PD-1在T细胞非霍金淋巴瘤外周血淋巴细胞中的研究

发布时间：2018-06-15 15:10

本文选题：程序性死亡因子-1 + T细胞非霍奇金淋巴瘤　；参考：《北京协和医学院》2017年博士论文

【摘要】：背景T细胞非霍奇金淋巴瘤(T cell non-Hodgkin lymphoma,T-NHL)是一组异质性疾病,其恶性程度高,5年总生存率仅为20-40%。目前常规治疗方案都易复发,并且一旦复发缺乏有效的挽救治疗方案。因此积极推动T-NHL临床研究,探究其发病机制及药物靶点应当成为淋巴瘤研究的重点。肿瘤微环境和肿瘤相关免疫是近年的研究和治疗热点,外周血循环淋巴细胞是肿瘤患者机体免疫系统的重要组成部分,已有研究证实淋巴瘤患者外周血T细胞表面的共刺激分子中常存在异常表达,与肿瘤的免疫逃逸相关。其中程序性死亡因子-1(programmed death-1,PD-1)与肿瘤免疫关系最为密切,然而循环淋巴细胞表面PD-1在T-NHL中的研究相对较少,本文对PD-1对于T-NHL患者的机体免疫的影响及作用进行初步探讨。目的1.明确PD-1及相关共刺激分子在初治T-NHL外周血Treg细胞表面的表达及其治疗后变化。2.探究T-NHL患者与健康人外周血PD-1(+)淋巴细胞功能的差异性。3.初步揭示T-NHL患者外周血PD-1(+)淋巴细胞功能差异性的分子基础。方法本研究共纳入2015年9月至2017年4月在北京协和医院诊治的60例T-NHL患者,和在健康体检中心查体的65名健康志愿者。采用流式细胞技术分析外周血Treg细胞表面PD-1及相关共刺激分子的表达水平,分析表达与患者预后间的关系。并在患者治疗第2-6疗程后,再次测定患者外周血T淋巴细胞表面PD-1及相关共刺激分子的表达情况,探究表达与患者疗效的关系。利用磁珠分选10例初治T-NHL及10例健康受试者的外周血PD-1(+)淋巴细胞,进行体外培养,采用MTT试验对比患者及健康受试者外周血PD-1(+)淋巴细胞功能,此外利用数字化基因表达谱测序技术分析患者及健康受试者外周血PD-1(+)淋巴细胞基因表达谱差异,探究功能差异的分子基础。结果1.1)初治T-NHL患者外周血Treg细胞表面CD279及CD152平均荧光强度较健康对照组明显升高(P0.001);CD152及CD28表达比例较健康对照明显降低(P0.001);2)中/高危患者外周血Treg细胞表面CD279及CD152平均荧光强度较低危患者组明显升高(P0.05),CD28表达比例较低危患者组明显降低(P0.05);3)血管免疫母细胞性T细胞淋巴瘤患者外周血Treg细胞表面CD279及CD28表达平均荧光强度较结外NK/T细胞淋巴瘤(鼻型)患者明显降低(PP0.05);4)T-NHL患者治疗后外周血Treg细胞表面CD279及CD152平均荧光强度较治疗前明显降低(P0.05),CD28平均荧光强度较治疗前明显升高(P0.05)。5)T-NHL患者外周血Treg细胞表面PD-1及CD152表达与患者无事件生存时间相关(P0.05)。2.T-NHL患者外周血PD-1(+)淋巴细胞转化率低于健康受试者(P0.001)。3.与健康对照相比,T-NHL患者外周血PD-1(+)淋巴细胞具有明显不同的基因表达谱,与差异基因相关的生物学过程包括细胞增殖/凋亡、免疫反应、信号传导、物质代谢及细胞粘附等,主要涉及IFN、MYD88以及Toll样受体相关通路。结论1、初治T-NHL患者外周血Treg表面PD-1及相关共刺激分子表达较健康人具有明显差异,与疾病分层及预后相关;经过治疗后,T-NHL患者外周血Treg细胞表面PD-1及相关共刺激分子表达向健康人水平转变。2、T-NHL患者外周血PD-1(+)淋巴细胞功能较健康人明显减弱。3、T-NHL患者外周血PD-1(+)淋巴细胞较健康人基因表达谱具有明显差异,差异表达的基因可能与T-NHL的发生及肿瘤免疫相关。
[Abstract]:Background T cell non Hodgkin lymphoma (T cell non-Hodgkin lymphoma, T-NHL) is a heterogeneous group of heterogeneous diseases with a high degree of malignancy. The 5 year total survival rate is only 20-40%. and the conventional therapy is easy to relapse, and once a relapse lacks effective salvage therapy, T-NHL clinical research is actively promoted to explore its pathogenesis and drug. The target point should be the focus of lymphoma research. Tumor microenvironment and tumor related immunity are the focus of research and treatment in recent years. Peripheral blood circulating lymphocytes are an important part of the immune system of the tumor patients. The abnormal expression of the co stimulators in the T cell surface of the peripheral blood of the lymphoma patients has been proved to be abnormal. The immune escape of the tumor is related. The procedural death factor -1 (programmed death-1, PD-1) is most closely related to the tumor immunity. However, the study of PD-1 in the circulating lymphocyte surface is relatively less in T-NHL. The effect and effect of PD-1 on the immunity of T-NHL patients were preliminarily discussed in this paper. Objective 1. PD-1 and associated co stimulation were clearly defined. The expression of molecules on the surface of Treg cells in the primary T-NHL peripheral blood and the change of.2. to explore the differential.3. of PD-1 (+) lymphocyte function in peripheral blood of T-NHL patients and healthy people preliminarily reveal the molecular basis of the functional difference of PD-1 (+) lymphocyte in peripheral blood of T-NHL patients. Methods this study was included in the Beijing association from September 2015 to April 2017. 