TCR基因重排在T淋巴细胞浸润亲表皮疾病中的应用

目的:探讨具有亲表皮现象的疾病T细胞受体γ链基因重排的情况.方法:用免疫组化标记筛选17例T细胞淋巴瘤、30例可疑为T细胞淋巴瘤和10例副银屑病,采用聚合酶链式反应扩增方法检测T细胞受体γ链基因重排.结果:17例T细胞淋巴瘤中12例出现T细胞受体γ链基因重排,30例可疑为T细胞淋巴瘤和10例副银屑病均无T细胞受体γ链基因重排.结论:T细胞受体γ链基因重排检测是区分T淋巴细胞浸润的亲表皮疾病鉴别诊断的有效方法,且对T细胞淋巴瘤的确诊有重要参考价值与显著意义.     

蕈样肉芽肿患者皮损及外周血淋巴细胞CD28分子表达的研究

目的探讨CD28分子在蕈样肉芽肿发病机制中的作用。方法15例蕈样肉芽肿病人,分别采用免疫组织化学ABC方法检测皮损组织,流式细胞仪检测外周血淋巴细胞上的CD28分子表达水平。结果免疫组化结果显示蕈样肉芽肿组皮损中CD28表达明显增多,与正常人对照组相比差异均有显著性(t=4.53,P<0.01)。流式细胞仪检测显示MF外周血淋巴细胞上CD28 T淋巴细胞低于正常对照组(t=16.12,P<0.001)。MF外周血CD8 -CD28 T淋巴细胞低于正常对照组(t=4.925,P<0.001)。结论CD28分子在蕈样肉芽肿发病过程中起一定的作用。     

HHV-8与非Kaposi肉瘤组织相关性的初步研究

目的:检测人类疱疹病毒8型(HHV-8)在四种非Kaposi肉瘤皮损组织中的感染情况,以初步探讨HHV-8与皮肤鳞状细胞癌、蕈样肉芽肿、Castleman病和化脓性肉芽肿相关性研究.方法:使用酚-氯仿-异戊醇方法对新疆维吾尔自治区人民医院皮肤性病科、病理科从2004年到2009年收集的30例鳞状细胞癌、20例Castleman病(其中5例为多中性Castleman病)、13例蕈样肉芽肿、20例化脓性肉芽肿石蜡包埋组织进行HHV-8DNA抽提;然后使用巢式PCR方法对抽提的DNA进行ORF26基因片段的扩增,扩增产物在1.5%的琼脂糖凝胶上电泳,并在凝胶成像系统下观察电泳结果,从而确定ORF26基因阳性标本:然后计算HHV-8在各种标本中的阳性率.结果:30例皮肤鳞状细胞癌皮损组织有5份为阳性(16.7%).13例蕈样肉芽肿石蜡组织有1例为阳性(7.7%),20例化脓性肉芽肿和20例Castleman病石蜡组织中均未检出HHV-8.结论:①HHV-8可能是新疆皮肤鳞状细胞癌发病原因之一.②HHV-8与蕈样肉芽肿的发病可能不具有相关性,HHV-8可能是MF发生后偶发的一种附带感染.(③Castleman 病和化脓性肉芽肿的发病可能均与HHV-8无关.     

N-甲基亚硝基脲诱导小鼠胸腺淋巴瘤TCR基因重排分析

目的探讨N-甲基亚硝基脲（MNU）诱导的小鼠胸腺淋巴瘤的单克隆起源。方法采用巢式PCR方法,对8例MNU诱导的胸腺淋巴瘤组织进行T细胞受体β链（TCRβ）和γ链（TCRγ）克隆性基因重排分析,并对TCRγ基因重排的PCR产物直接测序。结果 8例胸腺淋巴瘤检测TCRβ和TCRγ均呈克隆性基因重排。DNA序列测定证实TCRγ基因PCR扩增产物为基因重排产物。结论巢式PCR TCR基因重排检测及DNA序列分析证实,MNU诱导的小鼠胸腺淋巴瘤是来源于T细胞的肿瘤。     

