重组IL-12腺病毒载体对肺炎小鼠NK细胞功能的影响

目的:探讨重组IL-12腺病毒载体(AdCMV IL-12)对肺炎小鼠NK细胞功能的影响.方法:90只BALB/c小鼠随机分为4组,A组:空白对照组15只,不给予病毒,滴鼻生理盐水.B组:对照组25只,每只鼻腔滴入10~5TCID_(50)/0.1 mL的Ad3悬液100 μL.C组:AdCMV IL-12组25只,每只鼻腔滴入10~5TCID_(50)的Ad3/0.1 mL悬液100 μL,3 d后滴鼻给药吸入AdCMV IL-12(5×10~8 PFU/只)100 μL.D组:AdCMVLacZ组25只,每只鼻腔滴入10~5TCID_(50)/0.1 mL的Ad3悬液100 μL,3 d后滴鼻给药吸入AdCMVLacZ(5×108 PFU/只)100 μL.开始滴入病毒定为0 d.于第5天,摘眼球取血,颈椎脱臼处死,无菌条件下取肺,用于转染及病毒毒力检测.无菌条件下取脾脏做NK细胞活性检测.测定血清IL-12及FIN-γ含量.结果:含有目的基因的腺病毒载体能有效转染支气管及肺组织并在局部表达目的基因.血清IL-12浓度为C组>A组>B组、D组,NK、FIN-γ浓度为C组>B组、D组>A组.病毒滴度:A组细胞生长良好.B、D组于10~(-4)出现细胞病变,C组于原液中出现病变.结论:重组IL-12腺病毒载体可通过滴鼻给药成功转染至肺组织,升高血液IL-12浓度,通过增加NK细胞活性而增加了IFN-γ的释放,从而导致病毒滴度下降,限制CVB病毒复制的作用.     

NK细胞新概念——趋化因子活化的杀伤细胞(CHAK)

趋化因子是新近发现的一类蛋白质 ,主要参与趋化、激活白细胞亚群 ,调节机体非特异免疫 ,特别是与爱滋病和肿瘤的关系引起了国内外极大关注。国内已发表了不少趋化因子方面的综述 ,但趋化因子与NK细胞的关系却鲜为人知 ,本文重点介绍CHAK的国内外研究进展     

094 NK细胞新概念--趋化因子活化的杀伤细胞(CHAK)

趋化因子是新近发现的一类蛋白质,主要参与趋化、激活白细胞亚群,调节机体非特异免疫,特别是与爱滋病和肿瘤的关系引起了国内外极大关注.国内已发表了不少趋化因子方面的综述,但趋化因子与NK细胞的关系却鲜为人知,本文重点介绍CHAK的国内外研究进展.     

NK细胞新概念——趋化因子活化的杀伤细胞（CHAK）

趋化因子是新近发现的一类蛋白质,主要参与趋化、激活白细胞亚群、调节机体非特异免疫,特别是与爱滋病和肿瘤的关系引起了国内外极大关注。国内已发表了不少趋化因子方面的综述,但趋化因子与NK细胞的关系却鲜为人知,本文重点介绍CHAK的国内外研究进展。     

神经肽P物质对小鼠脾NK细胞迁移及趋化因子受体表达的影响

探讨神经肽P物质(substance P,SP)对小鼠NK细胞迁移能力和趋化因子受体表达的影响,揭示SP对NK细胞迁移功能的调节作用。采用Transwell法检测SP对C57BL/6小鼠脾NK细胞迁移活性的影响;实时荧光定量PCR检测趋化因子受体(CCR5、CXCR4)的mRNA表达水平;流式细胞术检测趋化因子受体(CCR5、CXCR4)的膜表达水平。结果显示,选用10~(-14)～10~(-6)mol/L SP处理小鼠脾NK细胞,在较低浓度范围内,NK细胞迁移作用随着SP浓度的升高而增强;在较高浓度范围内,NK细胞迁移作用随着SP浓度的升高而回落至基础水平。在所选浓度范围内,SP均可增加CCR5 mRNA和CXCR4 mRNA表达;受较低浓度SP刺激后,CCR5和CXCR4表达阳性细胞数随SP浓度升高而逐渐增加,而受较高浓度SP刺激后,CCR5和CXCR4表达阳性细胞数随SP浓度升高而逐渐回降,该趋势与SP促进NK细胞趋化活性的作用趋势基本一致。这些结果提示SP对NK细胞具有直接趋化作用,且SP对某些趋化因子受体的表达有一定的调节作用,这些趋化因子受体的表达可能参与了SP对NK细胞迁移活动的调控。     

小鼠肝脏NK细胞特有亚群:DX5-NK细胞

近来在小鼠中发现一群特殊的DX5-NK(DX5-NK1.1+CD3-)细胞,其具有不成熟NK细胞的表型(Ly49-CD11blowCD94hiNKG2+),且杀伤及分泌细胞因子能力明显低于经典的DX5+NK1.1+CD3-细胞.在不同脏器或不同年龄阶段,DX5-NK细胞的表型和功能均存在差别.成年小鼠体内的DX5-NK细胞主要分布于肝脏,占肝脏总NK细胞(CD3-NK1.1+)的50%左右.研究DX5-NK细胞将为了解NK细胞的组织特异性分布和NK细胞发育分化提供新线索.     