60 patients with T-NHL, and 65 healthy volunteers who were examined in a health check-up center, were used to analyze the expression level of PD-1 and related co stimulators on the surface of peripheral blood Treg cells by flow cytometry, and to analyze the relationship between the expression and the prognosis of the patients. After the treatment of the 2-6 course of treatment, the T lymph nodes were again measured in the peripheral blood of the patients. The expression of PD-1 and related costimulatory molecules on the cell surface was used to explore the relationship between the expression and the curative effect of the patients. Using magnetic beads, the peripheral blood PD-1 (+) lymphocytes of 10 patients with T-NHL and 10 healthy subjects were selected and cultured in vitro. The function of PD-1 (+) lymphocyte in peripheral blood of the patients and healthy subjects was compared with those of the healthy subjects, and the digitalization was also used. The gene expression spectrum sequencing technique was used to analyze the difference of PD-1 (+) lymphocyte gene expression profiles in patients and healthy subjects and to explore the molecular basis of functional differences. Results 1.1) the average fluorescence intensity of CD279 and CD152 on the surface of peripheral blood Treg cells in the early treatment of T-NHL patients was more than that of the healthy control group of Xian Shenggao (P0.001); the expression ratio of CD152 and CD28 was healthier than that of healthy controls. The average fluorescence intensity of CD279 and CD152 on the surface of peripheral blood Treg cells in the middle / high risk patients was significantly higher than that in the low risk group (P0.05), and the proportion of CD28 expression was significantly lower than that of the low risk group (P0.05), and the average fluorescence intensity of CD279 and CD28 expression of the Treg cell surface of peripheral blood on the peripheral blood of the patients with angioimmunocyte T cell lymphoma was significantly increased (P0.05) in the middle / high risk patients. (2) the average fluorescence intensity of CD279 and CD152 in the peripheral blood of the middle / high risk patients was significantly higher than that in the low risk group (P0.05) The patients with NK/T cell lymphoma (nasal type) were significantly lower (PP0.05); 4) the average fluorescence intensity of CD279 and CD152 on the Treg cell surface of the peripheral blood of T-NHL patients was significantly lower than that before treatment (P0.05), and the average fluorescence intensity of CD28 was significantly higher than that before treatment (P0.05).5). The transformation rate of PD-1 (+) lymphocyte in peripheral blood of patients with P0.05.2.T-NHL was lower than that of healthy subjects (P0.001).3. and healthy controls. The PD-1 (+) lymphocyte in peripheral blood of T-NHL patients had distinct gene expression profiles. The biological processes related to differential genes included cell proliferation / apoptosis, immune response, and signal transduction IFN, MYD88 and Toll like receptor related pathways are mainly involved in material metabolism and cell adhesion. Conclusion 1, the expression of PD-1 and related co stimulators on the Treg surface of the peripheral blood of T-NHL patients is significantly different from that of the healthy people, and is related to the stratification and prognosis of the disease. After treatment, the PD-1 and associated co stimulation of the peripheral Treg cells on the peripheral blood of the patients with T-NHL are stimulated. The expression of PD-1 (+) lymphocyte function in peripheral blood of T-NHL patients was significantly lower than that of healthy people. The expression of PD-1 (+) lymphocyte in peripheral blood of T-NHL patients was significantly different from that of healthy people. The differentially expressed genes may be related to the occurrence of T-NHL and tumor immunity in patients with T-NHL.
【学位授予单位】：北京协和医学院
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R733.1

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