Neonatal dendritic cells are intrinsically biased against Th-1 immune responses

Dendritic cells (DCs) were derived from human peripheral blood monocytes or cord blood monocytes cultured in the presence of IL-4 and GM-CSF. Adult and cord DCs were observed to have comparable immature phenotypes. However, the increase in surface expression of HLA-DR and CD86 after addition of LPS was significantly attenuated in cord DCs, with CD25 and CD83 expression also markedly reduced. Cord DCs were also unable to produce IL-12p70, failed to down-regulate expression of the chemokine receptor CCR5 and induced lower levels of IFN-gamma production from allogeneic naive CD4+ T cells than their adult counterparts. In contrast, the kinetics of the production of TNF-alpha and IL-10 in response to LPS stimulation was comparable to adult DCs. The reduced ability of cord DCs to attain a fully mature adult phenotype, and to activate naive CD4+ T cells to produce IFN-gamma, suggests that they are intrinsically preprogrammed against the generation of Th-1 immune responses.     

Role of c-Jun N-terminal kinase on lipopolysaccharide induced maturation of human monocyte-derived dendritic cells

Dendritic cells (DCs) are potent antigen-presenting cells that play a pivotal role in the initiation of T cell-dependent immune responses. Immature DCs obtained from peripheral blood CD14+ monocytes by culture with granulocyte macrophage-colony stimulating factor (GM-CSF) and interleukin-4 (IL-4) differentiate into mature DCs upon stimulation with lipopolysaccharide (LPS). At least three families of mitogen-activated protein kinases (MAPKs), that is, extracellular signal-regulated kinases (ERK), c-Jun N-terminal kinases (JNK) and p38 MAPK, are involved in the DC maturation process. We report investigations of the role of JNK in the maturation of human monocyte-derived DCs. SP600125, a specific inhibitor of JNK, inhibited the LPS-induced up-regulation of CD80, CD83, CD86 and CD54, but augmented the up-regulation of HLA-DR. SP600125 slightly inhibited the down-regulation of FITC-dextran uptake during DC maturation. However, SP600125 did not affect the LPS induced up-regulation of allostimulatory capacity of DCs. SP600125 inhibited the release of IL-12 p70 and TNF-alpha from mature DCs. Although autologous T cells primed by the ovalbumin (OVA)-pulsed mature DCs produced IFN-gamma, but not IL-4, OVA-pulsed SP600125-treated mature DCs could initiate IL-4 production from autologous T cells. In contrast, a p38 MAPK inhibitor, SB203580, profoundly inhibited the phenotypic and functional maturation of DCs, while an ERK inhibitor, PD98059, had little or no effect. Taken together, the JNK signaling pathway appears to have a role that is distinct from the p38 MAPK and ERK cascades in the maturation process of DCs, and may be involved in the augmentation of Th2-prone T cell responses when it is suppressed.     

Diverse functional activity of CD83+ monocyte-derived dendritic cells and the implications for cancer vaccines

Antigen-loaded dendritic cells (DCs) provide key regulatory signals to T cells during a developing antitumor response. In addition to providing costimulation, mature DC provides cytokine and chemokine signals that can define the T1 vs T2 nature of the antitumor T-cell response as well as whether T cells engage in direct interactions with tumor cells. In serum-free culture conditions that hasten the differentiation of monocytes into mature DCs, certain agents, such as CD40L, accelerate phenotypic maturation (e.g., CD83 and costimulatory molecule expression) without influencing the acquisition of Dc1/Dc2 characteristics. In contrast, exposure to serum-free medium and interferon-gamma (IFN-gamma) rapidly influences CD83+ DCs to secrete high levels of IL-12, IL-6, and MIP-1beta, and promotes Dcl differentiation. In contrast, CD83+ DCs matured in serum-free medium in the absence of IFN-gamma, or in the presence of calcium signaling agents, prostaglandin-E2, or IFN-alpha, produce no IL-12, scant IL-6, and prodigious IL-8, MDC, and TARC, and promote Dc2 differentiation. T cells sensitized via IL-12-secreting, peptide-pulsed DCs secrete cytokines when subsequently exposed to relevant peptide-pulsed antigen-presenting cells (APCs) or to HLA-compatible tumor cells endogenously expressing the peptide. In contrast, T cells sensitized via IL-12 nonsecreting DC were limited to antigenic reactivation through APC contact rather than tumor cell contact. Therefore, the development of antitumor responses can be dramatically influenced not only by costimulation, but also by the cytokine and chemokine production of DCs, which must be considered in the development of cancer vaccines.     

