咳嗽变异性哮喘与典型支气管哮喘患者外周血树突状细胞的变化

目的:计算咳嗽变异性哮喘(CVA)、典型支气管哮喘(CA)患者,其外周血中树突状细胞(DCs)及亚型髓样树突状细胞(m DC)、浆细胞样树突状细胞(p DC)比例和数量的变化,了解咳嗽变异性哮喘与典型支气管哮喘之间是否有类似的免疫学发病机制.方法:选取CA患者20例,CVA患者17例,对照组健康人17例,使用流式细胞术检测和计算CA组与CVA组受试者外周血中DCs及其两个亚群:p DC和m DC所占比例和数量的变化.结果:咳嗽变异性哮喘与典型支气管哮喘患者相比,其外周血树突状细胞及其亚型的数量和比例均无明显差异,咳嗽变异性哮喘、典型支气管哮喘患者较健康对照组,p DC数量及比例的增高明显,m DCs/p DCs的比值降低有统计学差异.结论:咳嗽变异性哮喘与支气管哮喘的发病机制在免疫学上类似,由于p DCs比例和数量的增多引起Th2细胞数量增多,造成Th1/Th2平衡向Th2细胞偏移.     

树突状细胞和细菌的相互作用

越来越多的证据表明树突状细胞（DC）在机体针对细菌产生的免疫应答中发挥着极为重要的作用。本书全面地介绍了树突状细胞（DC）在免疫系统中是怎么和入侵机体的细菌相互作用，诱发针对细菌的免疫应答的。     

人树突状细胞的体外培养和鉴定

目的:探索在体外分离培养人外周血树突状细胞的方法,同时为进一步研究树突状细胞工作奠定实验基础.方法:采用Ficoll、Precoll和Panning法分离和纯化人外周血DC,再加GM-CSF(100μg/mL)和IL-4(10μg/mL)诱生出大量树突状细胞,并进行免疫组化鉴定.结果:培养到第3天的DC扫描电镜可见DC伸出的树突状突起,呈毛刺状.培养到第7天的DC扫描电镜可见细胞胞体体积变大.这些毛刺状细胞表面有丰富的呈分叉的树突状胞质突起,某些突起形成薄片状结构,培养后的DC细胞呈S-100蛋白阳性染色.结论:人外周血树突状细胞可以在体外大量培养.     

树突状细胞瘤苗抗肿瘤功能的研究进展

树突状细胞(DC)是目前已知功能最强的专职抗原提呈细胞,DC在机体免疫应答中的作用尤其是其与肿瘤免疫激活、免疫耐受及相关治疗性疫苗的研究正受到高度关注,文章就DC瘤苗的实验与临床研究进展作一论述.     

猪树突状细胞的体外扩增

树突状细胞(DC)是一类专职的抗原提呈细胞,在激发机体初始免疫反应和调节T细胞介导的免疫反应中都发挥十分重要的作用,但至今对猪DC的了解较少.本研究从猪外周血中分离获得单核细胞,分别加入150ng/mLpGM-CSF和100U/mL pIL-4,体外培养6 d后诱导出大量DC,通过显微镜观察可见其细胞表面具有典型的树突状突起,呈毛刺状.猪DC的体外获得将为进一步研究许多病毒性疾病的致病机理奠定坚实的基础.     

健康恒河猴外周血及肠系膜淋巴结中树突状细胞(Dendritic Cell)的亚群及分布特点

树突状细胞(Dendritic Cell,DC)作为天然免疫的重要组成部分,在抗原识别和抗原呈递过程中发挥重要作用.对DC细胞在恒河猴外周血及淋巴结组织中的细胞亚群及其分布频率进行初步观察.取健康源恒河猴外周血和肠系膜淋巴结,从中分离出淋巴细胞.利用流式细胞术检测分析不同亚群的DC细胞在其间的分布.结果表明,外周血及淋巴结中DC细胞比例均较低,但外周血中DC比例略高于肠系膜淋巴结.mDC亚群占总DC细胞数比例高于pDC细胞亚群,而肠系膜淋巴结中mDC比例相较外周血中有明显下降,统计学差异显著.测定了健康源恒河猴各亚群DC细胞在外周血和淋巴组织中的基础数值,为相关模型研究奠定了基础.     

树突状细胞与妇科肿瘤的免疫治疗

树突状细胞(DC)是目前所知机体内功能最强的专职抗原提呈细胞,可在体内外向T细胞提呈抗原并诱发CTL反应,在抗肿瘤免疫中发挥重要作用.近年采用DC疫苗进行抗肿瘤治疗已成为当今肿瘤生物治疗领域备受关注的焦点之一.针对DC抗肿瘤机制、妇科肿瘤的免疫逃逸及在妇科肿瘤上的应用进行了研究.     