蕈样肉芽肿与扁平苔藓、慢性皮炎湿疹浸润细胞的免疫组化比较研究

目的探讨免疫表型对蕈样肉芽肿与扁平苔藓、慢性皮炎湿疹鉴别诊断的意义。方法本研究应用ABC免疫组化技术检测15例蕈样肉芽肿,17例慢性皮炎湿疹,17例扁平苔藓,6例正常皮肤的CD1a、CD8、ICAM-1和CD7的表达情况。结果1.蕈样肉芽肿表皮CD1a,CD8,ICAM-1的阳性细胞密度明显高于扁平苔藓、皮炎湿疹和正常皮肤。2.蕈样肉芽肿真皮中CD1a阳性细胞的线性密度高于扁平苔藓、慢性皮炎湿疹和正常皮肤,蕈样肉芽肿真皮内CD8阳性细胞百分比较慢性皮炎湿疹增多,蕈样肉芽肿真皮表达ICAM-1抗原的细胞百分比高于扁平苔癣和慢性皮炎湿疹。3.慢性皮炎湿疹、扁平苔藓真皮内CD7阳性细胞百分比高于蕈样肉芽肿和正常皮肤。结论CD1a、CD8、ICAM-1、CD7免疫表型的研究可能为蕈样肉芽肿和扁平苔藓、慢性皮炎湿疹等一些炎症性皮肤病的鉴别诊断提供一定的线索。     

蕈样肉芽肿和湿疹中Survivin 和Bcl-xL的表达分析

目的:探讨蕈样肉芽肿、湿疹、正常皮肤中Survivin、Bcl-xL蛋白的表达差异性及其临床意义。方法:运用免疫组织化学法检测30例蕈样肉芽肿、20例湿疹及15例正常人皮肤组织中Survivin、Bcl-xL蛋白的表达情况。结果:蕈样肉芽肿表皮中及真皮中Survivin、Bcl-xL的阳性表达率显著高于湿疹及正常皮肤,差异均有统计学意义(P0.05)。各时期蕈样肉芽肿表皮、真皮中Survivin、Bcl-xL的表达差异有统计学意义(P0.05)。结论:Survivin和Bcl-xL在蕈样肉芽肿中高表达其表达异常在蕈样肉芽肿的发生中起着重要作用,Survivin、Bcl-xL可能作为鉴别诊断蕈样肉芽肿、皮肤炎症性疾病有价值的参考指标。     

儿童急性淋巴细胞白血病特异性标记物的精准检测方法研究

目的：T细胞和免疫球蛋白重链基因重排是微小残留病灶水平的特异性标记物,而微小残留病灶的水平与儿童急性淋巴细胞白血病的复发强烈相关。应用传统的聚合酶链式反应方法来监测IgH／TCR基因重排不仅耗时、耗人力,而且敏感度较低。本研究旨在探索一种更为高效和敏感与实用的监测IgH／TCR基因重排的精准检测方法。方法：应用多重PCR技术检测26个患有急性淋巴细胞白血病的儿童的外周血样品中的标记物,这些儿童是在中国哈尔滨市最近两年内被诊断的患者。分别应用基因扫描和毛细血管电泳方法检测IgH（FRI,FRII,FRⅢ）／TCR（TCRB,TCRγ）基因重排和分析PCR产物的片段。结果：IgH／TCR基因重排和对IgH基因重排的阳性率分别为92．3％和75％,在26个病例中,4个复发病人的IgH的三个片段（FRI,FRII,FRⅢ）基因重排显示阳性。进一步分析显示复发与ign基因重排呈线性相关。结论：实验与临床应用表明,基因扫描这种方法对于IgH／TCR基因重排的检测是可靠的、实用的,因而可用于儿童急性淋巴细胞白血病的诊断和随访。     

T细胞抗原受体的结构与功能研究进展

T细胞抗原受体(T ceIl receptor,TCR)是T细胞表面关键的受体分子。TCR特异性地识别各种多肽抗原并通过胞内区ITAM磷酸化传递抗原刺激信号,进而引发T细胞的免疫效应。TCR的活性异常将会导致自身免疫病和免疫缺陷病的发生。对于TCR结构和功能的深入研究有助于我们更好地理解免疫反应的分子机理,从而为相关免疫疾病的预防和治疗提供重要的理论依据。该文对TCR的分类、基因重排机制、受体组装方式及其结构基础、TCR对抗原的识别以及活化机制等方面的研究成果进行了总结,综述了近几年来的最新研究进展。     

