慢性乙型肝炎患者外周血CD8+/CD28+淋巴细胞及其亚型的研究

目的 探讨慢性乙型肝炎患者外周血CD8+/CD28+淋巴细胞及其亚型与其临床状态和HBV复制的关系.方法 采用流式细胞技术多色荧光分析法,检测研究对象的外周血CD8+淋巴细胞、CD45RO、CD45RA以及CD28的表达及HBV病毒载量.结果 ①慢性HBV携带组CD8+淋巴细胞CD28的表达率(10.64±5.09%)明显低于慢性乙肝组和正常对照组;而CD45RA和CD45RO的表达差异无统计学意义;②慢性乙肝组CD8+/CD45RO+/CD28+的表达(10.99±7.33%)明显高于正常对照组,而慢性乙肝组和慢性HBV携带组CD8+/CD45RA+/CD28+表达率均明显低于正常对照组;③HBeAg阴性慢性乙肝CD8+/CD45RO+/CD28+表达率明显高于阳性组,而CD8+/CD45RA+/CD28+表达率两组无差别.结论 慢性HBV携带者处于病毒携带状态可能与CD8细胞上协同分子CD28的表达低下有关;CD8+/CD45RO+/CD28+亚型淋巴细胞数的升高与慢性乙肝的病情进展及血清HBeAg转换有关.     

GD患者外周血CD4~+CD28-T细胞亚群的表型特征及临床意义

检测Graves病(GD)患者外周血CD4~+CD28-T细胞水平及其表面CD45RO/CD45RA及ICOS的表达,探讨CD4~+ CD28-T细胞亚群在GD免疫致病机制中的作用。采用三色荧光抗体染色及流式细胞术检测了42例初发GD患者和30例健康者外周血中CD4~+CD28-T细胞的百分率及其表面CD45RO/CD45RA和ICOS表达水平,同时检测其甲状腺功能并进行相关性分析。结果GD患者外周血中CD4~+CD28-T细胞百分率明显高于健康对照组,并高表达ICOS分子,与FT3水平显著正相关;与健康对照组相比,GD患者CD4~+CD28~-CD45RO~+T细胞百分率也显著增高,而CD4~+CD28~-CD45RA~+T细胞呈下降趋势,FT3、FT4水平与CD4~+CD28-T细胞表面CD45RO的表达率呈正相关,而FT3水平与CD45RA表达呈负相关。结论GD患者外周血CD4~+CD28~-T细胞异常增高,表面高表达ICOS分子,具有记忆性细胞的表型特征,与甲状腺功能异常有一定的相关性,CD4~+CD28-T细胞可能是参与GD免疫病理反应的自身反应性T细胞。     

桥本氏甲状腺炎患者外周血中CD4+ CD45RO+ 记忆性T 细胞的表达及意义

目的:检测初诊桥本氏甲状腺炎(Hashimoto’s thyroiditis,HT)患者外周血CD4+ CD45RO+ 记忆性T 细胞比例,探讨其在HT 发病中的意义。方法:收集初诊的HT 患者作为病例组(HT,n =53),另外选取年龄、性别相匹配的正常人作为对照组(HC,n =43),并根据甲状腺功能状态将HT 组分为甲状腺功能正常组(HT-A,n =15)、亚临床甲减组(HT-B,n =14)和临床甲减组(HT-C,n =24)。流式细胞仪检测外周静脉血中CD4+ CD45RO+记忆性T 细胞比例,酶联免疫吸附法检测血清中IFN 、IL-17 水平,化学发光法检测甲状腺功能及甲状腺特异性抗体滴度。结果:HT 组外周血中CD4+ CD45RO+ 记忆性T 细胞比例、IFN 、IL-17、TPOAb 与TgAb 水平均显著高于HC 组,P<0.01。双变量相关分析显示HT 患者外周血CD4+ CD45RO+记忆性T 细胞的比例与IFN 、TPOAb、TgAb 均呈正相关(P<0.01,P =0.015,P<0.01)。结论:CD4+ CD45RO+ 记忆性T 细胞在HT 患者外周血中呈高表达,CD4+ CD45RO+记忆性T 细胞可能参与了HT 的发病过程。     

