was necessary for the priming of naive mice with an inhaled, harmless Ag. These data indicate that blockade of IL-13 may have therapeutic potential for controlling the immune response to inhaled Ags.     

Directed differentiation of human embryonic stem cells into functional dendritic cells through the myeloid pathway

We have established a system for directed differentiation of human embryonic stem (hES) cells into myeloid dendritic cells (DCs). As a first step, we induced hemopoietic differentiation by coculture of hES cells with OP9 stromal cells, and then, expanded myeloid cells with GM-CSF using a feeder-free culture system. Myeloid cells had a CD4+CD11b+CD11c+CD16+CD123(low)HLA-DR- phenotype, expressed myeloperoxidase, and included a population of M-CSFR+ monocyte-lineage committed cells. Further culture of myeloid cells in serum-free medium with GM-CSF and IL-4 generated cells that had typical dendritic morphology; expressed high levels of MHC class I and II molecules, CD1a, CD11c, CD80, CD86, DC-SIGN, and CD40; and were capable of Ag processing, triggering naive T cells in MLR, and presenting Ags to specific T cell clones through the MHC class I pathway. Incubation of DCs with A23187 calcium ionophore for 48 h induced an expression of mature DC markers CD83 and fascin. The combination of GM-CSF with IL-4 provided the best conditions for DC differentiation. DCs obtained with GM-CSF and TNF-alpha coexpressed a high level of CD14, and had low stimulatory capacity in MLR. These data clearly demonstrate that hES cells can be used as a novel and unique source of hemopoietic and DC precursors as well as DCs at different stages of maturation to address essential questions of DC development and biology. In addition, because ES cells can be expanded without limit, they can be seen as a potential scalable source of cells for DC vaccines or DC-mediated induction of immune tolerance.     

IFN-alpha skews monocyte differentiation into Toll-like receptor 7-expressing dendritic cells with potent functional activities

IFN-alpha is an important cytokine for the generation of a protective T cell-mediated immune response to viruses. In this study, we asked whether IFN-alpha can regulate the functional properties of dendritic cells (DCs). We show that monocytes cultured in the presence of GM-CSF and IFN-alpha can differentiate into DCs (IFN-alpha-derived DCs (IFN-DCs)). When compared with DCs generated in the presence of GM-CSF and IL-4 (IL-4-derived DCs), IFN-DCs exhibited a typical DC morphology and expressed, in addition to DC markers CD1a and blood DC Ag 4, a similar level of costimulatory and class II MHC molecules, but a significantly higher level of MHC class I molecules. After maturation with CD40 ligand, IFN-DCs up-regulated costimulatory, class I and II MHC molecules and expressed mature DC markers such as CD83 and DC-lysosome-associated membrane protein. IFN-DCs were endowed with potent functional activities. IFN-DCs secreted large amounts of the inflammatory cytokines IL-6, IL-10, TNF-alpha, IL-1beta, and IL-18, and promoted a Th1 response that was independent of IL-12p70 and IL-18, but substantially inhibited by IFN-alpha neutralization. Furthermore, immature IFN-DCs induced a potent autologous Ag-specific immune response, as evaluated by IFN-gamma secretion and expansion of CD8(+) T cells specific for CMV. Also, IFN-DCs expressed a large number of Toll-like receptors (TLRs), including acquisition of TLR7, which is classically found on the natural type I IFN-producing plasmacytoid DCs. Like plasmacytoid DCs, IFN-DCs could secrete IFN-alpha following viral stimulation or TLR7-specific stimulation. Taken together, these results illustrate the critical role of IFN-alpha at the early steps of immune response to pathogens or in autoimmune diseases.     