NK／T细胞淋巴瘤的细胞遗传学研究

淋巴瘤属国内前十大常见恶性肿瘤之一。研究发现在NK／T胞淋巴瘤存在1，6，7，8，9，11，13，14，15，16，17，19，20，22，X等多个染色体畸变，对其进一步研究有助于发现NK／T细胞淋巴瘤是否存在特征性的改变。     

体外抗原诱导小鼠T细胞对NK细胞功能的抑制及其可能分子机制研究

目的:为了探讨抗原诱导T细胞抑制对NK细胞功能的抑制及其可能机制.方法:在体外,分别利用抗原、抗原 抗CD80抗体、刀豆蛋白A、抗CD3抗体先行诱导小鼠T细胞,并于诱导后24、48、72、96小时分别引入NK细胞,通过51Cr释放试验动态观察NK细胞的功能状况,激光共聚焦显微镜观察T细胞抑制NK细胞功能的作用模式,同时用RT-PCR方法检测经诱导的T细胞Bc1与Bc2表达情况.结果:①各组在24、48、72小时相点,上清液对NK功能均无显著影响,而到96小时,抗原激活组与抗原 抗CD80抗体诱导组两组上清液对NK细胞的杀伤能力表现出促进作用,与其它各时相点相比有统计学意义(P＜0.05),且后者低于前者,二者相比也具统计学意义(P＜0.05).②刀豆蛋白A与抗CD3抗体刺激组T细胞从T细胞接受刺激24小时起至96小时,均表现了抑制NK细胞功能.抗原 抗CD80抗体耐受诱导组与抗原激活组T细胞则是在诱导后48小时方表现出对NK细胞的抑制,与其它两组相比差异具有统计学意义(P＜0.05).③只有同种抗原刺激48小时的细胞培养物可检出Bc1、Bc2.其他时相、其他细胞刺激剂作用下均未能检出Bc1,Bc2的表达.结论:经诱导的T细胞能以直接接触的方式抑制NK细胞功能,但不同诱导剂活化的T细胞引起NK细胞抑制的机制可能不同.小鼠HLA-G的类似物-Bc1与Bc2的表达与抗原信号有关,抗原诱导的小鼠T细胞可能通过表达Bc1与Bc2来抑制NK细胞的功能,从而参与移植耐受的形成,这一结论提示这类非经典MHC Ⅰ类分子有可能成为移植耐受的检测标志.     

绿素酊对小鼠T淋巴细胞及NK细胞活性的影响

为研究绿素酊对正常和免疫抑制小鼠T淋巴细胞及NK细胞活性的影响。采用MTT还原法检测PHA诱导的T淋巴细胞增殖功能和NK细胞活性。结果显示 ,绿素酊对正常小鼠T淋巴细胞和NK细胞活性无明显影响 ,但能显著增强免疫抑制小鼠T淋巴细胞增殖及NK细胞活性。绿素酊能增强免疫抑制小鼠T淋巴细胞功能及NK细胞活性     

人外周血NK T细胞体外扩增的初步研究

自然杀伤Ｔ细胞 (ＮａｔｕｒｅＫｉｌｌｅｒＴｃｅｌｌｓ,ＮＫＴｃｅｌｌｓ)是近来被人们所认识的不同于普通Ｔ细胞及ＮＫ细胞的一类新的免疫调节细胞 ,在肿瘤治疗 ,自身免疫病 ,以及ＧＶＨＤ方面有着广泛的应用前景[1 3] 。该类细胞有着特殊的表型 ,它既可表达ＮＫ相关的表面标志物 ,又具有明显选择性倾向的αβ型ＴＣＲ受体。鼠为Ｖα1 4Ｖβ8,人则为Ｖα2 4Ｖβ1 1 ,其配体为递呈脂类抗原的ＣＤ1ｄ分子[4 ] 。由于ＮＫＴ主要存在于胸腺、骨髓和肝脏中 ,外周血含量极少[5] ,单纯依靠原位分离的方法很难获得足够的人ＮＫＴ细胞。本研究利…     

The Role of NK Cell in T Cell Recruitment in Murine Liver Infected with Adenovirus