CHOP方案治疗前后外周T细胞淋巴瘤患者外周血CD45RA~+ T和CD45RO~+ T的变化特点

目的:探讨CHOP方案对外周T细胞淋巴瘤(PTCL)患者外周血中初始和记忆T细胞水平的变化及其临床意义。方法:采用流式细胞术检测20例PTCL患者CHOP方案化疗前后外周血中CD4+CD45RA+、CD4+CD45RO+、CD8+CD45RA+和CD8+CD45RO+T细胞的比例,分析疗效与T细胞亚群的关系。结果:PTCL患者化疗前外周血中CD4+T细胞、CD4+CD45RO+细胞的比例明显降低,CD4+CD45RA+、CD8+、CD8+CD45RO+和CD8+CD45RA+T细胞的比例均明显升高(P<0.05),而化疗后PTCL患者CD4+、CD4+CD45RO+细胞的比例较治疗前升高,CD4+CD45RA+、CD8+、CD8+CD45RO+和CD8+CD45RA+T细胞比例则较治疗前稍下降(P<0.05)。治疗前后化疗有效组的CD4+CD45RA+均明显高于化疗无效组(P<0.05)。结论:CHOP治疗对PTCL患者胸腺输出功能有一定的影响,伴有较高胸腺输出功能者对化疗效果更好。     

重症肌无力CD45RA、CD45RO胸腺细胞亚群及相关细胞因子表达分析

目的:探讨胸腺细胞异常分化与重症肌无力发生的关系.方法:采用基因芯片对多种白细胞介素、干扰素及其受体mRNA表达进行分析, 采用流式细胞术测定胸腺细胞CD45RA、 CD45RO的表达率, 采用免疫组化对胸腺组织切片CD45RA、 CD45RO表达及分布进行检测.结果:MG患者IL-1R、 IL-4R、 IFNγR1、 IFNγR2、 IL-6、 IL-8表达水平显著低于对照组; IL-10RB表达水平显著高于对照组; IL-1、 IL-2、 IL-4、 IL-10、 IL-7、 IFN-α、 IFN-β、 IFN-γ表达水平在MG和对照组均很低, 无显著差异; MG患者CD56 胸腺细胞百分率(0.56±0.33)显著低于对照组(1.78±0.69), MG患者CD45RO 、 CD1a 细胞显著高于对照组.免疫组化和RT-PCR也有相同结果.结论:MG患者胸腺细胞发育过程中CD45RO 细胞向CD45RA T细胞转变存在异常.     

CD45RA／CD45RO临床研究进展

目的：探讨CD45RA／CD45RO的研究现状从而有效的指导临床工作。方法：查阅近年来国内外CD45RA／CIM5RO的相关文献，对其最新研究进行总结。结果：作为跨膜酪氨酸磷酸蛋白酶，CIM5RA／CD45RO在哺乳动物淋巴细胞信号传导、增殖和活化中有重要作用，是区分初始和记忆T细胞的重要标志。结论：CD45RA／CD45RO的检测在人类自身免疫疾病和病毒感染性疾病的诊断方面有重要意义。     

