Vα24 NKT细胞的体外扩增和生物学功能研究

发布时间：2018-04-06 07:42

本文选题：Vα24　切入点：NKT细胞　出处：《中国协和医科大学》2005年博士论文

【摘要】：NKT细胞兼备固有性和适应性免疫细胞特性，被脂类或糖脂抗原激活后能迅速产生大量细胞因子，并上调表达细胞毒分子。在调节肿瘤免疫和移植耐受中可能有着重要作用。人NKT细胞T细胞受体库的α链为Vα24 JαQ，与之优势配对的β链为Vβ11，称为Vα24 NKT细胞。本研究拟建立体外诱导扩增Vα24 NKT细胞的方法，探讨特异抗原和细胞因子对扩增的影响；其次评价扩增的Vα24 NKT细胞的特性以及细胞因子对其特性的影响；确定Vα24 NKT细胞体外肿瘤杀伤活性以及对同种反应性T细胞免疫应答的抑制；初步论证Vα24 NKT细胞在人-小鼠异种移植中的作用。 34例外周血或脐带血单个核细胞(mononuclear cells，MNC)在α-半乳糖酰基鞘氨醇(α-galactosylceramide，α-GalCer)以及IL-2、IL-15诱导下培养12d，Vα24NKT细胞扩增343.5(95～2207)倍。其中以脐带血前体细胞来源树突状细胞(dendritic cells，DC)负载α-GalCer诱导脐带血Vα24 NKT细胞扩增221.5(95～501)倍(A1组，n=6)，以成人单核细胞来源DC诱导Vα24 NKT细胞扩增339(168～1263)倍(A2组，n=6)，成人外周血直接添加α-GalCer诱导其中NKT细胞扩增达413.5(101～2207)倍(B组，n=22)。B组效果明显优于A1组(P=0.020)，B组与A2组差异无显著性(P=0.576)，A2组似乎优于A1组，但无统计学意义(P=0.132)。所有34例中16例第二次经α-GalCer刺激，其中7例d20扩增达722～28301倍，其余的在培养过程中凋亡，，7例中的2例第三次经α-GalCer刺激继续生长，其余凋亡。 4例脐带血CD34细胞以TPO、IL-3、SCF、Flt3L、G-CSF、IL-2、IL-15、IL-7诱导扩增5周，细胞增长1748.5(826～3558)倍；Vα24NKT细胞比例为6.67％(5.66％～18.6％)。将3例外周血MNC分别分5组培养12d。α-GalCer组(a组)细胞很快凋亡，α-GalCer+细胞因子(d和e组)扩增效果明显优于单用细胞因子(b和c组)(P=0.002)；α-GalCer+IL-2+IL-15(e组)在所有组中效果最佳(P=0.037)。将7例外周血MNC标本每例分为若干组，在不同细胞因子组合条件下经
[Abstract]:NKT cells possess both innate and adaptive immunocytic properties. Activated by lipids or glycolipid antigens, NKT cells can rapidly produce a large number of cytokines and up-regulate the expression of cytotoxic molecules.It may play an important role in the regulation of tumor immunity and transplantation tolerance.The 伪 chain of T cell receptor library of human NKT cell is V 伪 24 J 伪 Q, and the 尾 chain of T cell receptor library is V 尾 11, which is called V 伪 24 NKT cell.The aim of this study was to establish a method to induce the amplification of V 伪 24 NKT cells in vitro, to investigate the effects of specific antigen and cytokines on the amplification, and then to evaluate the characteristics of the amplified V 伪 24 NKT cells and the effects of cytokines on the amplification of V 伪 24 NKT cells.To determine the tumor killing activity of V 伪 24 NKT cells and the inhibition of immune response to alloreactive T cells in vitro, and to demonstrate the role of V 伪 24 NKT cells in human mouse xenotransplantation.34 mononuclear cells (MNCs) from peripheral blood or umbilical cord blood were amplified 343.5 times by 伪 -galactosylceramide (伪 -Gal-Cerl) and IL-2- (IL-15) -induced proliferation of mononuclear cells (MNCs) from 34. 5% to 95% (2207) times.Among them, dendritic cells derived from dendritic cells derived from umbilical cord blood progenitor cells were loaded with 伪 -GalCer to induce the expansion of V 伪 24 NKT cells in umbilical cord blood by 221.5 rm95 / 501). In A1 group, V 伪 24 NKT cells were induced to expand by dendritic cells derived from adult monocytes, V 伪 24 NKT cells from adult monocytes were induced to expand by 3391681263), and adult peripheral blood was directly added to A2 group.伪 -GalCer induces the expansion of NKT cells up to 413.5n 101U 2207) the effect of group B is significantly better than that of group A 1 (P 0. 020) and group A 2. There is no significant difference between group A 2 and group A 2. The effect of group A 2 seems to be better than that of group A1.However, there was no statistical significance (P < 0. 132).Among the 34 cases, 16 cases were stimulated by 伪 -GalCer for the second time, and 7 cases were amplified 722 ~ 28301 times on the 20th day, 2 of the other 7 cases continued to grow by 伪 -GalCer stimulation for the third time, and the remaining apoptosis continued.Four umbilical cord blood CD34 cells were induced and amplified by TPO-IL-3SCFT flt3LG-CSFG IL-2 IL-15 IL-7 for 5 weeks, and the cell growth rate was 1748.5% 8263558) and the ratio of V 伪 24NKT cells was 6.67% 5.66% and 18.6% respectively.Seven cases of peripheral blood MNC were divided into several groups under different cytokine combinations.
【学位授予单位】：中国协和医科大学
【学位级别】：博士
【学位授予年份】：2005
【分类号】：R329.2

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