Infections of the liver with virus usually induced host re sponses that damaged hepatocytes and concomitant increaseof liver enzymes such as ALT in serum. In some cases liv er injury caused liver failure. Therefore, it was importantto elucidate the molecular mechanisms of liver injury anddesigned approaches to prevent and treat liver injurycaused by viral infection. It was known that liver injurycaused by virus was mainly due to action of host cell medi ated immune system[1 3]. For …     

NK cells can generate from precursors in the adult human liver

Hepatic NK cells constitute ≈ 40% of hepatic lymphocytes and are phenotypically and functionally distinct from blood NK cells. Whether hepatic NK cells derive from precursors in the BM or develop locally from hepatic progenitors is still unknown. Here, we identify all five known sequential stages of NK-cell development in the adult human liver and demonstrate that CD34(+) hepatic progenitors can generate functional NK cells. While early NK-cell precursors (NKPs) were similar in liver and blood, hepatic stage 3 NKPs displayed immunophenotypical differences, suggesting the onset of a liver-specific NK-cell development. Hepatic stage 3 NKPs were RORC(neg) and did not produce IL-17 or IL-22, excluding them from the lymphoid tissue-inducer (LTi) subset. In vitro culture of hepatic NKPs gave rise to functional NK cells exhibiting strong cytotoxicity against K562 targets. To determine whether hepatic NKPs are stably residing in the liver, we analyzed donor and recipient-derived cells in transplanted livers. Shortly after liver transplantation all donor NKPs in liver grafts were replaced by recipient-derived ones, indicating that hepatic NKPs are recruited from the bloodstream. Together, our results show that NKPs are continuously recruited from peripheral blood into the liver and can potentially differentiate into liver-specific NK cells.     

小鼠NKT细胞在5型腺病毒感染所致肝损伤早期的免疫调节作用

目的 使用5型腺病毒(AdS)感染小鼠模型来研究肝脏NKT细胞(natural killer Tcell)在肝损伤早期的免疫调节机制.方法 C57BL/6小鼠尾静脉注射1.5×109PFU和3×109PFUAdS病毒以构建两个剂量组病毒感染的小鼠肝损伤模型,通过观察病毒感染后5 d内小鼠肝组织病理学及小鼠血清丙氨酸转氨酶/天门冬氨酸转氨酶(ALT/AST)水平改变来判断肝损伤程度,使用流式细胞术(FACS)分析感染5 d内肝单个核细胞亚群比例、NKT细胞表面FasL表达水平以及NKT细胞合成IL-4和IFN-γ水平的变化,应用RT-PCR检测小鼠肝内趋化因子及趋化因子受体表达水平.结果 高滴度(3×109 PFU)的AdS病毒感染小鼠1 d后,小鼠肝脏内NKT细胞明显增加,其表面FasL表达上调,肝脏NKT细胞合成IL-4和IFN-γ的水平明显增加,肝组织内淋巴细胞浸润明显;低滴度AdS病毒(1.5×109PFU)感染小鼠后,肝脏NKT细胞比例变化不明显,CD8+T细胞在肝脏的浸润明显弱于高滴度AdS病毒感染;RT-PCR检测结果 显示:3×109PFU AdS病毒感染2 d后,小鼠肝内活化后可调节的及正常的T细胞分泌的趋化因子(RANTES)、人干扰素诱导蛋白10(IP-10)以及巨噬细胞炎症蛋白(MIP)-1β表达增加,3d后相关趋化因子受体CCR5、CCR1、CXCR3表达上调.结论 NKT细胞在淋巴细胞向肝脏趋化的过程中起重要的作用,这种作用与病毒感染诱导NKT细胞合成IL-4和IFN-γ及上调其表面的FasL,从而促进肝细胞内IP-10、Mig等趋化因子的产生有关.     

Characterization of the role of TCRγδ in NK cell accumulation during viral liver inflammation

Polyinosinic-polyctidylic acid (Poly I:C) is a viral RNA mimic that can induce immune responses similar to that seen during viral infection. Although poly I:C administration into mice is associated increased NK cell infiltrates in the liver, the mechanisms underlying increased hepatic NK cell accumulation in response to poly I:C administration are incompletely defined. In the current study, we have identified a novel and important role for γδT cells in driving the accumulation and activation of NK cells in the liver during poly I:C-mediated viral liver infection. Specifically, NK cell accumulation but not activation in γδT cell deficient mice following poly I:C administration was significantly attenuated in comparison to that seen in poly I:C-treated wildtype mice. The ability of γδT cells to promote NK cell accumulation and activation in the liver may be virus-specific since NK cell accumulation in the liver was not altered by TCRγδ deficiency following adenovirus administration.     