乙型肝炎病毒感染者外周血CD4+T细胞CD25、CD127不同亚群表达的检测及临床意义

目的 研究乙型肝炎病毒(HBV)感染者外周血CD4+T细胞表面CD25、CD127不同亚群的表达情况及临床意义。方法 用荧光抗体CD127-FITC、CD4-PECY5、CD25-PE标记T细胞。用流式细胞仪分别测定53例慢性乙型肝炎患者和53例HBV携带者CD4+T细胞表面CD25、CD127不同亚群的表达情况。对20例HBV-DNA阳性乙型肝炎病毒感染者干扰素治疗进行随访。结果与健康对照组[7.26％(6.15％,8.50％)]比较,慢性乙型肝炎患者[11.23％(9.10％,14.86％)]、HBV携带者[13.34％( 10.73％,18.90％)]CD25-CD 127-均显著升高,差异均有统计学意义(Q=4.559,P＜0.05;Q=6.230,尸＜0.05)。慢性乙型肝炎患者CD25hiCD127low/-[8.78％ (7.62％,10.44％)]显著高于健康对照[6.76％(5.73％,8.23％)]和HBV携带者[6.99％(5.77％,9.34％)],差异均有统计学意义(Q=3.497,P＜0.05;Q=3.103,P＜0.05)。HBV-DNA阳性组CD25-CD127-显著低于阴性组,两者差异有统计学意义[(12.92±5.20)％比(15.78±6.91)％,t=2.290,P=0.024],而CD25+/-CD127+显著高于阴性组,两者差异有统计学意义[(79.27±5.20)％比(76.02±7.04)％,t=2.194,P=0.030]。与治疗前比较,干扰素治疗12周CD25hiCD127low/-显著升高[(9.29±2.51)％比(11.08±2.38)％,t=2.820,P=0.011],而CD4+CD25-CD127-显著降低,两者差异有统计学意义[(13.86±5.72)％比( 10.86±3.60)％,t=2.469,P=0.024]。结论 HBV感染者外周血CD4+T细胞中CD25-CD127-亚群的表达与病毒的感染和清除有关;CD25hiCD 127low/-亚群的表达升高与发病有关。外源性干扰素可升高CD25hiCD127low/-的表达,降低CD25-CD127-的表达,从而抑制免疫反应。     

类风湿关节炎患者Th17和CD161+Th17细胞水平的变化

目的 观察类风湿关节炎(RA)患者和健康对照组外周血中CD4+ IL-17+T细胞(Th17)和CD4+ CD161+ IL-17+T细胞(CD161+Th17)的水平,探讨其在RA发病过程中的意义.方法 采用流式细胞术测定36例RA患者和11例健康对照组外周血中Th17细胞及CD161+ Th17的细胞百分率.结果 RA患者外周血Th17及CD161+ Th17细胞水平明显高于健康对照组(P＜0.01)且与疾病活动指数(DAS28)、血沉和C-反应蛋白水平呈显著正相关(P ＜0.05);RA患者和健康对照组外周血中CD161+ Th17细胞百分率明显高于Th17细胞百分率(P＜0.01);Th17细胞百分率与CD161+ Th17细胞百分率显著正相关(P＜0.01).结论 Th17及CD161+ Th17细胞在RA患者外周血中均增高且与疾病活动度正相关.     

卵巢癌患者CD4+CD25+调节性T细胞的测定以及与细胞因子关系的研究

目的 探讨卵巢癌患者外周血CD4+CD25+调节性T细胞(Treg)在CD4+T细胞的分布,分析CD4+CD25+Treg与卵巢癌患者临床病理与生理特征的关系;研究CD4+CD25+Treg与TH1、TH2类细胞因子的关系.方法 流式细胞仪测定53例卵巢癌患者、44例卵巢良性疾患、45例健康人外周血CD4+CD25+Treg比例,TH1类细胞因子白细胞介素-2(IL-2)、γ-干扰素(IFN-γ)、肿瘤坏死因子-α(TNF-α)和TH2类细胞因子IL-4、IL-6、IL-10的水平.结果 (1)卵巢癌患者外周血CD4+CD25+Treg(21.95%±6.80%)明显高于卵巢良性疾患(12.57%±5.12%)和健康对照组(10.77%±3.02%),P均＜0.001;良性疾患与健康对照组比较差异无统计学意义;手术前的卵巢癌患者CD4+CD25+Treg(21.95%±6.80%)明显高于手术后的患者(16.41%±5.84%),P＜0.001.(2)CD4+CD25+Treg与患者的年龄、职业没有关系;卵巢浆液性腺癌CD4+CD25+Treg为23.58%±7.08%,明显高于混合性瘤(18.30%±5.46%),差异有统计学意义(P＜0.05);与黏液性腺癌(22.06%±6.08%)差异无统计学意义;Ⅳ期卵巢癌患者CD4+CD25+Treg比值(28.79%±4.26%)明显高于Ⅲ期(20.01%±6.27%)和Ⅰ～Ⅱ期患者(17.97%±4.50%),P均＜0.001;CD4+CD25+Treg与肿瘤的远处转移(23.61%±6.52%)、淋巴结转移(24.05%±6.74%)、腹水的形成(24.06%±6.73%)有关(P＜0.05);低分化的卵巢癌患者CD4+CD25+Treg(25.05%±5.96%)与高分化患者(16.03%±0.80%)比较,差异有统计学意义(P＜0.05),而与中分化(21.82%±6.53%)比较差异无统计学意义.(3)卵巢癌患者TH1类细胞因子IL-2、TNF-α、IFN-γ的水平[(0.87±0.25)pg/ml、(1.09±0.34)pg/ml)、(7.56±2.25)pg/ml]明显低于健康对照[(2.38±1.09)pg/ml、(3.28±1.36)pg/ml、(19.04±4.59)pg/ml],P均＜0.001.而TH2类细胞因子IL-6的水平[(86.28±55.92)pg/ml]与对照组[(5.04±2.69)pg/ml)]比较显著升高(P＜0.001),而IL-4[(4.86±2.94)pg/ml]、IL-10[(1.07±2.30)pg/ml与健康人[(5.42±3.59)pg/ml、(4.46±2.31)pg/ml]之间差异无统计学意义.通过相关分析,卵巢癌患者外周血CD4+CD25+Treg与TH1类细胞因子IL-2、TNF-α、IFN-γ存在负相关,相关系数分别为r=-0.658,P＜0.05;r=-0.591,P＜0.05;r=-0.695,P＜0.05.与TH2类细胞因子IL-6存在正相关,相关系数r=0.784,P＜0.001.结论 (1)卵巢癌患者外周血CD4+CD25+Treg数量显著增加,这可能是卵巢癌患者免疫功能下降的一个重要原因;(2)CD4+CD25+Treg数量与肿瘤的分期、组织学类型、分化程度、浸润、淋巴结转移、腹水的形成有关.(3)卵巢癌患者外周血CD4+CD25+Treg与TH1类细胞因子之间存在着负相关,而与TH2类细胞因子IL-6之间存在正相关.CD4+CD25+Treg可能通过TH1、TH2类细胞因子的调节发挥对效应性T细胞的抑制作用.     