香加皮羽扇豆烷乙酸酯(CPLA)对树突状细胞分化成熟的影响

目的：探讨香加皮羽扇豆烷乙酸酯（CPLA）对人外周血单个核细胞（PBMC）来源的树突状细胞（DC）在体外分化成熟及免疫活性的影响。方法：从人外周血分离单个核细胞，与细胞因子GM—CSF、IL-4共培养，于第5天加入DC的促成熟刺激剂TNF-α（阳性对照组）或CPLA。倒置显微镜和透射电镜下观察DC的形态；应用流式细胞术检测成熟DC的表面标志CD1a、CD83、CD80和CD86的表达情况；用ELISA检测DC培养上清中IL-12和IFN-γ的含量；用MTT法测定DC刺激T细胞增殖的能力。结果：培养10d后，经CPLA刺激的PBMC呈现出典型DC的形态学特征；成熟DC的特征性表面分子CD1a、CD83、CD80和CD86表达水平均明显上调（P〈0．05）；细胞培养上清中IL-12和IFN-γ含量明显增高（P〈0．05）；刺激T细胞增殖的能力明显增强（P〈0．05）。结论：CPLA可诱导PBMC来源的DC分化成熟，并可促进其细胞因子的分泌，增强DC的免疫调节活性。     

Polarization of naive T cells into Th1 or Th2 by distinct cytokine-driven murine dendritic cell populations: implications for immunotherapy

Dendritic cells (DCs) activate T cells and regulate their differentiation into T helper cell type 1 (Th1) and/or Th2 cells. To identify DCs with differing abilities to direct Th1/Th2 cell differentiation, we cultured mouse bone marrow progenitors in granulocyte macrophage-colony stimulating factor (GM), GM + interleukin (IL)-4, or GM + IL-15 and generated three distinct DC populations. The GM + IL-4 DCs expressed high levels of CD80/CD86 and major histocompatibility complex (MHC) class II and produced low levels of IL-12p70. GM and GM + IL-15 DCs expressed low levels of CD80/CD86 and MHC class II. The GM + IL-15 DCs produced high levels of IL-12p70 and interferon (IFN)-gamma, whereas GM DCs produced only high levels of IL-12p70. Naive T cells stimulated with GM + IL-4 DCs secreted high levels of IL-4 and IL-5 in addition to IFN-gamma. In contrast, the GM + IL-15 DCs induced higher IFN-gamma production by T cells with little or no Th2 cytokines. GM DCs did not induce T cell polarization, despite producing large amounts of IL-12p70 following activation. A similar pattern of T cell activation was observed after in vivo administration of DCs. These data suggest that IL-12p70 production alone, although necessary for Th1 differentiation, is not sufficient to induce Th1 responses. These studies have implications for the use of DC-based vaccines in immunotherapy of cancer and other clinical conditions.     

Airway epithelial IL-15 transforms monocytes into dendritic cells

IL-15 has recently been shown to induce the differentiation of functional dendritic cells (DCs) from human peripheral blood monocytes. Since DCs lay in close proximity to epithelial cells in the airway mucosa, we investigated whether airway epithelial cells release IL-15 in response to inflammatory stimuli and thereby induce differentiation and maturation of DCs. Alveolar (A549) and bronchial (BEAS-2B) epithelial cells produced IL-15 spontaneously and in a time- and dose-dependent manner after stimulation with IL-1beta, IFN-gamma, or TNF-alpha. Airway epithelial cell supernatants induced an increase of IL-15Ralpha gene expression in ex vivo monocytes, and stimulated DCs enhanced their IL-15Ralpha gene expression up to 300-fold. Airway epithelial cell-conditioned media induced the differentiation of ex vivo monocytes into partially mature DCs (HLA-DR+, DC-SIGN+, CD14+, CD80-, CD83+, CD86+, CCR3+, CCR6(+), CCR7-). Based on their phenotypic (CD123+, BDCA2+, BDCA4+, BDCA1(-), CD1a-) and functional properties (limited maturation upon stimulation with LPS and limited capacity to induce T cell proliferation), these DCs resembled plasmacytoid DCs. The effects of airway epithelial cell supernatants were largely blocked by a neutralizing monoclonal antibody to IL-15. Thus, our results demonstrate that airway epithelial cell-conditioned media have the capacity to differentiate monocytes into functional DCs, a process substantially mediated by epithelial-derived IL-15.     

