慢性乙肝患者树突状细胞对Th1/Th2分化的影响

探讨慢性乙肝患者树突状细胞(dendritic cells,DC)对CD4+Th细胞亚群分化的影响。分离慢性乙肝患者外周血单个核细胞(PBMC),以rhIL-4(50 ng/ml)、rhGM-CSF(10 ng/ml)和rhTNF-α(100 u/ml)诱导培养DC。以流式细胞仪检测DC表面CD1a、CD83、CD80、CD86、HLA-DR分子表达情况。MTT法检测DC刺激同种异体淋巴细胞增殖能力。免疫磁珠分离外周血CD4+T细胞亚群,PMA+Ionomycin刺激后胞内荧光染色,流式细胞仪检测辅助性T细胞(helper T cell,Th)内特征性细胞因子IFN-γ/IL-4以判断Th1/Th2分化。ELISA法检测DC或Th细胞培养上清中IL-6、IL-12、IFN-γ和IL-4的含量。结果:慢性乙肝患者的DC表达CD1a、CD83、CD80、CD86、HLA-DR分子水平明显低于正常人(P<0.01);培养至第7天,慢性乙肝患者DC分泌的IL-12水平低于正常人(P<0.01),而分泌的IL-6水平增高(P<0.05)。与正常人相比,慢性乙肝患者外周血中Th1细胞占CD4+T细胞的百分比较低(P<0.01),其Th细胞培养上清中IFN-γ的量也较低(P<0.01)。患者DC与同种异体的健康人Th细胞共培养,刺激Th1型细胞因子IFN-γ产生的能力低于正常人(P<0.01)。慢性乙肝患者体内DC功能的异常可能导致了外周血Th1细胞分化不足。     

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。     

负载滋养层细胞抗原对小鼠树突状细胞表型及功能的影响

目的 研究负载滋养层细胞抗原对小鼠髓源性树突状细胞(DC)分化成熟过程的影响,获得致耐受性DC.方法 体外使用粒细胞巨细胞集落刺激因子(GM-CSF)诱导小鼠骨髓细胞定向分化、经LPS刺激获得成熟DC;通过外胎盘锥组织块培养法获得滋养层细胞,制备可溶性抗原,加入DC培养体系.流式细胞术检测DC表面共刺激分子及MHC-Ⅱ的表达,ELISA法检测DC分泌IL-10和IL-12的浓度,混合淋巴细胞培养评估 DC刺激同种T细胞增殖、活化的功能.结果 成熟DC表型为CD40high CD80highCD86highMHC-Ⅱhigh,分泌大量的IL-12和极少量的IL-10 ,体外能有效刺激T细胞的增殖;负载滋养层细胞抗原的DC表型为CD40midCD80lo wCD86lowMHC-Ⅱlow,在分泌大量IL-12的同时IL-10也明显升高,不能有效刺激T细胞增殖,并使T细胞分泌细胞因子呈现明显Th2偏倚.结论 负载滋养层细胞抗原后的DC表面共刺激分子及MHC-Ⅱ表达降低,刺激T细胞增殖能力下降;其自分泌和促使T细胞旁分泌的细胞因子呈现Th2偏倚,是一种耐受性DC.     

天花粉蛋白及其肽段诱导的免疫抑制依赖APC表面Notch配体的表达

新近发现,Notch信号途径参与调节外周成熟T细胞及其亚群的分化和功能发挥。本研究应用天花粉蛋白及其衍生肽处理骨髓来源的小鼠树突状细胞(DC),检测Notch配体家族分子的表达及DC对CD8+T细胞分泌细胞因子的影响。结果表明,天花粉蛋白或其衍生肽PB处理DC可使Notch配体Jagged1、Delta1分子表达明显增加,并改变CD8+T细胞细胞因子分泌格局,明显抑制Th1相关细胞因子IFN-γ的分泌,而Th2相关细胞因子IL-4和IL-10分泌明显增加。Notch信号的阻断剂可以部分逆转Tk及肽段的抑制作用。表明天花粉蛋白及其衍生肽可诱导一群具有抑制能力的CD8+T细胞,该作用依赖于DC表面Notch配体的表达。     