慢性乙型肝炎患者外周血CD4~+CD25~+Treg与CD4~+和CD8~+ T淋巴细胞亚群的相关研究

目的:探讨慢性乙型肝炎患者外周血中CD4+CD25+调节性T细胞的含量和CD4+CD8+T淋巴细胞亚群分布,两者之间相关性及与HBV的相关性。方法:采用流式细胞术检测50例慢性乙型肝炎患者和20例健康对照者外周血中CD4+CD25high、CD4+CD25+Foxp3+Treg细胞表达及CD3/CD4/CD8 T淋巴细胞亚群,荧光定量PCR法检测HBV DNA含量。结果:慢性乙型肝炎患者外周血中CD4+CD25highTreg明显高于健康对照组(P0.01),且随HBV DNA载量增加,患者外周血中CD4+CD25highTreg细胞的水平逐渐升高。慢性乙型肝炎患者外周血中CD4+CD25+Foxp3+Treg细胞也相应增高,且与CD4+CD25highTreg细胞的变化成正相关(r=0.890,P0.001)。与健康对照组比较,患者组CD4+T细胞百分率及CD4+/CD8+比值均降低,而CD3+T细胞和CD8+T细胞百分率差异无显著性(P0.05)。CD4+CD25highTreg细胞与HBV DNA取对数后成正相关(r=0.782,P0.001),与谷丙转氨酶(ALT)成正相关(r=0.432,P0.005);与CD3+、CD4+、CD8+T细胞水平及CD4+/CD8+比值均无相关性(P0.05)。CD3+、CD4+、CD8+T淋巴细胞及CD4+/CD8+比值与HBV DNA载量之间亦无相关性(P0.05)。结论:慢性乙型肝炎患者外周血中CD4+CD25+Treg细胞增高,且与HBV的复制水平及ALT增高具有一致性,而T细胞亚群是否可作为监测CHB患者免疫状态的指标需进一步探讨。     

