Monocyte derived dendritic cell responses in common variable immunodeficiency

The phenotype and function of monocyte derived dendritic cells (MdDC) were investigated in 25 patients with common variable immunodeficiency (CVID) to test for abnormalities that might help explain the failure of antibody production. Using MHC class II DR and CD86 as markers of maturation, DCs from the majority of CVID patients were normal. However 5 patients, the majority of whom had affected family members who had previously been shown to have a susceptibility genetic locus in the MHC region, expressed abnormally low levels of DR on repeated testing, in some cases associated with a reduced capacity to support antigen stimulated T cell proliferation; nevertheless costimulatory molecules for production of IL-13, IL-10 and IFN-gamma from T cells were intact. In contrast to DCs from healthy donors, DCs from many CVID patients had high spontaneous production of IL-8 and lipopolysaccharide stimulation often caused a reduction in DR expression. Expression of other cytokines (IL-1a, IL-6 and IL-12), either before or after LPS stimulation, was normal. The data suggests there is a fundamental defect in the maturation of MdDCs in a subset of CVID patients that may compromise antigen presentation and subsequent antibody production.     

Stimulatory and inhibitory differentiation of human myeloid dendritic cells

Dendritic cells (DCs) play a critical obligate role in presenting antigens to T cells for activation. In the process, upon antigen capture, DCs undergo maturation and become more stimulatory. Human myeloid DCs can be generated from various sources, including blood, bone marrow, and CD34(+) stem cells. As such, plastic-adherent monocytes from circulation have served as a ready source for generating myeloid DCs in culture in granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4) for translational research in active specific immunotherapy, especially in cancer, with the belief that they are essentially stimulatory or "immunogenic." Here we show that in vitro cultures of plastic-adherent circulating monocytes in GM-CSF and IL-4 followed by further maturation in interferon-gamma plus bacterial superantigens (DC maturing agents) can give rise to two diametrically opposite types of DCs-one stimulatory and another inhibitory. The stimulatory DCs express higher amounts of costimulatory molecules, synthesize IL-12, and efficiently stimulate naive allogeneic T cells in mixed lymphocyte reaction (MLR). The inhibitory DCs, in contrast, express lower concentrations of the critical costimulatory molecules, synthesize large amounts of IL-10, and are nonstimulatory in allogeneic primary MLR. Moreover, while the stimulatory DCs further amplify proliferation of T cells in lectin-driven proliferation assays, the inhibitory DCs totally block T cell proliferation in similar assays, in vitro. Most interestingly, neutralization of the endogenously derived IL-10 with anti-IL-10 antibody in DC cultures repolarizes the inhibitory DCs toward stimulatory phenotype. Accordingly, these observations have important implications in translational research involving myeloid DCs.     

Azithromycin drives in vitro GM-CSF/IL-4-induced differentiation of human blood monocytes toward dendritic-like cells with regulatory properties

Azithromycin, a macrolide antibacterial, has been shown to modify the phenotype of macrophages. We have investigated whether azithromycin in vitro is able to modulate the differentiation of human blood monocytes to DCs. iA-DCs appear to have a unique phenotype, characterized by increased granularity, adherence, and a surface molecule expression profile similar to that of MDCs, namely, CD1a?CD14?CD71?CD209(high), as well as high CD86 and HLA-DR expression. The iA-DC phenotype is associated with increased IL-6 and IL-10 release, increased CCL2 and CCL18 expression and release, and M-CSF expression, as well as reduced CCL17 expression and release. Upon maturation with LPS, A-DCs and MDCs exhibit decreased expression of HLA-DR and costimulatory molecules, CD40 and CD83, as well as an increase in IL-10 and a decrease in CCL17 and CXCL11 secretion. These modulated responses of iA-DCs were associated with the ability to reduce a MLR, together with enhanced phagocytic and efferocytotic properties. Azithromycin, added 2 h before activation of iDCs with LPS, enhanced IL-10 release and inhibited IL-6, IL-12p40, CXCL10, CXCL11, and CCL22 release. In conclusion, azithromycin modulates the differentiation of blood monocyte-derived DCs to form iA-DCs with a distinct phenotype similar to that of iMDCs, accompanied by enhanced phagocytic and efferocytic capabilities. It also modifies LPS-induced DC maturation by decreasing surface molecule expression required for T cell activation, increasing IL-10 production, and inducing MLR-reducing properties.     