IL-21 enhances SOCS gene expression and inhibits LPS-induced cytokine production in human monocyte-derived dendritic cells

Dendritic cells (DCs) play an important role in innate and adaptive immune responses. In addition to their phagocytic activity, DCs present foreign antigens to na?ve T cells and regulate the development of adaptive immune responses. Upon contact with DCs, activated T cells produce large quantities of cytokines such as interferon-gamma (IFN-gamma) and interleukin (IL)-21, which have important immunoregulatory functions. Here, we have analyzed the effect of IL-21 and IFN-gamma on lipopolysaccharide (LPS)-induced maturation and cytokine production of human monocyte-derived DCs. IL-21 and IFN-gamma receptor genes were expressed in high levels in immature DCs. Pretreatment of immature DCs with IL-21 inhibited LPS-stimulated DC maturation and expression of CD86 and human leukocyte antigen class II (HLAII). IL-21 pretreatment also dramatically reduced LPS-stimulated production of tumor necrosis factor alpha, IL-12, CC chemokine ligand 5 (CCL5), and CXC chemokine ligand 10 (CXCL10) but not that of CXCL8. In contrast, IFN-gamma had a positive feedback effect on immature DCs, and it enhanced LPS-induced DC maturation and the production of cytokines. IL-21 weakly induced the expression Toll-like receptor 4 (TLR4) and translation initiation region (TIR) domain-containing adaptor protein (TIRAP) genes, whereas the expression of TIR domain-containing adaptor-inducing IFN-beta (TRIF), myeloid differentiation (MyD88) 88 factor, or TRIF-related adaptor molecule (TRAM) genes remained unchanged. However, IL-21 strongly stimulated the expression of suppressor of cytokine signaling (SOCS)-1 and SOCS-3 genes. SOCS are known to suppress DC functions and interfere with TLR4 signaling. Our results demonstrate that IL-21, a cytokine produced by activated T cells, can directly inhibit the activation and cytokine production of myeloid DCs, providing a negative feedback loop between DCs and T lymphocytes.     

CCK-8对LPS诱导树突状细胞成熟的影响

目的: 观察八肽胆囊收缩素(CCK-8)对脂多糖(LPS)诱导树突状细胞(DCs)成熟的影响。方法: 采用粒-巨噬细胞集落刺激因子(GM-CSF)诱导法培养小鼠骨髓来源DCs,在LPS诱导其成熟过程中用不同浓度的CCK-8进行干预,采用流式细胞分析技术检测DCs表面主要组织相容性复合物II(MHC II)、分化群80(CD80)和分化群86(CD86)的表达;ELISA法检测DCs培养上清中白细胞介素1β(IL-1β)、白细胞介素6(IL-6)和肿瘤坏死因子α(TNF-α)的含量;MTT法检测CCK-8处理DCs对同种异体T细胞增殖反应的影响。结果: CCK-8剂量依赖性地抑制LPS诱导DCs表面CD80、CD86和MHC II表达(P<0.01, P<0.05 );CCK-8抑制LPS诱导DCs分泌IL-1β、IL-6和TNF-α(P<0.01);并且,CCK-8降低LPS诱导DCs刺激同种异体T淋巴细胞增殖的活性(P<0.05,P<0.01)。结论: CCK-8对LPS诱导DCs成熟过程中的细胞表型、细胞因子分泌和抗原提呈功能有抑制作用,提示CCK-8有可能在抗感染和抵抗自身免疫性疾病过程中发挥重要调节作用。     

Galectin-9 induces maturation of human monocyte-derived dendritic cells]