High mobility group box protein 1: an endogenous signal for dendritic cell maturation and Th1 polarization

High mobility group box protein 1 (HMGB1), a DNA binding nuclear and cytosolic protein, is a proinflammatory cytokine released by monocytes and macrophages. This study addressed the hypothesis that HMGB1 is an immunostimulatory signal that induces dendritic cell (DC) maturation. We show that HMGB1, via its B box domain, induced phenotypic maturation of DCs, as evidenced by increased CD83, CD54, CD80, CD40, CD58, and MHC class II expression and decreased CD206 expression. The B box caused increased secretion of the proinflammatory cytokines IL-12, IL-6, IL-1alpha, IL-8, TNF-alpha, and RANTES. B box up-regulated CD83 expression as well as IL-6 secretion via a p38 MAPK-dependent pathway. In the MLR, B box-activated DCs acted as potent stimulators of allogeneic T cells, and the magnitude of the response was equivalent to DCs activated by exposure to LPS, nonmethylated CpG oligonucleotides, or CD40L. Furthermore, B box induced secretion of IL-12 from DCs as well as IL-2 and IFN-gamma secretion from allogeneic T cells, suggesting a Th1 bias. HMGB1 released by necrotic cells may be a signal of tissue or cellular injury that, when sensed by DCs, induces and/or enhances an immune reaction.     

A dendritic cell population generated by a fusion of GM-CSF and IL-21 induces tumor-antigen-specific immunity

We have previously shown that the fusion of GM-CSF and IL-21 (GIFT-21) possesses a potent immune stimulatory effect on myeloid cells. In this study, we define the effect of GIFT-21 on naive murine monocytes (GIFT-21 dendritic cells [DCs]), which express increased levels of Gr-1, CD45R, MHC class I, CD80, CD86, and CXCR4 and suppress CD11c and MHC class II. Compared with conventional dendritic cells, GIFT-21 DCs produced substantially more CCL2, IL-6, TNF-α, and IFN-α and induced significantly greater production of IFN-γ by CD8(+) T cells in MHC class I-restricted Ag presentation assays. B16 melanoma and D2F2 Neu breast cancer growth was inhibited in mice treated with Ag-naive GIFT-21 DCs. This effect was lost in CD8(-/-) and CCR2(-/-) mice and when mice were treated with β(2)-microglobulin-deficient GIFT-21 DCs, indicating that GIFT-21 DCs migrated to and sampled from the tumors to present tumor Ags to CCL2 recruited CD8(+) T cells via MHC class I. We propose that autologous GIFT-21 DCs may serve as a cell therapy platform for the treatment of cancer.     

Synergistic induction of antigen-specific CTL by fusions of TLR-stimulated dendritic cells and heat-stressed tumor cells

Dendritic cell (DC)/tumor cell fusion cells (FCs) can induce potent CTL responses. The therapeutic efficacy of a vaccine requires the improved immunogenicity of both DCs and tumor cells. The DCs stimulated with the TLR agonist penicillin-killed Streptococcus pyogenes (OK-432; OK-DCs) showed higher expression levels of MHC class I and II, CD80, CD86, CD83, IL-12, and heat shock proteins (HSPs) than did immature DCs. Moreover, heat-treated autologous tumor cells displayed a characteristic phenotype with increased expression of HSPs, carcinoembryonic Ag (CEA), MUC1, and MHC class I (HLA-A2 and/or A24). In this study, we have created four types of FC preparation by alternating fusion cell partners: 1) immature DCs fused with unheated tumor cells; 2) immature DCs fused with heat-treated tumor cells; 3) OK-DCs fused with unheated tumor cells; and 4) OK-DCs fused with heat-treated tumor cells. Although OK-DCs fused with unheated tumor cells efficiently enhanced CTL induction, OK-DCs fused with heat-treated tumor cells were most active, as demonstrated by: 1) up-regulation of multiple HSPs, MHC class I and II, CEA, CD80, CD86, CD83, and IL-12; 2) activation of CD4+ and CD8+ T cells able to produce IFN- gamma at higher levels; 3) efficient induction of CTL activity specific for CEA or MUC1 or both against autologous tumor; and 4) superior abilities to induce CD107+ IFN-gamma+ CD8+ T cells and CD154+ IFN-gamma+ CD4+ T cells. These results strongly suggest that synergism between OK-DCs and heat-treated tumor cells enhances the immunogenicity of FCs and provides a promising means of inducing therapeutic antitumor immunity.