TRAF6在人PBMC来源的树突状细胞成熟中的作用

目的: 探讨肿瘤坏死因子受体相关因子6(TRAF6)在不同成熟度树突状细胞(DCs)中的表达及其对DCs免疫刺激活性的影响。方法: 重组人粒细胞-巨噬细胞集落刺激因子(rhGM-CSF)和重组人白细胞介素4(IL-4)体外诱导的人外周血单个核细胞来源的DCs,经TNF-α、LPS和细胞因子鸡尾酒法诱导成熟,分别以流式细胞术、ELISA和混合淋巴细胞反应(MLR)检测DCs的表面标志、IL-12分泌和刺激淋巴细胞增殖能力。Western blotting和real-time PCR分别检测各组TRAF6蛋白水平及mRNA的表达。结果: 各组DCs均表达TRAF6,成熟状态最佳的鸡尾酒诱导下DCs的TRAF6蛋白表达最高。高表达TRAF6的DCs可促进IL-12的分泌,增强刺激淋巴细胞增殖的能力。结论: TRAF6是调节DCs激活和成熟的关键因素。     

Stimulatory and inhibitory maturation of human macrophage-derived dendritic cells.

Circulating human macrophages are often used to generate dendritic cells (DCs) by culturing them in granulocyte macrophage-colony stimulating factor (GM-CSF) and interleukin-4 (IL-4). As DCs are superb antigen-presenting cells, these types of myeloid DCs are now used in many DC-based vaccination protocols, especially in cancer, with the belief that they are essentially stimulatory or 'immunogenic'. Here we show that just as peripheral macrophage-derived myeloid DCs can be stimulatory, in vitro cultures of myeloid DCs in GM-CSF and IL-4 followed by further maturation in interferon-gamma plus bacterial superantigens (as DC maturing agents) can give rise to DCs that are functionally inhibitory. The stimulatory DCs express higher amounts of costimulatory molecules, synthesize IL-12, and efficiently stimulate naive allogeneic T cells in mixed lymphocyte reaction (MLR). The inhibitory DCs, in contrast, express lower concentrations of the critical costimulatory molecules, synthesize large amounts of IL-10, and are either marginally stimulatory or nonstimulatory in MLR. Moreover, while the stimulatory DCs further amplify proliferation of T cells in lectin-driven proliferation assays, the inhibitory DCs suppress T cell proliferation in similar assays, in vitro. Most interestingly, neutralization of the endogenously derived IL-10 with anti-IL-10 antibody with DC cultures as well as exposure of the inhibitory DCs to CpG oligonucleotides or to in vitro activated autologous CD4+ T helper cells repolarize them into stimulatory phenotype. Accordingly, these observations have important implications in translational research involving myeloid DCs.     

Selective deficits in blood dendritic cell subsets in common variable immunodeficiency and X-linked agammaglobulinaemia but not specific polysaccharide antibody deficiency

Myeloid and plasmacytoid dendritic cells (MDCs, PDCs) play critical roles in B cell development and antibody production. Primary antibody deficiencies in humans might therefore reflect a deficit in MDCs and/or PDCs. We tested this hypothesis by measuring dendritic cell (DC) subset numbers in patients with common variable immunodeficiency (CVID), X-linked agammaglobulinaemia (XLA) and specific polysaccharide antibody deficiency (SPAD). In CVID both MDC and PDC numbers were markedly reduced. There was a graded reduction in all DC subsets across the Freiburg CVID groups (memory B cell classification) and the greatest deficit was seen in group Ia cases with the most severe disease. In contrast, MDC numbers alone were reduced in XLA whilst in SPAD the DC numbers were normal. In CVID, the number of MDCs correlated strongly with switched memory B cell percentage and total B cell count. Low numbers of DCs correlated with a greater incidence of autoimmunity, splenomegaly and granulomatous disease, and a higher incidence of clinical complications. Measurement of MDC and PDC numbers provides both prognostic information for clinical management and classification of CVID cases for future pathogenetic research. Our findings are consistent with the hypothesis that deficits in DC subsets are a critical feature in CVID.     