We investigated the role of galectin-9 (Gal-9) in maturation of dendritic cells (DC). Culture of immature DCs with exogenous Gal-9 markedly increased the surface expression of CD40, CD54, CD80, CD83, CD86, and HLA-DR in a concentration-dependent manner, although Gal-9 had no effect on differentiation of human monocytes into immature DCs. Gal-9-treated DCs secreted IL-12 but not IL-10, and they elicited the production of Th1 cytokines (IFN-gamma and IL-2), but not that of the Th2 cytokines (IL-4 and IL-5) by allogeneic CD4(+) T cells. These effects of Gal-9 on immature DCs were not essentially dependent on its lectin properties, given that they were only slightly inhibited by lactose. We further found that a Gal-9 mutant that lacks beta-galactoside binding activity reproduced the above activities, and that an anti-Gal-9 mAb suppressed them. Gal-9 induced phosphorylation of the p38 MAPK and ERK1/2 in DCs, and an inhibitor of p38 signaling, but not inhibitors of signaling by either ERK1/2 or phosphatidylinositol 3-kinase, blocked Gal-9-induced up-regulation of costimulatory molecule expression and IL-12 production. These findings suggest that Gal-9 plays a role not only in innate immunity but also in acquired immunity by inducing DC maturation and promoting Th1 immune responses.     

Nontoxic Shiga toxin derivatives from Escherichia coli possess adjuvant activity for the augmentation of antigen-specific immune responses via dendritic cell activation

Shiga toxin (Stx) derivatives, such as the Stx1 B subunit (StxB1), which mediates toxin binding to the membrane, and mutant Stx1 (mStx1), which is a nontoxic doubly mutated Stx1 harboring amino acid substitutions in the A subunit, possess adjuvant activity via the activation of dendritic cells (DCs). Our results showed that StxB1 and mStx1, but not native Stx1 (nStx1), resulted in enhanced expression of CD86, CD40, and major histocompatibility complex (MHC) class II molecules and, to some extent, also enhanced the expression of CD80 on bone marrow-derived DCs. StxB1-treated DCs exhibited an increase in tumor necrosis factor alpha and interleukin-12 (IL-12) production, a stimulation of DO11.10 T-cell proliferation, and the production of both Th1 and Th2 cytokines, including gamma interferon (IFN-gamma), IL-4, IL-5, IL-6, and IL-10. When mice were given StxB1 subcutaneously, the levels of CD80, CD86, and CD40, as well as MHC class II expression by splenic DCs, were enhanced. The subcutaneous immunization of mice with ovalbumin (OVA) plus mStx1 or StxB1 induced high titers of OVA-specific immunoglobulin M (IgM), IgG1, and IgG2a in serum. OVA-specific CD4+ T cells isolated from mice immunized with OVA plus mStx1 or StxB1 produced IFN-gamma, IL-4, IL-5, IL-6, and IL-10, indicating that mStx1 and StxB1 elicit both Th1- and Th2-type responses. Importantly, mice immunized subcutaneously with tetanus toxoid plus mStx1 or StxB1 were protected from a lethal challenge with tetanus toxin. These results suggest that nontoxic Stx derivatives, including both StxB1 and mStx1, could be effective adjuvants for the induction of mixed Th-type CD4+ T-cell-mediated antigen-specific antibody responses via the activation of DCs.     

Regulatory role of nitric oxide on monocyte-derived dendritic cell functions.

Nitric oxide (NO) has an established role in the defense against bacterial infections and exerts multiple modulatory activities on both inflammatory and immune responses. However, the relevance of NO on dendritic cell (DC) functions has been poorly investigated. In this study, we found that addition of the NO donor S-nitrosoglutathione (GSNO) to monocyte-derived DCs matured by lipopolysaccharide (LPS) or soluble CD40 ligand led to a decreased capacity to activate naive allogeneic T cells but a more prominent Th1 polarization, with increased interferon-gamma (IFN-gamma) secretion and reduced interleukin-5 (IL-5) release. The presence of GSNO during maturation of DCs caused a reduced expression of surface CD86, whereas CD80, CD83, and MHC molecule expression was not affected. Moreover, GSNO induced a dose-dependent decrease of IL-10 and enhancement of tumor necrosis factor-alpha (TNF-alpha) release from mature DCs. In parallel, a marked reduced production of IL-12 p40 subunit but no significant perturbation of the bioactive IL-12 p70 production was observed. Finally, GSNO significantly reduced the release of IP-10/CXCL10 and RANTES/CCL5 but not IL-8/CXCL8 by mature DCs. Although GSNO can strengthen the capacity of mature DCs to induce type 1 polarization of T lymphocytes, our data suggest that it elicits distinct anti-inflammatory functions, eventually reducing T lymphocyte proliferation and recruitment.     