自体肝癌细胞裂解物致敏的树突状细胞诱导抗肝癌免疫的体外研究

从术后肝癌病人的外周血中诱导树突状细胞（DC），并经自体肝癌细胞裂解物致敏DC，用流式细胞仪、^3H-TdR掺入法及MTT法检测了DC表面分子的表达、DC刺激T细胞的增殖效应及DC诱导的T细胞对肝癌细胞的杀伤作用，进而比较经自体肝癌细胞裂解物致敏的DC与其它条件下的DC功能的差异．结果显示：肝癌细胞裂解物致敏DC的功能较未致敏DC显著提高，其可诱导自体混合淋巴细胞强的增殖效应，同时诱导的T细胞对自体肝癌细胞有较强的杀伤率．     

大鼠不同成熟状态骨髓源树突状细胞的培养与鉴定

分离、纯化大鼠骨髓单个核细胞,在培养体系中添加重组大鼠粒-巨噬细胞集落刺激因子(rrGMCSF)和重组大鼠白细胞介素(rrIL-4)刺激获得未成熟树突状细胞(imDC),再分别以肿瘤坏死因子-α(TNF-α)和脂多糖(LPS)刺激获得半成熟DC(smDC)和成熟DC(mDC).并对imDC、smDC和mDC的形态、表型、细胞因子分泌情况进行鉴定,检测不同DC对异基因淋巴细胞的激活功能.检测结果符合文献对DC的介定,培养所得为imDC、smDC和mDC.可见,体外利用GM-CSF、IL-4、TNF-α和LPS诱导骨髓单核细胞是获得大鼠3种不同成熟DCs的有效方法.     

人外周血DC细胞的冻存

目的:探讨人外周血DC细胞的冻存方法和意义。方法:通过树突状细胞的分离和培养,抗原标本前处理,抗原的装载,实验分组,细胞冻存等,研究树突状细胞冻存情况。结果:冷冻DC细胞保持了它的特性,但会因冻溶过程中生物特性有所减低。结论:我们在应用冻存DC细胞时应考虑增加输入细胞的量。     

SAMHD1 is the dendritic- and myeloid-cell-specific HIV-1 restriction factor counteracted by Vpx

The primate lentivirus auxiliary protein Vpx counteracts an unknown restriction factor that renders human dendritic and myeloid cells largely refractory to HIV-1 infection. Here we identify SAMHD1 as this restriction factor. SAMHD1 is a protein involved in Aicardi-Goutières syndrome, a genetic encephalopathy with symptoms mimicking congenital viral infection, that has been proposed to act as a negative regulator of the interferon response. We show that Vpx induces proteasomal degradation of SAMHD1. Silencing of SAMHD1 in non-permissive cell lines alleviates HIV-1 restriction and is associated with a significant accumulation of viral DNA in infected cells. Concurrently, overexpression of SAMHD1 in sensitive cells inhibits HIV-1 infection. The putative phosphohydrolase activity of SAMHD1 is probably required for HIV-1 restriction. Vpx-mediated relief of restriction is abolished in SAMHD1-negative cells. Finally, silencing of SAMHD1 markedly increases the susceptibility of monocytic-derived dendritic cells to infection. Our results demonstrate that SAMHD1 is an antiretroviral protein expressed in cells of the myeloid lineage that inhibits an early step of the viral life cycle.     

Vpx relieves inhibition of HIV-1 infection of macrophages mediated by the SAMHD1 protein

Macrophages and dendritic cells have key roles in viral infections, providing virus reservoirs that frequently resist antiviral therapies and linking innate virus detection to antiviral adaptive immune responses. Human immunodeficiency virus 1 (HIV-1) fails to transduce dendritic cells and has a reduced ability to transduce macrophages, due to an as yet uncharacterized mechanism that inhibits infection by interfering with efficient synthesis of viral complementary DNA. In contrast, HIV-2 and related simian immunodeficiency viruses (SIVsm/mac) transduce myeloid cells efficiently owing to their virion-associated Vpx accessory proteins, which counteract the restrictive mechanism. Here we show that the inhibition of HIV-1 infection in macrophages involves the cellular SAM domain HD domain-containing protein 1 (SAMHD1). Vpx relieves the inhibition of lentivirus infection in macrophages by loading SAMHD1 onto the CRL4(DCAF1) E3 ubiquitin ligase, leading to highly efficient proteasome-dependent degradation of the protein. Mutations in SAMHD1 cause Aicardi-Goutières syndrome, a disease that produces a phenotype that mimics the effects of a congenital viral infection. Failure to dispose of endogenous nucleic acid debris in Aicardi-Goutières syndrome results in inappropriate triggering of innate immune responses via cytosolic nucleic acids sensors. Thus, our findings show that macrophages are defended from HIV-1 infection by a mechanism that prevents an unwanted interferon response triggered by self nucleic acids, and uncover an intricate relationship between innate immune mechanisms that control response to self and to retroviral pathogens.     

Soluble CD4 blocks the infectivity of diverse strains of HIV and SIV for T cells and monocytes but not for brain and muscle cells

The CD4 antigen has been subverted as a receptor by the human and simian immunodeficiency viruses (HIV-1, HIV-2 and SIV). Several groups have reported that recombinant, soluble forms of the CD4 molecule (sCD4) block the infection of T lymphocytes by HIV-1, as CD4 binds the HIV envelope glycoprotein, gp120, with high affinity. We now report that sCD4 blocks diverse strains of HIV-1, HIV-2 and SIV, but is less effective for HIV-2. The blocking effect is apparent even after adsorption of virions to CD4 cells. Soluble CD4 prevents HIV infection of T-lymphocytic and myelomonocytic cell lines, but neither sCD4 nor anti-CD4 antibodies inhibit infection of glioma and rhabdomyosarcoma cell lines.     