Streptococcus pyogenes activates human plasmacytoid and myeloid dendritic cells

Human peripheral blood contains two major dendritic cell (DC) populations, namely CD11c(-)CD123+ plasmacytoid DCs (PDCs) and CD11c+CD123(-) myeloid DCs (MDCs). Although the activation of these DC types by various TLR ligands has been relatively well-characterized, less is known about the ability of whole live bacteria to induce PDC and MDC activation. In the present report, we have compared the activation of human PDCs and MDCs in response to major human bacterial pathogen Streptococcus pyogenes (group A streptococci) and influenza A virus. S. pyogenes stimulation resulted in the maturation of both DC types, as evidenced by enhanced expression of costimulatory molecules and production of proinflammatory cytokines and chemokines. Furthermore, S. pyogenes-stimulated PDCs and MDCs activated na?ve CD4+ T cells and enhanced their Th1 cytokine production. Influenza A virus infection induced rapid PDC activation, whereas MDCs were extremely sensitive to influenza A virus-induced cell death. The most significant differences between DC types were seen in the production of IL-10 and IL-12, which were only produced by S. pyogenes-stimulated MDCs. Although S. pyogenes was able to induce PDC activation, only influenza A virus infection resulted in detectable IFN-alpha production. Our results show that depending on the infecting microbe, the functions of PDCs and MDCs may be partially overlapping, suggesting a considerable flexibility of the human DC system.     

Engagement of human monocyte-derived dendritic cells into interleukin (IL)-12 producers by IL-1beta + interferon (IFN)-gamma

Dendritic cells (DCs) are potent antigen-presenting cells and can induce tumour- or pathogen-specific T cell responses. For adoptive immunotherapy purposes, immature DCs can be generated from adherent monocytes using granulocyte macrophage colony stimulating factor (GM-CSF) and interleukin (IL)-4, and further maturation is usually achieved by incubation with tumour necrosis factor (TNF)-alpha. However, TNF-alpha-stimulated DCs produce low levels of IL-12. In this study, we compared the effects of TNF-alpha, interferon (IFN)-gamma, IL-1beta or IFN-gamma + IL-1beta on the phenotypic and functional maturation of DCs. Our results show that IFN-gamma, but not IL-1beta, augmented the surface expression of CD80, CD83 and CD86 molecules without inducing IL-12 production from DCs. However, IL-1beta, but not IFN-gamma, induced IL-12 p40 production by DCs without enhancing phenotypic maturation. When combined, IFN-gamma + IL-1beta treatment profoundly up-regulated the expression of CD80, CD83, CD86 and major histocompatibility complex (MHC) class II antigens. Furthermore, IFN-gamma + IL-1beta-treated DCs produced larger amounts of IL-12 and induced stronger T cell proliferation and IFN-gamma secretion in primary allogeneic mixed lymphocyte reaction (MLR) than did TNF-alpha-treated DCs. Our results show that IFN-gamma + IL-1beta induced human monocyte-derived DCs to differentiate into Th1-prone mature DCs.     

The effect of conditional EFNB1 deletion in the T cell compartment on T cell development and function

Background

Mannan-binding lectin (MBL) is a pattern-recognition molecule present in serum, which is involved in the innate immune defense by activating complement and promoting opsonophagocytosis. Dendritic cells (DCs) are professional antigen presenting cells (APCs) that are crucial for the initiation of adaptive immunity. Lipopolysaccharide (LPS) has been shown to be a strong activator of the inflammatory response and immune regulation. We first examined whether MBL modulated LPS-induced cellular responses, then investigated possible mechanisms of its inhibitory effect.