非霍奇金淋巴瘤患者T细胞亚群、NK细胞检测的临床意义

目的：研究非霍奇金淋巴瘤（NHL）患者外周血T淋巴细胞亚群、NK细胞检测结果的变化与该病的关系及与慢性淋巴腺炎患者细胞免疫功能的不同变化。方法：采用流式细胞仪（FCM）检测非霍奇金淋巴瘤（NHL）患者、慢性淋巴腺炎及正常人外周血T淋巴细胞亚群比例、NK细胞的变化。结果：非雹奇金淋巴瘤患者与正常人比较总的T淋巴细胞、辅助性T淋巴细胞及CD4^＋／CD8^＋比值明显下降（P〈0．05），细胞毒性T淋巴细胞明显升高（P〈0．05），NK细胞则无明显变化（P〉0．05）。非霍奇金淋巴瘤患者与慢性淋巴腺炎患者比较，细胞毒性T淋巴细胞、NK细胞明显升高（P〈0.05），而总的T淋巴细胞、辅助性T淋巴细胞无明显改变（P〉0．05），CD4^＋／CD8^＋比值略有下降但无明显统计学意义。结论：非霍奇金淋巴瘤患者细胞免疫功能明显受到抑制，T细胞亚群及NK细胞的检测对NHL的诊断、治疗、预后判断有一定的临床价值。     

导入NKG5—SV基因增强细胞杀伤活性的研究

目的研究免疫效应蛋白NKG5的拼接异构体基因NKG5-SV导入外周血自然杀伤细胞(NK细胞)对其杀伤活性的影响.方法Percoll细胞分离液分离外周血NK细胞,流式细胞仪检测分离后CD56阳性细胞浓度.以逆转录病毒为载体将NKG5-SV基因和对照基因LacZ导入NK细胞,Northernblot检测NKG5-SV的表达,K562细胞杀伤实验检测病人NK细胞转染NKG5基因前后和正常人NK细胞的NK活性.结果Percoll分离后的淋巴细胞CD56阳性细胞为77.44%,分离前为15.88%.病人NK细胞经NKG5-SV基因转染后NKG5-SVmRNA表达增高.病人NK细胞、NK-LacZ细胞、NK-NKG5-SV细胞和正常人NK细胞对K562的杀伤活性分别为5.96%±0.38%、6.03%±0.42%(P＞0.05)、27.67%±0.18%(P＜0.01)、30.33%±0.83%(P＜0.01).结论免疫效应蛋白基因导入NK细胞,可增强NK细胞的杀伤活性.     

Diversity of tissue-resident NK cells

Although natural killer (NK) cells were initially named for their spontaneous tumor-killing capacity, their concept has been greatly expanded with more than 40 years of extensive investigation. Currently, NK cells are known as a heterogeneous population of innate lymphoid cell (ILC) family, consisting of different subsets with unique phenotypic and functional features. Recent studies have shown that tissue-resident NK (trNK) cells, which are distinct from conventional NK (cNK) cells, preferentially distribute in non-lymphoid tissues, such as the liver, uterus, salivary gland, and adipose. In this review, we provide a comprehensive overview of the current knowledge about the phenotype, function and development of trNK cells across different tissues and describe the similarities and differences between diverse trNK cells and cNK cells, with a particular focus on the tissue-specific characteristics of different trNK cells.     

血小板T细胞活化抗原1分子在NK细胞上的作用研究

目的 研究血小板T细胞活化抗原1（PTA1）分子在NK细胞杀伤过程中的作用。方法 应用重民地向细胞毒实验,探讨了PTA1分子对混合淋巴细胞反应中活化的NK细胞杀伤作用的影响。结果 在重导向细胞毒实验中,PTA1 McAb能明显上调NK细胞及CTL的杀伤活性。结论 PTA1分子在NK细胞上具有刺激性受体的功能。     

Transient Notch signaling induces NK cell potential in Pax5-deficient pro-B cells

Unlike early B/T cell development, NK cell lineage commitment is not well understood, with a major limitation being the lack of a robust culture system to assay NK cell progenitors. Here we have exploited the multi-lineage potential of Pax5(-/-) pro-B cells to establish an effective system to direct differentiation of progenitors into the NK cell lineage. Cultivation of Pax5(-/-) pro-B cells on OP9 cells expressing the Notch ligand Delta-Like1 (OP9-DL1) in the presence of IL-7 efficiently induced T and NK cell potential. For NK cells, Notch was only transiently required, as prolonged signaling decreased NK and increased T cell development. Pure NK cell populations could be obtained by the culture of these Notch signal-experienced cells onto OP9 stroma and IL-15. A similar transient exposure to Notch was also compatible with the differentiation of NK cells from hematopoietic progenitors, while sustained Notch signaling impaired NK cell generation. Pax5(-/-) pro-B cell-derived NK cells were cytotoxic, secreted cytokines and expressed all the expected NK cell-specific surface markers examined except the Ly49 family, a phenotype similar to fetal NK cells. These data indicate that Notch signaling induces T/NK cell differentiation in Pax5(-/-) pro-B cells that is strikingly similar to early thymopoiesis.