新型肿瘤细胞膜重组脂质体体外免疫激活作用的实验研究

将肿瘤细胞膜与人工脂质体重组,制备新型重组复合体并探索其体外免疫活性.热处理S180细胞并收集裂解细胞膜,重组于人工脂质体得重组复合体,电镜和Weston杂交鉴定重组复合体.体外培养观察该复合体对小鼠树突状细胞及淋巴细胞的影响,流式细胞仪分析CD11c,CD80,CD86等表面标志,3H-TdR掺入测定淋巴细胞增殖.设立单纯细胞膜碎片及单纯脂质体组为对照.裂解肿瘤细胞膜可以重组复合于人工脂质体,形成稳定的复合物,可刺激DC成熟,促进淋巴细胞增殖,与对照组均有显著差异(P<0.05).肿瘤细胞膜重组脂质体可以显著改善肿瘤的体外免疫原性,为快速的个体化肿瘤免疫治疗提供了新思路.     

Viral infection switches non-plasmacytoid dendritic cells into high interferon producers

Type I interferons (IFN-I) are important cytokines linking innate and adaptive immunity. Plasmacytoid dendritic cells make high levels of IFN-I in response to viral infection and are thought to be the major source of the cytokines in vivo. Here, we show that conventional non-plasmacytoid dendritic cells taken from mice infected with a dendritic-cell-tropic strain of lymphocytic choriomeningitis virus make similarly high levels of IFN-I on subsequent culture. Similarly, non-plasmacytoid dendritic cells secrete high levels of IFN-I in response to double-stranded RNA (dsRNA), a major viral signature, when the latter is introduced into the cytoplasm to mimic direct viral infection. This response is partially dependent on the cytosolic dsRNA-binding enzyme protein kinase R and does not require signalling through toll-like receptor (TLR) 3, a surface receptor for dsRNA. Furthermore, we show that sequestration of dsRNA by viral NS1 (refs 6, 7) explains the inability of conventional dendritic cells to produce IFN-I on infection with influenza. Our results suggest that multiple dendritic cell types, not just plasmacytoid cells, can act as specialized interferon-producing cells in certain viral infections, and reveal the existence of a TLR-independent pathway for dendritic cell activation that can be the target of viral interference.     

HIV-1 Nef intersects the macrophage CD40L signalling pathway to promote resting-cell infection

All primate lentiviruses (HIV-1, HIV-2, SIV) encode Nef proteins, which are important for viral replication and pathogenicity in vivo. It is not known how Nef regulates these processes. It has been suggested that Nef protects infected cells from apoptosis and recognition by cytotoxic T lymphocytes. Other studies suggest that Nef influences the activation state of the infected cell, thereby enhancing the ability of that cell to support viral replication. Here we show that macrophages that express Nef or are stimulated through the CD40 receptor release a paracrine factor that renders T lymphocytes permissive to HIV-1 infection. This activity requires the upregulation of B-cell receptors involved in the alternative pathway of T-lymphocyte stimulation. T lymphocytes stimulated through this pathway become susceptible to viral infection without progressing through the cell cycle. We identify two proteins, soluble CD23 and soluble ICAM, that are induced from macrophages by Nef and CD40L, and which mediate their effects on lymphocyte permissivity. Our results reveal a mechanism by which Nef expands the cellular reservoir of HIV-1 by permitting the infection of resting T lymphocytes.     

HIV-1 Nef inhibits ASK1-dependent death signalling providing a potential mechanism for protecting the infected host cell

In vivo infection of lymphatic tissues by the human immunodeficiency virus type 1 (HIV-1) leads to enhanced apoptosis, which prominently involves uninfected bystander cells. Increased killing of such bystander cells is mediated in part through Nef induction of Fas ligand (FasL) expression on the surface of the virally infected T cells. The subsequent interaction of FasL with Fas (CD95) displayed on neighbouring cells, including HIV-1-specific cytotoxic T lymphocytes, may lead to bystander cell killing and thus forms an important mechanism of immune evasion. As HIV-1 also enhances Fas expression on virally infected cells, it is unclear how these hosts avoid rapid cell-autonomous apoptosis mediated through cis ligation of Fas by FasL. Here we show that HIV-1 Nef associates with and inhibits apoptosis signal-regulating kinase 1 (ASK1), a serine/threonine kinase that forms a common and key signalling intermediate in the Fas and tumour-necrosis factor-alpha (TNFalpha) death-signalling pathways. The interaction of Nef with ASK1 inhibits both Fas- and TNFalpha-mediated apoptosis, as well as the activation of the downstream c-Jun amino-terminal kinase. Our findings reveal a strategy by which HIV-1 Nef promotes the killing of bystander cells through the induction of FasL, while simultaneously protecting the HIV-1-infected host cell from these same pro-apoptotic signals through its interference with ASK1 function.