Results

MBL at higher concentrations (10-20 μg/ml) significantly attenuated LPS-induced maturation of monocyte-derived DCs (MDCs) and production of proinflammatory cytokines (e.g., IL-12 and TNF-α), and inhibited their ability to activate allogeneic T lymphocytes. It bound to immature MDCs at physiological calcium concentrations, and was optimal at supraphysiological calcium concentrations. MBL also bound directly to immature MDCs and attenuated the binding of LPS to the cell surfaces, resulting in decreased LPS-induced nuclear factor-κB (NF-κB) activity in these cells.

Conclusion

All these data suggest that MBL could affect the functions of DCs by modifying LPS-induced cellular responses. This study supports an important role for MBL in the regulation of adaptive immune responses and inflammatory responses.     

Complement protein C1q induces maturation of human dendritic cells

Maturation of dendritic cells (DCs) is known to be induced by several stimuli, including microbial products, inflammatory cytokines and immobilized IgG, as demonstrated recently. Since immune complexes formed in vivo also contain C1q, moreover apoptotic cells and several pathogens fix C1q in the absence of antibodies, we undertook to investigate whether this complement protein has an impact on various functions of human DCs. Maturation of monocyte-derived immature DCs (imMDCs) cultured on immobilized C1q was followed by monitoring expression of CD80, CD83, CD86, MHCII and CCR7. The functional activity of the cells was assessed by measuring cytokine secretion and their ability to activate allogeneic T lymphocytes. Cytokine production by T cells co-cultured with C1q-matured DCs was also investigated. C1q, but not the structurally related mannose-binding lectin was found to bind to imMDC in a dose-dependent manner and induced NF-kappaB translocation to the nucleus. Immobilized C1q induced maturation of MDCs and enhanced secretion of IL-12 and TNF-alpha, moreover, elevated their T-cell stimulating capacity. As IFN-gamma levels were increased in supernatants of MDC-T cell co-cultures, our data suggest that C1q-induced DC maturation generates a Th1-type response. Interestingly, IL-10 levels were elevated by C1q-treated MDCs but not in the supernatant of their co-cultures with allogeneic T cells. Taken together, these results indicate that C1q-opsonized antigens may play a role in the induction and regulation of immune response. Moreover our data are relevant in view of the role of C1q in removal of apoptotic cells and the association between C1q-deficiency and autoimmunity.     

Altered phenotype and function of blood dendritic cells in multiple sclerosis are modulated by IFN-beta and IL-10.

Multiple sclerosis (MS) is assumed to result from autoaggressive T cell-mediated immune responses, in which T helper type 1 (Th1) cells producing cytokines, e.g. IFN-gamma and lymphotoxin promote damage of oligodendrocyte-myelin units. Dendritic cells (DCs) as potent antigen presenting cells initiate and orchestrate immune responses. Whether phenotype and function of DCs with respect to Th1 cell promotion are altered in MS, are not known. This study revealed that blood-derived DCs from MS patients expressed low levels of the costimulatory molecule CD86. In addition, production of IFN-gamma by blood mononuclear cells (MNCs) was strongly enhanced by DCs derived from MS patients. IFN-beta and IL-10 inhibited the costimulatory capacity of DCs in mixed lymphocyte reaction (MLR) and showed additive effects on suppression of IL-12 production by DCs. Correspondingly, DCs pretreated with IFN-beta and IL-10 significantly suppressed IFN-gamma production by MNCs. IFN-beta in vitro also upregulated CD80 and, in particular, CD86 expression on DCs. In vitro, anti-CD80 antibody remarkably increased, while anti-CD86 antibody inhibited DC-induced IL-4 production in MLR. We conclude that DC phenotype and function are altered in MS, implying Th1-biased responses with enhanced capacity to induce Th1 cytokine production. In vitro modification of MS patients' DCs by IFN-beta and IL-10 could represent a novel way of immunomodulation and of possible usefulness for future immunotherapy of MS.     