蕈样肉芽肿患者外周血单一核细胞CD45RA及CD45RO表达的研究

目的探讨蕈样肉芽肿（Mycosis fungoides,MF）患者外周血单个核细胞CD45RA及CD45RO的表达及其与MF发病的关系。方法应用双荧光抗体标记、流式细胞仪检测15例MF患者外周血单个核细胞CD45RA及CD45RO的表达。结果（1）MF患者外周血CD3^＋、CD3^＋CD8^＋细胞与正常对照比较差异不显著（P〉0.05）。（2）MF患者外周血CD4^＋细胞低于正常对照,差异非常显著（P〈0.001）。（3）MF患者外周血T细胞CD3^＋CD4^＋/CD3^＋CD8^＋比值低于正常对照,差异显著（P〈0.001）。（4）MF患者外周血CD45RA^＋细胞低于正常对照,差异非常显著（P〈0.001）,CD45RO^＋细胞高于正常对照,差异非常显著（P〈0.001）。（5）MF患者外周血CD45RO^＋/CD45RA^＋比值高于正常对照,差异非常显著（P〈0.001）。（6）MF患者外周血CD4^＋CD45RA^＋细胞低于正常对照,差异非常显著（P〈0.001）。（7）MF患者外周血CD4^＋CD45RO^＋细胞及CD8^＋CD45RO^＋细胞均高于正常对照,差异非常显著（均P〈0.001）。（8）MF患者外周血CD4^＋CD45RO^＋/CD4^＋CD45RA^＋比值及CD8^＋CD45RO^＋/CD8^＋CD45RA^＋比值均高于正常对照,差异非常显著（P〈0.01及P〈0.001）。结论MF患者外周血中,不仅存在CD4^＋亚群失调和CD4^＋/CD8^＋比值降低,而且在CD4^＋和CD8^＋亚群中也存在CD45RA^＋、CD45RO^＋亚群失调和CD45RO^＋/CD45RA^＋比值升高,从而导致的机体免疫功能紊乱,可能与MF的发病或病情加剧有关。     

慢性乙型肝炎患者外周血CD45RA+、CD45RO+T淋巴细胞亚群的特点及临床意义

目的：探讨慢性乙型肝炎患者外周血CD4^＋CD45RA^＋、CD4^＋CD45RO^＋、CD8^＋CD45RA^＋和CD8^＋CD45RO^＋T淋巴细胞亚群的特点及其与肝病病情的关系。方法：采集46例轻中度慢性乙型肝炎患者、58例重度慢性乙型肝炎患者和30例健康人的外周抗凝血，应用流式细胞技术三色荧光分析法对其外周血中CD4^＋CD45RA^＋、CD4^＋CD45RO^＋、CD8^＋CD45RA^＋和CD8^＋CD45RO＋T淋巴细胞亚群进行检测。结果：轻中、重度慢性乙型肝炎患者与正常人相比，其外周血中CD4^＋、CD8^＋T细胞均无明显改变；CD8^＋CD45RA^＋T细胞均明显降低，CD8^＋CD45RO^＋T细胞均明显增高，而CD4^＋CD45RA^＋、CD4^＋CD45RO^＋T细胞均无明显改变；重度慢性乙型肝炎患者与轻中度慢性乙型肝炎患者相比，CD8^＋CD45RA^＋T细胞明显降低（P〈0.05），CD8^＋CD45RO^＋T细胞明显升高（P〈0.05）。结论：乙型肝炎慢性化过程中，CD8^＋CD45RO^＋T细胞起重要作用且与慢性乙型肝炎患者病情的进展呈正相关；检测CD4^＋CD45RA^＋、CD4^＋CD45RO^＋、CD8^＋CD45RA^＋和CD8^＋CD45RO^＋T淋巴细胞亚群比检测CD4^＋和CD8^＋T细胞亚群更能正确、充分、全面地了解慢性乙型肝炎的发病机制和预后，从而有效地指导临床治疗。     

Immune memory in CD4+ CD45RA+ T cells.