HBeAg对小鼠骨髓源性树突状细胞成熟的影响

目的 探讨HBe Ag对LPS诱导小鼠骨髓源性树突状细胞(DC)成熟的影响。方法 体外诱导C57BL/6小鼠骨髓细胞分化成未成熟树突状细胞,经CD11c磁珠分选纯化后将DCs随机分为空白对照组、LPS刺激组、HBe Ag+LPS刺激组。流式检测DC表型变化,混合淋巴反应(MLR)检测DC促T淋巴细胞增殖能力,酶联免疫法(ELISA)检测细胞上清液中IL-12的分泌水平,Western blot检测p38磷酸化水平,并设置SB203580组为阳性对照探讨细胞IL-12分泌的可能调节机制。结果 LPS刺激未成熟DC引起细胞表面MHC-Ⅱ、CD86表达升高,刺激同种异体淋巴细胞增殖能力增强,IL-12分泌量增高。HBe Ag可抑制LPS促进DC表面MHC-Ⅱ、CD86表达升高和促淋巴细胞增殖能力增强的作用。LPS刺激DC可引起p38磷酸化水平升高,并呈时间依赖性;HBe Ag或SB203580预处理细胞再予LPS刺激,磷酸化p38表达和IL-12分泌较单纯LPS刺激组明显下降。结论 HBe Ag对LPS引起的树突状细胞的成熟有一定的抑制作用,且HBe Ag可能通过抑制p38MAPK信号通路下调LPS诱导的树突状细胞IL-12的产生。     

Graves病患者外周血树突状细胞和CD4+CD25+CD127low/-Tr细胞的变化

目的 观察Graves病患者外周血树突状细胞(dendritic cells,DC)和CD4+ CD25+CD127low/-调节性T细胞(Tr)在疾病过程中的水平变化.方法 根据临床表现以及血清游离甲功三项(FT3、FT4、TSH),将患者分为治疗前组、临床缓解组和稳定组,并设正常对照组.采用流式细胞术测定研究对象外周血(EDTA-K2抗凝)中DC及髓样DC(myeloid dendritic cell,MDC)和浆细胞样DC(plasmacytoid dendritic cell,PDC)两个亚群以及CD4+CD25+CD127low/-Tr细胞占CD4+T细胞的百分比,观察它们在病程中的变化,再进一步探讨检测指标与游离甲功三项的相关性.结果 (1)治疗前组、临床缓解组、稳定组和正常对照组的总DC、MDC和MDC/PDC逐渐下降,各组间比较均有统计学差异;(2)治疗前组、临床缓解组、稳定组和正常对照组的PDC依次呈下降趋势,治疗前组仅与正常对照组有统计学差异;(3)治疗前组、临床缓解组、稳定组和正常对照组的CD4+ CD25+ CD127low/-Tr细胞逐渐升高,治疗前组仅与正常对照组有统计学差异;(4)治疗前组PDC和CD4+CD25+CD127kw/-Tr细胞有相关性;(5)在治疗前组,DC及其亚群与游离甲功三项有相关性,而CD4+CD25+CD127low/-Tr细胞只与FT3和FT4有相关性.结论 Graves病患者总DC、MDC、MDC/PDC比值明显增高,抗甲亢治疗随着病情好转而下降,因此DC及其亚群的检测有望用来监测Graves病患者病情;而CD4+CD25+CD127low/-Tr细胞可作为Graves病发病的新指标.     