Intrinsic ability of GM+IL-4 but not Flt3L-induced rat dendritic cells to promote allogeneic T cell hyporesponsiveness

The influence of GM+IL-4 and Flt3 ligand (FL) on phenotype and function of BM-derived DC from Lewis rats was investigated. GM+IL-4-induced DC, despite expression of CD80/CD86, were less stimulatory than FL-induced DC that expressed low CD80/CD86 and were efficient stimulators of allogeneic T cells. GM+IL-4 DC were CD11b+ OX62lo, whereas FL DC were CD11blo OX62+. Following activation, GM+IL-4 DC produced IL-10 and IL-6, but no IL-12p70, and were resistant to further maturation. FL DC produced IL-12p70, IFN-alpha/beta, IL-10 and IL-6 and underwent maturation. Repeated stimulation of T cells with GM+IL-4 DC inhibited proliferation, cytokine production and induced early T cell apoptosis. FL DC-activated T cells produced large amounts of IFN-gamma/IL-10 and exhibited late T cell apoptosis/necrosis. In vivo, GM+IL-4 DC induced alloAg-specific hyporesponsiveness following T cell restimulation. These results demonstrate that GM+IL-4 DC display intrinsic regulatory properties, inducing passive-cell-death in T cells with potential for inactivation/regulation of alloreactive T cells in transplantation.     

Dendritic cell activating peptides induce distinct cytokine profiles

High-mobility group box 1 protein (HMGB1), a DNA-binding nuclear and cytosolic protein, is a pro-inflammatory cytokine released by monocytes and macrophages. HMGB1 as well as its B box domain induce maturation of human dendritic cells (DCs). This report demonstrates that the B box domain induces phenotypic maturation of murine bone marrow-derived dendritic cells (BM-DCs) as evidenced by increased CD86, CD40 and MHC-II expression. The B box domain enhanced secretion of pro-inflammatory cytokines and chemokines: IL-1beta, IL-2, IL-5, IL-8, IL-12 and tumor necrosis factor (TNF)-alpha, but not IL-6 and IL-10. Furthermore, four peptides whose sequences correspond to different regions of HMGB1 induced production of IL-1beta, IL-2 and IL-12 (p70), but not IL-10 and IL-6 in mouse BM-DCs. Interestingly, these peptides differed in their capacity to induce TNF-alpha, IL-5, IL-18 and IL-8. B box domain as well as peptide-activated DCs acted as potent stimulators of allogeneic T cells in a mixed leukocyte reaction. DCs exposed to HMGB1 peptides induced proliferation of ovalbumin-specific syngeneic T cells. These DC-activating peptides could serve as an adjuvant in immunotherapeutic or vaccine context and the selective activity of these different peptides suggests a means to customize the functional properties of DCs.     

Plasmodium falciparum-free merozoites and infected RBCs distinctly affect soluble CD40 ligand-mediated maturation of immature monocyte-derived dendritic cells

Free plasmodium merozoites released from the parasitized hepatocytes and erythrocytes represent a transitory, extracellular stage in its mammalian host. In this study, we compared the effect of Plasmodium falciparum-free merozoites with infected RBCs (iRBCs) on the maturation of human monocyte-derived dendritic cells (DCs) in vitro. Phagocytosed-free merozoites prevented soluble CD40 ligand (sCD40L)-induced, phenotypic maturation of DCs and secretion of IL-12p70 but enhanced IL-10 production and primed, naive CD4+ cells to produce a high level of IL-10 compared with IFN-gamma. Free merozoites augmented sCD40L-induced ERK1/2 activation, and inhibition of ERK1/2 with its inhibitor PD98059 markedly abrogated IL-10 production and rescued IL-12 production. Therefore, the molecular mechanisms by which free merozoites antagonized sCD40L-induced DC maturation appeared to involve the activation of the ERK pathway. In contrast, phagocytosed iRBCs by itself induced DCs to semi-maturation, responded to CD40 signaling by maturing and secreting increased levels of TNF-alpha, IL-6, and also IL-12p70, and led to a pronounced, proinflammatory response by the allogenic CD4+ T cells. iRBCs regulate CD40-induced p38MAPK. Studies using inhibitors selective for p38MAPK (SB203580) showed that p38MAPK played an essential role in the maturation and function of DCs. Our results reveal the ability of free merozoites and iRBCs to distinctly alter the sCD40L-induced DC functioning by regulating the activation of the MAPK pathway that can inactivate or exacerbate immune responses to promote their survival and the development of parasite-specific pathologies.     