蕈样肉芽肿和湿疹中p53, Cox-2 的表达分析

目的:探讨蕈样肉芽肿、湿疹、正常皮肤中p53、Cox-2蛋白的表达差异性及其临床意义。方法:运用免疫组织化学法检测30例蕈样肉芽肿、20例湿疹及15例正常人皮肤组织中p53、Cox-2蛋白的表达情况。结果:蕈样肉芽肿表皮中及真皮中p53、Cox-2的阳性表达率显著高于湿疹及正常皮肤,差异均有统计学意义(P0.05)。各时期蕈样肉芽肿表皮、真皮中p53、Cox-2的表达比较差异均无统计学意义(P0.05)。结论:p53、Cox-2在蕈样肉芽肿和湿疹、正常皮肤的表达有显著差异,提示p53和Cox-2的表达异常在蕈样肉芽肿的发生中起着重要作用,p53、Cox-2可能作为鉴别诊断MF、皮肤炎症性疾病有价值的参考指标。     

树突状表皮T细胞在胸腺内发育的研究进展

树突状表皮T淋巴细胞(DETC),特异性分布在表皮组织内,在皮肤免疫监视,皮肤伤面愈合中发挥重要作用.小鼠DETC发育仅涉及胚胎14.5 ～ 18 d这一短暂时间窗,而后则不再产生这类细胞.本综述拟从DETC T细胞受体(TCR)基因重排特点及调控机制,DETC在胸腺中的阳性选择及调控机制等方面进行论述,以期对DETC...     

TCR‐β repertoire analysis of antigen‐specific single T cells using a high‐density microcavity array

The antigen specificity of cytotoxic T cells, provided by T‐cell receptors (TCRs), plays a central role in human autoimmune diseases, infection, and cancer. As the TCR repertoire is unique in individual cytotoxic T cells, a strategy to analyze its gene rearrangement at the single‐cell level is required. In this study, we applied a high‐density microcavity array enabling target cell screening of several thousands of single cells for identification of functional TCR‐β gene repertoires specific to melanoma (gp100) and cytomegalovirus (CMV) antigens. T cells expressing TCRs with the ability to recognize fluorescent‐labeled antigen peptide tetramers were isolated by using a micromanipulator under microscopy. Regularly arranged cells on the microcavity array eased detection and isolation of target single cells from a polyclonal T‐cell population. The isolated single cells were then directly utilized for RT‐PCR. By sequencing the amplified PCR products, antigen‐specific TCR‐β repertoires for gp100 and human cytomegalovirus antigens were successfully identified at the single‐cell level. This simple, accurate, and cost‐effective technique for single‐cell analysis has further potential as a valuable and widely applicable tool for studies on gene screening and expression analyses of various kinds of cells. Biotechnol. Bioeng. 2010;106: 311–318. © 2010 Wiley Periodicals, Inc.     

正常人外周血αβ T细胞TCR β链CDR3多态性和长度分析

采用免疫扫描谱型分析技术,分析正常人外周血T细胞TCR β链CDR3的多态性和长度分布.提取6例正常人外周血单个核细胞（peripheral blood mononuclear cell PBMC）的总RNA,逆转录成cDNA,以24个TCR BV基因家族为上游引物,共同的TCR BC基因为下游引物（荧光标记）,6例样品PCR产物的基因扫描分析表明,24个TCR BV家族的CDR3谱型分析在1.5%琼脂糖凝胶电泳图上显示1个模糊条带.序列测定结果显示,各TCR BV家族的CDR3谱型均超过8个条带.GeneScan分析证明,正常人外周血T细胞的24个TCR BV家族的CDR3谱型为标准的高斯分布,各家族的CDR3表达频率相近,呈现不同的CDR3多态性和不同的CDR3长度.多数TCR BV家族产物为框架内重排（相邻2个PCR产物相差3个碱基）,少数TCR BV家族表现为两个峰群的重排.该研究对正常人TCR β链CDR3谱系的详细分析将为T细胞应答、TCR重组等研究提供基础.利用免疫谱型分析技术能监测到TCR CDR3谱型分布特征和表达频率的变化.     