IL-10、TNF-α对慢性乙型肝炎患者树突状细胞的影响

目的 研究IL-10和TNF-α对慢性乙型肝炎患者树突状细胞的分化与功能的影响.方法 健康志愿者和慢性乙型肝炎患者各15例,取外周静脉血,分离获得外周血单个核细胞(PBMC),常规培养获得DC组.培养期间用IL-10刺激的为imDC组;TNF-α刺激的为mDC组;先后用IL-10、TNF-α刺激的为imDC+TNF-α组.各组收获细胞后分别检测反映DC成熟度的各种分子及其体外诱导的自体淋巴细胞增殖效应.结果 源于慢性乙型肝炎患者的常规培养的Dc在数量,HLA-DR、HLA-A、B、C和CD86表达,IL-12p70分泌及其诱导的淋巴细胞增殖效应,均低于健康者组(P＜0.01).源于慢性乙型肝炎患者的mDC与健康者mDC表达的HLA-DR、HLA-A、B、C和CD86比较无显著性差异;2组mDC诱导自体淋巴细胞增殖效应和分泌的IL12p70均明显增加,比较有显著性差异(P＜0.01);2组imDC的HLA-DR、HLA-A、B、C和CD86低表达,分泌的IL-12p70极低,不能强烈激活诱导淋巴细胞增殖效应,与DC组比较有显著性差异(P＜0.01).加入TNF-α刺激后HLA-A、B、C,HLA-DR,CD86仍低表达,分泌的IL-12pT0和淋巴细胞增殖效应仍较低.健康对照组、慢性乙型肝炎组的上清夜中IL-21的ELISA检测值均较低,无显著性差异.结论慢性乙型肝炎患者常规培养的DC在数量和功能上低于健康者.TNF-α可促进常规培养的DC成熟为mDC、IL-10可抑制DC成熟.慢性乙型肝炎患者的DC同健康者一样可进行有效的调控,分化成熟后功能强于健康者常规培养的DC,或可提高自体DC治疗性疫苗在不同免疫状态下的治疗效果.     

IL-12基因修饰DC中SOCS1基因沉默对体外诱导特异性CTL的影响

利用重组腺病毒载体pAd CMV/V5-DEST-IL-12转染小鼠骨髓来源的树突状细胞(IL-12/DC),探讨SOCS1基因沉默IL-12/DC在体外诱导细胞毒性T淋巴细胞(CTL)的效能,及其免疫杀伤肺癌细胞株LLC的能力。采用重组腺病毒介导IL-12基因和SOCS1SiRNA基因共同修饰C57BL/6小鼠骨髓来源的DC,经反复冻融法提取LLC抗原,致敏基因修饰的DC;用ELISA法检测各组DC分泌IL-12和IL-10的水平,及各组DC刺激后的T细胞分泌IFN-γ的水平;MTT法检测DC刺激同源小鼠T细胞的增殖能力,微量细胞毒法检测CTL的活性并收集刺激后的T细胞,流式细胞术分析CD8+/CD4+比例和CD4+CD25+Treg的水平;统计学分析各组间的差异。SOCS1SiRNA和IL-12基因共同修饰能有效下调DC中SOCS1蛋白的表达并上调IL-12蛋白的表达;IL-12的分泌水平也明显高于SOCS1SiRNA或IL-12单基因转染组;基因共同修饰的DC表型更加成熟,能明显促进CTL的增殖和活化,减少Treg的生成;CTL分泌高水平的IFN-γ,产生对LLC特异性的细胞免疫。     

白细胞介素-18干预诱导的树突状细胞表型和活性研究

目的：研究白细胞介素-18（IL-18）干预诱导的树突状细胞（DC）的表型和活性。方法:自人外周血单核细胞诱导DC，第5 d起分为IL-18组、TNF-α组和IL-18+TNF-α组，分别加IL-18、TNF-α及IL-18+TNF-α促成熟，用ELISA法测定上清中IL-12含量；用流式细胞仪测定培养8 d DC的CD1a、HLA-DR、CD83及CD86的表达；用MTT法检测3组DC诱导T细胞增殖的作用。用ELISA法测定3组DC刺激T细胞分泌干扰素γ(IFN-γ)的量。结果:IL-18组与TNF-α组CD1a、HLA-DR、CD83及CD86表达无差异，IL-18+TNF-α组CD1a、CD83及HLA-DR阳性率高于IL-18组。IL-18+TNF-α组IL-12量高于IL-18组和TNF-α组(P＜0.05)。IL-18组与TNF-α组DC刺激T细胞增殖作用无差异，IL-18+TNF-α组DC的作用强于IL-18组和TNF-α组。IL-18组和TNF-α组IFN-γ量无显著差异，IL-18+TNF-α组IFN-γ的量高于IL-18组和TNF-α组(P＜0.05)。结论:IL-18干预诱导的DC高表达表面分子，具有明显的免疫刺激活性，IL-18与 TNF-α合用作用更强。     