天花粉蛋白联合CD40信号激发的人MoDC介导Th2细胞分化的研究

探讨天花粉蛋白(Tk)联合CD40信号诱导人单核细胞来源DC(MoDC)的成熟活化及其介导Th2细胞分化的作用和机制。采用GM-CSF和IL-4联合方案体外诱导人MoDC,并利用鼠抗人CD40激发型单抗(5C11)刺激负载了Tk的DC制备成熟DC。采用免疫荧光标记和流式细胞术分析DC表型(CD80、CD83、CD86、CD14、PDL1)及其摄取FITC-Dextran的能力;ELISA法测定DC上清中IL-12p70的含量;胞内染色和流式细胞术检测经成熟DC活化的T细胞中CD4~+IFN-γ~+T和CD4~+IL-4~+T的比例。实验结果显示单独Tk不能刺激DC完全成熟,用Tk负载DC后必须联合外源性成熟刺激信号(如5C11),才能有效促进DC成熟,表现在CD80、CD83、CD86的上调表达,CD14下调表达,DC摄取FITC-Dextran的能力降低。与CD40信号单独诱导组相比,Tk联合CD40信号诱导的成熟DC表面PDL1分子呈下调表达,分泌IL-12的能力显著降低,明显提高T细胞中CD4~+IL-4~+T的比例。由此表明负载了Tk的成熟MoDC体外能有效介导Th2细胞分化。     

Differential regulation of DC function by Siphonodiol

Siphonodiol is a polyacetylene diol isolated from marine sponges Callyspongia sp. We demonstrate that the effect of Siphonodiol on the phenotypic and functional maturation of human monocyte derived DC in vitro. Human monocytes were exposed to Siphonodiol alone, or in combination with LPS and thereafter co-cultured with na?ve T cells. The expression levels of CD1a, CD80, CD83, CD86 and HLA-DR on LPS-primed DC were partially enhanced by Siphonodiol. Siphonodiol augmented the T cell stimulatory capacity in an allo MLR to LPS-primed DC. Siphonodiol dose-dependently enhanced the production of IL-12p70 by LPS-primed DC and this cytokine production was inhibited by anti-TLR4 mAb. IFN-gamma secretion from naive T cells co-cultured with DC differentiated with LPS was augmented by Siphonodiol. These results suggest that the enhancement of Th1 cells polarization to LPS-primed DC induced by Siphonodiol depends on TLR4 and via the activation of IL-12p70.     

Streptococcus pyogenes and Lactobacillus rhamnosus differentially induce maturation and production of Th1-type cytokines and chemokines in human monocyte-derived dendritic cells

Dendritic cells (DCs) are the most efficient antigen-presenting cells and thus, have a major role in regulating host immune responses. In the present study, we have analyzed the ability of Gram-positive, pathogenic Streptococcus pyogenes and nonpathogenic Lactobacillus rhamnosus to induce the maturation of human monocyte-derived DCs. Stimulation of DCs with S. pyogenes resulted in strong expression of DC costimulatory molecules CD80, CD83, and CD86 accompanied with a T helper cell type 1 (Th1) cytokine and chemokine response. S. pyogenes also induced interleukin (IL)-2 and IL-12 production at mRNA and protein levels. In addition, IL-23 and IL-27 subunits p40, p19, p28, and EBI3 were induced at mRNA level. In contrast, L. rhamnosus-stimulated DCs showed only moderate expression of costimulatory molecules and produced low levels of cytokines and chemokines. Furthermore, no production of IL-2 or IL-12 family cytokines was detected. Bacteria-induced DC maturation and especially cytokine and chemokine production were reduced when bacteria were heat-inactivated. Our results show that human monocyte-derived DCs respond differently to different Gram-positive bacteria. Although pathogenic S. pyogenes induced a strong Th1-type response, stimulation with nonpathogenic L. rhamnosus resulted in development of semi-mature DCs characterized by moderate expression of costimulatory molecules and low cytokine production.     