以T细胞受体为基础的免疫疗法研究进展

T细胞是机体抗肿瘤免疫的核心,以T细胞功能调控为基础的免疫检查点疗法已经在多种肿瘤的临床治疗中取得了重大突破,以基因工程化T细胞为基础的过继性免疫细胞疗法在血液瘤治疗中取得了重要进展,免疫治疗已经对肿瘤的临床治疗产生了深刻变革,成为肿瘤临床治疗策略的重要组成部分。T细胞受体（T cell receptor,TCR）赋予了T细胞识别肿瘤抗原的特异性,能够识别由主要组织相容性复合体（major histocompatibility complex,MHC）呈递的包括胞内抗原在内的广泛肿瘤抗原,具有高度的抗原敏感性,因而具有广泛的抗肿瘤应用前景。2022年第一款TCR药物的上市开启了TCR药物开发的新纪元,多项TCR药物临床研究表现出潜在的肿瘤治疗价值。本文综述了以TCR为基础的免疫治疗策略研究进展,包括T细胞受体工程化T细胞（T cell receptor-engineered T cell,TCR-T）和TCR蛋白药物,以及基于TCR信号的其他免疫细胞疗法,以期为以TCR为基础的免疫治疗策略开发提供参考。     

TCR transgenes and transgene cassettes for TCR gene therapy: status in 2008

The genetic introduction of T cell receptor genes into T cells has been developed over the past decade as a strategy to induce defined antigen-specific T cell immunity. With the potential value of TCR gene therapy well-established in murine models and the feasibility of infusion of TCR-modified autologous T cells shown in a first phase I trial, the next key step will be to transform TCR gene transfer from an experimental technique into a robust clinical strategy. In this review, we discuss the different properties of the TCR transgene and transgene cassette that can strongly affect both the efficacy and the safety of TCR gene transfer. This paper is a focussed research review based on a presentation given at the sixth annual meeting of the Association for Immunotherapy of Cancer (CIMT), held in Mainz, Germany,15–16 May 2008.     

Self-peptide/MHC and TCR antagonism: physiological role and therapeutic potential

TCR antagonists are peptides that bind MHC molecules and can specifically inhibit T cell activation induced by antigens. Studying TCR antagonism has taken an important place in immunology for both theoretical and practical reasons. Deciphering the mechanism(s) of action of TCR antagonists can yield important information about interactions of the TCR with ligands, T cell development, and TCR signaling. Moreover, microorganisms may employ TCR antagonism to elude the attention of the immune system. Finally, specificity of inhibition makes TCR antagonists an ideal tool to seek antigen-specific immunomodulation. Present state of knowledge on these topics is reviewed.     

Contribution of the T cell receptor BJ gene to recognition of the P91A tumor antigen in DBA/2 mice

 To understand specific immune responses against a tumor, it is important to characterize T cells that recognize the tumor antigen. The mouse P91A antigen is one of the well-defined tumor antigens that is expressed on the P911 cell line, and T cells responding to the antigen in DBA/2 mice were reported to be restricted to BV8S2/S3 families in their T cell receptor (TCR) BV gene usage. We have further characterized the P91A-responding T cells in DBA/2 mice, focusing on TCR BJ gene usage and using the polymerase chain reaction/enzyme-linked immunosorbent assay and DNA sequencing studies of their third complementarity-determining (CDR3) regions. As a result, T cells with cytotoxic activity to the P91A antigen, induced from murine spleen cells both in vivo and in vitro, showed predominant use of the BJ2S1 gene segment in both BV8S2 and BV8S3 T cells compared to unmanipulated murine spleen cells. Sequencing studies of the CDR3 regions in the BV8S3 T cells revealed clonal expansion of T cells with the BV8S3-BJ2S1 combination in two of three DBA/2 mice tested. In the remaining mouse, clonal expansion was not detected despite predominant use of the BJ2S1 segment by these T cells. These data suggest that P91A-recognizing T cells would predominantly use the BV8S2/S3-BJ2S1 combination. Analysis of T cells with these TCR BV-BJ gene combinations may contribute to the evaluation, monitoring and development of a T-cell-mediated immunotherapeutic strategy. Received: 3 July 1997 / Accepted: 17 November 1997     