IL-37通过作用树突状细胞调节CD8~+T细胞功能

将小鼠骨髓细胞诱导分化为DC,用IL-37处理后,流式细胞术检测细胞表面共刺激分子(CD86、CD80),RT-PCR法检测TNF-α、IFN-γ、IL-6、IL-12和TGF-βmRNA的表达,多重液相蛋白定量CBA试剂盒及ELISA试剂盒检测细胞上清中TNF-α、IL-6、IL-12、IFN-γ、MCP-1和TGF-β蛋白的表达。Western blotting方法检测DC中磷酸化蛋白的表达。将DC与T细胞共培养,流式细胞术检测CD8+T细胞增殖及活化程度(CFSE、CD69)。结果显示,IL-37能够降低表达CD80+/CD86+的DC数量。促炎因子TNF-α、IL-12和IL-6的表达被明显抑制,而T淋巴细胞抑制因子TGF-β明显增高。IL-37预处理的DC明显降低T淋巴细胞的增殖和活化能力。IL-37能够降低DC中磷酸化ERK和NF-κB的表达。IL-37处理的DC对CD8+T细胞产生明显抑制作用。结果表明,IL-37能够通过影响ERK和NF-κB依赖的信号通路抑制DC的成熟和免疫反应,从而抑制CD8+T细胞的活化及增殖。     

Toll-like receptor expression reveals CpG DNA as a unique microbial stimulus for plasmacytoid dendritic cells which synergizes with CD40 ligand to induce high amounts of IL-12

Human plasmacytoid dendritic cells (DC) (PDC, CD123+) and myeloid DC (MDC, CD11c+) may be able to discriminate between distinct classes of microbial molecules based on a different pattern of Toll-like receptor (TLR) expression. TLR1-TLR9 were examined in purified PDC and MDC. TLR9, which is critically involved in the recognition of CpG motifs in mice, was present in PDC but not in MDC. TLR4, which is required for the response to LPS, was selectively expressed on MDC. Consistent with TLR expression, PDC were susceptible to stimulation by CpG oligodeoxynucleotide (ODN) but not by LPS, while MDC responded to LPS but not to CpG ODN. In PDC, CpG ODN supported survival, activation (CD80, CD86, CD40, MHC class II), chemokine production (IL-8, IP-10) and maturation (CD83). CD40 ligand (CD40L) and CpG ODN synergized to activate PDC and to stimulate the production of IFN-alpha and IL-12 including bioactive IL-12 p70. Previous incubation of PDC with IL-3 decreased the amount of CpG-induced IFN-alpha and shifted the cytokine response in favor of IL-12. CpG ODN-activated PDC showed an increased ability to stimulate proliferation of naive allogeneic CD4 T cells, butTh1 polarization of developing T cells required simultaneous activation of PDC by CD40 ligation and CpG ODN. CpG ODN-stimulated PDC expressed CCR7, which mediates homing to lymph nodes. In conclusion, our studies reveal that IL-12 p70 production by PDC is under strict control of two signals, an adequate exogenous microbial stimulus such as CpG ODN, and CD40L provided endogenously by activated T cells. Thus, CpG ODN acts as an enhancer of T cell help, while T cell-controlled restriction to foreign antigens is maintained.     