Type 1-polarized dendritic cells loaded with autologous tumor are a potent immunogen against chronic lymphocytic leukemia

Induction of active tumor-specific immunity in patients with chronic lymphocytic leukemia (CLL) and other hematologic malignancies is compromised by the deficit of endogenous dendritic cells (DCs). In attempt to develop improved vaccination strategies for patients with CLL and other tumors with poorly identified rejection antigens, we tested the ability of ex vivo-generated DCs to cross-present the antigens expressed by CLL cells and to induce CLL-specific, functional CTL responses. Monocyte-derived DCs from CLL patients were induced to mature using a "standard" cytokine cocktail (in IL-1beta, TNF-alpha, IL-6, and PGE2) or using an alpha-type 1-polarized DC (alphaDC1) cocktail (in IL-1beta, TNF-alpha, IFN-alpha, IFN-gamma, and polyinosinic:polycytidylic acid) and were loaded with gamma-irradiated, autologous CLL cells. alphaDC1 from CLL patients expressed substantially higher levels of multiple costimulatory molecules (CD83, CD86, CD80, CD11c, and CD40) than standard DCs (sDCs) and immature DCs, and their expression of CCR7 showed intermediate level. alphaDC1 secreted substantially higher (10-60 times) levels of IL-12p70 than sDCs. Although alphaDC1 and sDCs showed similar uptake of CLL cells, alphaDC1 induced much higher numbers (range, 2.4-38 times) of functional CD8+ T cells against CLL cells. The current demonstration that autologous tumor-loaded alphaDC1 are potent inducers of CLL-specific T cells helps to develop improved immunotherapies of CLL.     

Sphingosine kinase inhibitor suppresses a Th1 polarization via the inhibition of immunostimulatory activity in murine bone marrow-derived dendritic cells

Sphingosine kinase (Sphk) has been shown to be activated by growth factor and survival factors, and one of its products, sphingosine-1-phosphate, plays an important role in the regulation of various cellular responses. However, the effect of Sphk on the maturation and immunostimulatory function of dendritic cells (DCs) still remains largely unknown. In this study, we examined whether sphingosine kinase inhibitor (SKI) can influence co-stimulatory molecules (CD40, CD80, CD86 and MHC class II) and cytokine production (IL-12 and IL-10) in murine bone marrow-derived DCs. SKI significantly inhibited co-stimulatory molecules in DCs. SKI suppressed IL-12 production by DCs and IFN-gamma production by T cells. In addition, SKI-inhibited LPS induced the translocation of nuclear factor-kappaB, whereas it did not affect the degradation of IL-1 receptor-associated kinase-1 by LPS. These novel findings provide new insight into the immunopharmacological role of SKI in terms of its effects on DCs. These findings open a possibility for further understanding of the immunopharmacological functions of SKI, as well as therapeutic adjuvants for the treatment of DC-related acute and chronic diseases.     

Human epidermal Langerhans cells differ from monocyte-derived Langerhans cells in CD80 expression and in secretion of IL-12 after CD40 cross-linking

Langerhans cells (LCs) represent an immature population of myeloid dendritic cells (DCs). As a result of their unique Birbeck granules (BGs), langerin expression, and heterogeneous maturation process, they differ from other immature DCs. Monocyte-derived LCs (MoLCs) mimic epidermal LCs. MoLCs with characteristic BGs are generated by culturing blood-derived monocytes with granulocyte macrophage-colony stimulating factor, interleukin (IL)-4, and transforming growth factor-beta1. Here, we compare maturation-induced antigen expression and cytokine release of LCs with MoLCs. To achieve comparable cell populations, LCs and MoLCs were isolated by CD1c cell sorting, resulting in high purity. In unstimulated cells, CD40 was expressed at equal levels. After stimulation with CD40 ligand (CD40L), LCs and MoLCs acquired CD83 and increased CD86. High CD80 expression was exclusively detected in CD1c-sorted MoLCs. Human leukocyte antigen-DR and CD54 expression was found in all cell populations, however, at different intensities. CD40 triggering increased the potency of LCs and MoLCs to stimulate CD4+ T cell proliferation. Activated MoLCs released IL-12p70 and simultaneously, anti-inflammatory IL-10. The application of the Toll-like receptor ligands peptidoglycan, flagellin, and in particular, lipopolysaccharide (LPS) increased the corelease of these cytokines. LCs secreted IL-10 at a comparable level with MoLCs but failed to produce high amounts of IL-12p70 after application of danger signals. These data indicate that MoLCs as well as LCs display no maturation arrest concerning CD83 and CD86 expression. In difference to MoLCs, LCs resisted activation by CD40L and LPS in terms of IL-12 production. This shows that natural and generated LCs share similar features but differ in relevant functions.