Probiotic modulation of dendritic cell function is influenced by ageing

Dendritic cells (DCs) are critical for the generation of T-cell responses. DC function may be modulated by probiotics, which confer health benefits in immunocompromised individuals, such as the elderly. This study investigated the effects of four probiotics, Bifidobacterium longum bv. infantis CCUG 52486, B. longum SP 07/3, Lactobacillus rhamnosus GG (L.GG) and L. casei Shirota (LcS), on DC function in an allogeneic mixed leucocyte reaction (MLR) model, using DCs and T-cells from young and older donors in different combinations. All four probiotics enhanced expression of CD40, CD80 and CCR7 on both young and older DCs, but enhanced cytokine production (TGF-β, TNF-α) by old DCs only. LcS induced IL-12 and IFNγ production by DC to a greater degree than other strains, while B. longum bv. infantis CCUG 52486 favoured IL-10 production. Stimulation of young T cells in an allogeneic MLR with DC was enhanced by probiotic pretreatment of old DCs, which demonstrated greater activation (CD25) than untreated controls. However, pretreatment of young or old DCs with LPS or probiotics failed to enhance the proliferation of T-cells derived from older donors. In conclusion, this study demonstrates that ageing increases the responsiveness of DCs to probiotics, but this is not sufficient to overcome the impact of immunosenescence in the MLR.     

胰腺癌患者树突状细胞诱导I型调节性T细胞的特征及其意义

为了检测胰腺癌患者树突状细胞(dendritic cells,DC)诱导I型调节性T细胞(typeⅠregulatory T cells,Tr1)的特征、功能及其临床意义。采用外周血单核细胞来源的未成熟DC,诱导同种异体初始T细胞分化为Tr1,ELISA、流式细胞仪检测Tr1细胞因子表达水平。用混合淋巴细胞反应(mixed lymphocyte reaction,MLR)检测Tr1的免疫抑制功能。结果:经胰腺癌患者DC诱导分化的Tr1分泌IL-10(P<0.05)和TGF-β(P<0.01)水平高于正常对照组。IL-10胞内染色结果也表明,分泌IL-10的Tr1在胰腺癌组明显增加(P<0.01)。胰腺癌患者DC诱导的Tr1抑制MLR增殖的能力也明显增强(P<0.01)。胰腺癌患者DC诱导同种异体Tr1分化的能力明显增强,提示过度Tr1活化可能与胰腺癌的病理形成有关。     

抗CD47单抗对人树突状细胞的免疫调控作用及其机制探讨

目的:研究可溶性的抗CD47单抗(B6H12)对树突状细胞(DC)的免疫调控作用及分子机理。方法:分离人外周血单核细胞,联合应用rhGM-CSF、IL-4、细菌脂多糖(LPS)在体外诱导扩增DC,并在培养体系中添加抗CD47单抗(B6H12)。采用透射电镜观察DC形态,流式细胞仪检测DC膜表面分子,ELISA定量检测DC释放IL-12P70水平,Brdu-ELISA 法检测DC刺激同种异型淋巴细胞增殖,凝胶电泳迁移率改变实验(EMSA)检测NF-κB活性。结果:(1)经抗CD47单抗(B6H12)处理的DC,膜表面CD80、CD86、CD1a、CD83、DR表达显著降低(P＜0.05);其释放IL-12P70的水平及刺激同种异型淋巴细胞增殖的能力也显著低于对照组(P＜0.01)。(2) B6H12单抗处理DC组其NF-κB活性显著降低(P＜0.05),且该效应呈剂量依赖性。结论:抗CD47单抗可通过抑制NF-κB活性而影响DC的发育、成熟及功能。     