Combined cyclosporin-A /prednisone therapy of patients with active uveitis suppresses IFN-gamma production and the function of dendritic cells

In this study, we assessed the Th1/Th2 polarization of the immune response and the involvement of dendritic cells (DC) and Th1 lymphocytes in the pathogenesis of uveitis. Thirty-seven patients with chronic idiopathic uveitis were enrolled: 21 of them had active uveitis and the remaining 16 were in complete remission. Patients with active uveitis were characterized as follows: 5 had intermediate uveitis, 5 panuveitis and the remaining 11 posterior uveitis. Thirteen healthy subjects were also studied as controls. Patients with active uveitis were treated with cyclosporin-A (CsA) associated to low doses of prednisone (PDS) and studied at baseline and after 6 months of therapy. Analysis of cytokine-producing CD3+ lymphocytes revealed a strong Th1 polarization of the immune response in patients with active uveitis. Th1 lymphocytes paralleled serum IL-12 levels and the response to therapy, which greatly reduced both IFN-gamma+/CD3+ lymphocytes and serum IL-12 levels, associated with a general clinical improvement. In vitro studies demonstrated that DC from untreated patients with active uveitis were mature and functionally active. In fact, they showed a higher ability to stimulate cell proliferation of allogeneic T cells in primary mixed lymphocyte reaction (MLR) and produced larger amounts of IL-12 than DC from CsA/PDS-treated patients and those in remission. These results demonstrate that CsA/PDS therapy impairs the capacity of mature DC to secrete IL-12 and inhibits their MLR activity.     

Decreased blood dendritic cell counts in type 1 diabetic children

In this study DC numbers, phenotype and DC responses to the Toll-like receptor (TLR)-3 ligand, poly I:C, were examined in new-onset Type 1 diabetes (T1D) patients (ND) and in established T1D patients (ED). Absolute blood myeloid DC (MDC) and plasmacytoid DC (PDC) numbers were decreased in ND and ED patients compared to age-matched controls. The decrease in MDC and PDC counts was less evident in patients with a combination of T1D and coeliac disease (CD) or CD alone. The age-dependent decline in blood DC numbers, found in control children, was not evident in ND patients, such that 2-10 years old ND children had similar MDC and PDC numbers to 15-17 years old controls. In ED patients the t-score of MDC and PDC numbers related to the age of diagnosis but not to disease duration. Blood DC in T1D patients were not distinguished from those of controls by the levels of HLA-DR, CD40 and CD86 expression or the percentage of DC expressing cytokines, IL-12, IL-10, IL-6 and TNF-alpha, in responses to poly I:C. If low DC numbers are shown to contribute to the autoimmunity in T1D, interventions aimed to increase DC numbers may mitigate against beta-cell loss.     

调节性DC分泌的外体诱导免疫耐受功能的体外研究

为了探讨树突状细胞(DC)分泌的外体(Dex)在诱导T细胞免疫耐受中的作用,体外研究采用供体Dex降低同种异体移植排斥的可能性。从正常人外周血单个核细胞中诱导培养未成熟DC(imDC),用TGF-β1联合IL-10诱导调节性DC,LPS诱导DC成熟。采用流式细胞术方法观察TGF-β1和IL-10对DC表型、吞噬功能的影响;采用超速离心和超滤的方法提纯Dex;Western blot方法检测imDC分泌的Dex(imDex)与调节性DC分泌的Dex(rDex)表达的相关分子;通过CCK-8法分析异源iDex和mDex在混合淋巴细胞反应(MLR)中的生物学功能,并比较rDex与iDex诱导免疫耐受的能力。结果显示,TGF-β1和IL-10可下调DC表面的共刺激分子CD80、CD83、CD86的表达,并诱导调节性DC分泌更多的rDex;异源的mDC分泌的Dex(mDex)在mDC存在时增强MLR,而异源的imDex在imDC存在时一定程度上抑制MLR,rDex诱导的抑制T细胞增殖作用显著强于iDex;rDex表达更多的FasL,提示TGF-β1和IL-10诱导的调节性DC分泌的rDex在免疫耐受中发挥重要作用,有望应用于同种异体移植抗免疫排斥。     

Direct allorecognition promotes activation of bystander dendritic cells and licenses them for Th1 priming: a functional link between direct and indirect allosensitization