Regulating the discriminatory response to antigen by T-cell receptor

The cell-mediated immune response constitutes a robust host defense mechanism to eliminate pathogens and oncogenic cells. T cells play a central role in such a defense mechanism and creating memories to prevent any potential infection. T cell recognizes foreign antigen by its surface receptors when presented through antigen-presenting cells (APCs) and calibrates its cellular response by a network of intracellular signaling events. Activation of T-cell receptor (TCR) leads to changes in gene expression and metabolic networks regulating cell development, proliferation, and migration. TCR does not possess any catalytic activity, and the signaling initiates with the colocalization of several enzymes and scaffold proteins. Deregulation of T cell signaling is often linked to autoimmune disorders like severe combined immunodeficiency (SCID), rheumatoid arthritis, and multiple sclerosis. The TCR remarkably distinguishes the minor difference between self and non-self antigen through a kinetic proofreading mechanism. The output of TCR signaling is determined by the half-life of the receptor antigen complex and the time taken to recruit and activate the downstream enzymes. A longer half-life of a non-self antigen receptor complex could initiate downstream signaling by activating associated enzymes. Whereas, the short-lived, self-peptide receptor complex disassembles before the downstream enzymes are activated. Activation of TCR rewires the cellular metabolic response to aerobic glycolysis from oxidative phosphorylation. How does the early event in the TCR signaling cross-talk with the cellular metabolism is an open question. In this review, we have discussed the recent developments in understanding the regulation of TCR signaling, and then we reviewed the emerging role of metabolism in regulating T cell function.     

TCR基因免疫治疗中优化转TCR基因配对的研究进展

TCR基因修饰T细胞的过继性免疫治疗是指将识别肿瘤抗原的特异性TCR基因转导至外周血T细胞,经大量扩增后回输给患者,从而发挥抗肿瘤效应的一种治疗技术。目前TCR基因治疗所面临的关键问题之一是如何改造修饰转TCR基因使得转TCR α链和β链在T细胞表面优先配对以提高转T细胞的功能,并避免off-target反应毒性的产生。最近,各种基因修饰策略被用于优化转TCR基因配对和减少错配。介绍了近年来针对TCR基因进行修饰改造的各种策略及TCR基因治疗的临床试验。     

T-cell receptor Vbeta gene usage in CSF lymphocytes in X-linked adrenoleukodystrophy

X-linked adrenoleukodystrophy (ALD) is a peroxisomal disorder with impaired very-long-chain fatty acid (VLCFA) metabolism that produces a neurological disease with significant variability of clinical phenotypes even within kindred. The two most common forms are the cerebral form (CALD) with an important inflammatory reaction at the active edge of demyelinating lesions, resembling some aspects of multiple sclerosis pathology, and adrenomyeloneuropathy (AMN), which involves the spinal cord and in which the inflammatory reaction is mild or absent. One hypothesis is that the phenotypic variability is related to T cell-mediated immune mechanisms playing a primary role in the demyelinating pathogenic process of CALD. The present study aims to test the hypothesis that CSF of patients with the CALD form contains highly restricted T cell populations. The variable regions of the T cell receptor beta chains (TCR Vbeta) were studied in CSF from 29 ALD patients with different phenotypes. RNA was extracted and cDNA synthesized from CSF lymphocytes; TCR Vbeta gene segments were amplified from the cDNA by polymerase chain reaction (PCR) using 20 family-specific primers. PCR products were analyzed by Southern blot. Some amplified Vbeta products were sequenced. The majority of ALD patients (21/29), whatever their phenotype, exhibited oligoclonal T cell expansion. However the overexpression of some TCR Vbeta families was heterogeneous among the different patients without any preponderance of specific Vbeta families or any clustering according to clinical phenotype. In particular a dominant TCR Vbeta utilization was not found in patients with CALD.