人外周血分离树突状细胞前体分离培养及其分化成熟过程中生物特征的变化

用从人外周血分离、纯化、扩增树突状细胞的方法,对树突状细胞成熟过程中的功能分化作初步探讨.以酵母菌、HRP吞噬实验测试树突状细胞的内吞能力;以同种混合淋巴细胞反应检测细胞的免疫刺激能力.结果显示,在GM-CSF或GM-CSF与IL-4联合培养2～3天后,树突状细胞无或仅有极弱的吞噬酵母菌能力,但能活跃内吞HRP.培养5天的树突状细胞内吞HRP能力减弱.但能有效的刺激T细胞增殖,GM-CSF与IL-4联合作用后的树突状细胞,其内吞HRP能力及刺激T细胞能力均强于单用GM-CSF培养的树突状细胞.由此可见,树突状细胞在分化成熟的不同阶段表现出不同的功能,GM-CSF与IL-4联合培养能明显增强树突状细胞功能.     

Dendritic cells contribute to the development of atopy by an insufficiency in IL-12 production.

BACKGROUND: IL-12 is a crucial factor in the development and course of allergic diseases. By virtue of their IL-12 production, dendritic cells (DCs) are potent inducers of T(H)1 responses. However, distinct subsets of DCs have also been shown to induce T(H)2 differentiation. OBJECTIVE: We hypothesized that DCs from atopic and nonatopic individuals might differ in their propensity to skew T-cell responses to either the T(H)1 type or the T(H)2 type. To this end, we investigated the cytokine patterns produced by DCs from atopic and nonatopic individuals, and we attempted to clarify whether this could be due to different DC lineages or, alternatively, to different microenvironmental factors. METHODS: DCs were generated from lymphocyte-depleted PBMCs from atopic and nonatopic donors and fully matured with monocyte-conditioned medium. Production of IL-4, IL-5, IL-10, IL-12, and IL-13 in response to CD40 ligation was measured with ELISA. DC subsets were identified in PBMCs from freshly drawn blood by 3-color flow cytometry. RESULTS: Compared with DCs from healthy donors, monocyte-derived DCs from atopic patients produced less bioactive IL-12 and IL-10. DC production of IL-4, IL-13, and IL-5 was not detected. Relatively more CD123(+) DCs, corresponding to T(H)2-inducing "DC2s," were found in PBMCs from atopic patients. CONCLUSION: The data suggest that in addition to the described abnormalities in the patients' T-cell populations, DCs might also critically contribute to the atopic/allergic T(H)1 outcome in the patient and thus to the disease.     

Cord blood CD34+ cells cultured with FLT3L, stem cell factor, interleukin-6, and IL-3 produce CD11c+CD1a-/c- myeloid dendritic cells

Methods that allow expansion of myeloid dendritic cells (MDCs) from CD34(+) cells are potentially important for boosting anti-leukemic responses after cord blood (CB) hematopoietic stem cell transplantation (HSCT). We showed that the combination of early-acting cytokines FLT3-ligand (FL), stem cell factor (SCF), interleukin (IL)-3, and IL-6 supported the generation of CD11c(+)CD16() CD1a()/c() MDCs from CB CD34(+) cells or CB myeloid precursors. Early-acting cytokine-derived MDCs were maintained within the myeloid CD33(+)CD14()CD15() precursors with a mean of 4 x 10(6) cells generated from 1-4 x 10(4) CB CD34(+) cells or myeloid precursors after 2 weeks. After 8-12 days of culture the MDCs expressed higher levels of HLA-DR antigen but lower levels of CD40 and CD86 antigen, compared to adult blood MDCs. At this stage of differentiation, the early-acting cytokine-derived MDCs had acquired the ability to induce greater allogeneic T cell proliferation than monocytes or granulocytes derived from same culture. Early-acting cytokine-derived MDCs exposed to the cytokine cocktail (CC) comprising IL-1beta, IL-6, tumor necrosis factor (TNF)-alpha, and prostaglandin E (PGE)-2, upregulated the surface co-stimulatory molecules CD40 and CD86 and enhanced allogeneic T cell proliferation, as is characteristic of MDCs maturation. The reliable production of MDCs from CB CD34(+) cells provides a novel way to study their lineage commitment pathway(s) and also a potential means of enriching CB with MDCs to improve prospects for DC immunotherapy following CB HSCT.     