T-cell sensitization to indirectly presented alloantigens (indirect pathway of allorecognition) plays a critical role in chronic rejection. The usual very efficient priming of such self-restricted, T helper type 1 (Th1)-deviated CD4+ T cells obviously conflicts with the fact that allogeneic MHC molecules are poorly immunogenic per se. The aim of the present study is to elucidate whether direct allosensitization induces production of inflammatory mediators that may affect recruitment and activation of immature bystander (host) dendritic cells (DC). These potential mechanisms were studied in vitro by conducting primary allogeneic mixed leucocyte reactions (MLR), mimicking the priming phase in secondary lymphoid organs, and secondary MLR, mimicking the effector phase within the graft. Primary, and particularly secondary, MLR supernatants were found to contain high levels of monocyte/immature DC-recruiting CC chemokines and pro-inflammatory cytokines. Exposure of immature DC to primary or secondary MLR supernatants was found to upregulate CD40 expression and further enhanced lipopolysaccharide-induced interleukin-12 (IL-12) p70 production. Secondary MLR supernatants additionally induced upregulation of CD86 and deviated allogeneic T-cell responses towards Th1 (enhanced interferon-gamma production without concomitant induction of detectable IL-4 or IL-10 production). These findings indicate that direct allorecognition may act as a Th1-deviating adjuvant for indirect allosensitization.     

IL-2通过上调naive CD4+T细胞SOCS-3表达抑制CD4+T细胞的极化和增殖

目的 探讨IL-2预孵育naive CD4+T细胞后,对其极化(polarization)方向和增殖(proliferation)能力的影响.方法 用终浓度为50 U/ml的IL-2分别预孵育DOI 1.10 TCR转基因小鼠和C57BL/6N小鼠naYve CD+T细胞,在不同时间点用荧光实时定量PCR(real-time PCR)检测这两种细胞中SOCS-3(suppressor of cytokine signal-3)表达的变化.预孵育4 h后,洗去IL-2,分别加入卵清白蛋白(OVA)和灭活后的BALB/c脾细胞,在存在细胞因子IL-12或IL-4情况下共培养14 d后,用流式细胞仪检测TH1细胞活化和极化的标志IL-12R β1、IL-12Rβ2,对C57BL/6N小鼠nave CD4+2T细胞的极化中还榆测TH2极化的标志--细胞内IL-4的表达;同时将无IL-2预孵育的作为对照组.结果 IL-2预孵育后这两种鼠naive CD4+T细胞内的SOCS-3表达于6 h达高峰;在SOCS-3表达达高峰后分别给予特异性抗原和同种异基因抗原刺激,其向TH1方向的极化和增殖能力都受到明显的抑制(P<0.05).结论 IL-2预孵育naive CD+T细胞后,可以上调SOCS-3的表达;SOCS-3的上调表达可以抑制naive CD4+T细胞接受特异性抗原和同种异基因抗原刺激后向TH1方向的极化和增殖能力.     

Eminent role of ICOS costimulation for T cells interacting with plasmacytoid dendritic cells

CD4+ CD45RO+ T cells could mature freshly isolated human plasmacytoid dendritic cells (PDC) in a superantigen-driven culture in a similar way to recombinant interleukin-3 (IL-3). Mature PDC expressed significantly higher levels of inducible costimulator ligand (ICOS-L), but similar levels of CD80 and CD86, when compared to mature monocyte-derived DC (moDC). We therefore directly compared the capacities of mature PDC and moDC to activate T cells. A similar T helper type 1 (Th1)/Th2 pattern of cytokines was generated in both systems, but significantly higher levels of IL-3, IL-4 and IL-10 were induced by PDC. In T cells interacting with PDC, the ICOS/ICOS-L costimulatory pathway played a pre-eminent role in the generation of IL-3 and IL-10, CD28 was central to the induction of IL-2, and both pathways were equally important for the generation of other cytokines. In cocultures with moDC, the CD28 pathway was dominant over ICOS under all circumstances, except for the ICOS-mediated release of IL-10. In general, our data demonstrate an eminent role of ICOS in the interaction of T cells with PDC, and thus modify the current paradigm of CD28 dominance for the costimulation of T cells interacting with professional antigen-presenting cells. In particular, our data highlight the role of ICOS in the generation of IL-3, a factor central to the biology of human PDC.     

Reciprocal regulation of plasmacytoid dendritic cells and monocytes during viral infection.

Reciprocal regulation of opposing functions characterizes biological systems. We now show that adenovirus-infected plasmacytoid dendritic cells (PDC) inhibit monocyte to myeloid dendritic cell (MDC) differentiation and function, and that adenovirus-infected monocytes inhibit PDC type I interferon secretion. Adenovirus-infected PDC secreted IFN-alpha, beta and omega in an 86:2:1 ratio. PDC type I interferons inhibited MDC differentiation and function (reduced IL-12 secretion, IFN-gamma induction, MLR and CD40 expression, and increased CD1a(+)CD14(+) cells). Type I interferon receptor blocking antibody reversed all PDC effects, and recombinant IFN-alpha, beta or omega replicated all effects, except reduced CD40. Adenovirus-infected monocytes suppressed PDC type I interferon secretion, which was reversed with anti-IL-10 neutralizing antibodies. Exogenous IL-10 suppressed PDC type I interferon secretion without reducing PDC viability. Therefore, monocyte IL-10 regulates PDC type I interferon secretion, and PDC type I interferons inhibit MDC differentiation and function. Such reciprocal regulation of potentially opposing influences may help modulate anti-pathogen immunity.     

TGF-β1诱导的调节性T细胞体内输注延长小鼠皮肤移植物存活

目的 利用TGF-β1体外诱导naYve T细胞分化为凋节性T细胞(Treg),通过体内输注延长小鼠皮肤移植物存活时间,并研究其相关机制. 方法 根据诱导条件不同分为3组:对照组(加入IL-2培养的C57BL/6小鼠T细胞)、MLR组(即混合淋巴细胞反应组,经同种抗原刺激活化的C57BL/6小鼠T细胞)和TGF-β组(经同种抗原刺激活化的C57BL/6小鼠T细胞,同时加入5.0ng/ml TGF-β1诱导).利用FACS检测CD4+CD25+T细胞比例,并用RT-PCR检测Foxp3的表达水平.建立小鼠皮肤移植模型,并于第0、l、2和3天输注上述细胞,观察皮肤移植物存活时间.术后第9天,取部分受鼠行移植物病理检测,用FACS检测脾脏中THl、TH2和CD4+CD25+Treg比例,并用Alamar Blue法检测淋巴细胞增殖能力. 结果 TGF-β组中CD4+CD25+T细胞比例高于对照组和MLR组(P<0.05),且其高表达Foxp3.将培养的细胞输注给受鼠后发现,输注MLR组细胞的受鼠其移植物平均存活时间(mean survival time,MST)为(9.4±1.3)d,低于对照组(P<0.05);而输注TGF-13组细胞小鼠MST为(22.8±1.9)d,较对照组和MLR组明显延长(P<0.05).病理检测亦显示TGF-β组受鼠移植物结构完整,无明显淋巴细胞浸润.FACS结果显示TGF-β组小鼠体内TH1细胞(CD4+TIM-3+)比例低于对照组和MLR组(P0.05),但低于对照组(P<0.05);而且TGF-β组中CD4+CD25+Treg比例明显高于对照组和MLR组(P<0.05).用Alamar Blue法检测受鼠外周淋巴细胞增殖活性显示,TGF-β组受鼠淋巴细胞增殖能力被明显抑制,低于对照组(P<0.05). 结论 TGF-β1可诱导T细胞分化为具有抑制能力的Treg将诱导后的细胞进行过继输注可使小鼠体内CD4+CD25+Treg比例升高,同时抑制TH1和TH2细胞分化扩增,并使得外周淋巴细胞增殖能力减弱,从而延长小鼠皮肤移植物存活时间.