Milk-derived GM(3) and GD(3) differentially inhibit dendritic cell maturation and effector functionalities

Gangliosides are complex glycosphingolipids, which exert immune-modulating effects on various cell types. Ganglioside GD(3) and GM(3) are the predominant gangliosides of human breast milk but during the early phase of lactation, the content of GD(3) decreases while GM(3) increases. The biological value of gangliosides in breast milk has yet to be elucidated but when milk is ingested, dietary gangliosides might conceptually affect immune cells, such as dendritic cells (DCs). In this study, we address the in vitro effect of GD(3) and GM(3) on DC effector functionalities. Treatment of bone marrow-derived DCs with GD(3) before lipopolysaccharide-induced maturation decreased the production of interleukin-6 (IL-6), IL-10, IL-12 and tumor necrosis factor-alpha as well as reduced the alloreactivity in mixed leucocyte reaction (MLR). In contrast, only IL-10 and IL-12 productions were significantly inhibited by GM(3,) and the potency of DCs to activate CD4(+) cells in MLR was unaffected by GM(3). However, both gangliosides suppressed expression of CD40, CD80, CD86 and major histocompatibility complex class II on DCs. Because GD(3) overall inhibits DC functionalities more than GM(3), the immune modulating effect of the ganglioside fraction of breast milk might be more prominent in the commencement of lactation during which the milk contains the most GD(3).     

Maturation of murine bone marrow-derived dendritic cells with poly(I:C) produces altered TLR-9 expression and response to CpG DNA

Although poly(I:C) and LPS induced differential dendritic cell (DC) cytokine profiles and toll-like receptor (TLR) expression, all were capable of causing phenotypic and functional DC maturation. Both LPS and poly(I:C) downregulated TLR-4/MD-2 expression on DCs. Although poly(I:C) highly upregulated their cell surface TLR-9 expression, LPS upregulated the intracellular TLR-9 expression. LPS-treated DCs could not produce IL-12p70 in response to subsequent both LPS- and CpG DNA-stimulation. On the other hand, poly(I:C)-treated DCs retained to produce IL-12p70 by subsequent CpG DNA-stimulation, while subsequent LPS-stimulation did not induce IL-12p70 production. Chloroquine, inhibitor of endosomal maturation, completely inhibited cytokine production of LPS-treated DCs as well as unstimulated control in response to subsequent CpG DNA-stimulation, while it failed to delete the IL-12p40 and IL-10 production in poly(I:C)-treated DCs. These data suggest that poly(I:C) may induce a novel DC phenotype that preserves the capacity of cytokine production to subsequent CpG DNA-stimulation.     

糖基化终产物对人单核细胞源树突状细胞糖基化终产物受体表达的研究

目的：探讨糖基化终产物 (AGEs）)对人单核细胞源树突状细胞（MDCs）糖基化终产物受体（RAGE） 表达的影响。 方法： 用免疫磁珠分离人外周血CD14+单核细胞，经含rhGM-CSF（100 μg/L）和rhIL-4（50 μg/L）的RPMI-1640培养，使其分化为MDCs，采用RT-PCR和Western blotting法，观察糖基化-白蛋白（AGE-BSA）对MDCs RAGE mRNA和蛋白表达的影响，同时检测培养液上清中IFN-γ和IL-12的浓度。 结果： AGE-BSA诱导DCs RAGE mRNA和蛋白的表达（P＜0.05），高于空白对照组，并且明显促进了DCs IFN-γ和IL-12的分泌（P＜0.05）。BSA干预组与空白对照组相比差异无显著（P＞0.05）。 结论： AGEs能够上调DCs RAGE的表达，并且促进了DCs IFN-γ和IL-12的分泌，这可能是糖尿病通过DCs促进动脉粥样硬化发生的重要机制之一。