This study addresses the question of whether human peripheral CD4+ CD45RA+ T cells possess antigen-specific immune memory. CD4+ CD45RA+ T cells were isolated by a combination of positive and negative selection. Putative CD4+ CD45RA+ cells expressed CD45RA (98.9%) and contained < 0.1% CD4+ CD45RO+ and < 0.5% CD4+ CD45RA+ CD45RO+ cells. Putative CD45RO+ cells expressed CD45RO (90%) and contained 9% CD45RA+ CD45RO+ and < 0.1% CD4+ CD45RA+ cells. The responder frequency of Dermatophagoides pteronyssinus-stimulated CD4+ CD45RA+ and CD4+ CD45RO+ T cells was determined in two atopic donors and found to be 1:11,314 and 1:8031 for CD4+ CD45RA+ and 1:1463 and 1:1408 for CD4+ CD45RO+ T cells. The responder frequencies of CD4+ CD45RA+ and CD4+ CD45RO+ T cells from two non-atopic, but exposed, donors were 1:78031 and 1:176,903 for CD4+ CD45RA+ and 1:9136 and 1:13,136 for CD4+ CD45RO+ T cells. T cells specific for D. pteronyssinus were cloned at limiting dilution following 10 days of bulk culture with D. pteronyssinus antigen. Sixty-eight clones were obtained from CD4+ CD45RO+ and 24 from CD4+ CD45RA+ T cells. All clones were CD3+ CD4+ CD45RO+ and proliferated in response to D. pteronyssinus antigens. Of 40 clones tested, none responded to Tubercule bacillus purified protein derivative (PPD). No difference was seen in the pattern of interleukin-4 (IL-4) or interferon-gamma (IFN-gamma) producing clones derived from CD4+ CD45RA+ and CD4+ CD45RO+ precursors, although freshly isolated and polyclonally activated CD4+ CD45RA+ T cells produced 20-30-fold lower levels of IL-4 and IFN-gamma than their CD4+ CD45RO+ counterparts. Sixty per cent of the clones used the same pool of V beta genes. These data support the hypothesis that immune memory resides in CD4+ CD45RA+ as well as CD4+ CD45RO+ T cells during the chronic immune response to inhaled antigen.     

Differential infiltration by CD45RO and CD45RA subsets of T cells associated with human heart allograft rejection.

Subsets of T cells express different isoforms of the leukocyte common antigen CD45; those expressing the glycoprotein 220 isoform (CD45RA) have been characterized as naive in their response to antigens, and those expressing the glycoprotein 180 isoform (CD45RO) as memory T cells. The association between the rejection status of human cardiac allograft recipients and the relative infiltration of the CD45 subsets of both CD8+ and CD4+ T cells was examined using two-color immunohistological labeling techniques on 33 heart transplant biopsies, categorized by routine histological and clinical criteria as mild (requiring no treatment) or moderate (requiring antirejection therapy) rejection. Double-labeling was performed using pairs of monoclonal antibodies to define the following populations: CD4+ CD45RA+, CD4+ CD45RO+, CD8+ CD45RA+, and CD8+-CD45RO+. The number of cells per high-power field (HPF) for each of these cell subsets was counted in every biopsy. In cases with mild rejection, infiltration was predominant for CD4+ CD45RA+ cells (median = 5.0 cells/HPF) relative to CD4+ CD45RO+ (3.12 cells/HPF), CD8+ CD45RA+ (2.14 cells/HPF), and especially CD8+ CD45RO+ (1.22 cells/HPF) populations. In cases with moderate rejection, all four subpopulations increased but were essentially equivalent in intensity, such that in comparison to cases with mild rejection, the smallest increase was seen for CD4+ CD45RA+ cells (6.67 cells/HPF, P < 0.09) and the greatest for CD8+ CD45RO+ cells (7.00 cells/HPF, P < 0.002). A majority of CD8 cells expressed CD45RA in 14 of 16 (88%) cases of mild rejection compared to only 2 of 17 cases of moderate rejection. Moreover, the ratio of CD45RO+ to CD45RA+ cells in each biopsy was higher in moderate versus mild rejection for both CD4 (median ratios = 1.13 versus 0.68, respectively; P < 0.008) and CD8 (1.43 versus 0.58, respectively; P < 0.005) subsets. A majority of T cells expressed CD45RO in cases of moderate rejection (11 of 14 or 79%), compared to only 1 of 13 (8%) cases of mild rejection. These findings indicate that during generally self-limited mild acute cardiac allograft rejection there is a predominance of naive CD45RA+ T cells, especially of the CD4 phenotype, whereas during moderate rejection there is a significant shift toward activated CD45RO+ T cells, especially in the CD8 population.     

Dual expression of CD45RA and CD45RO isoforms on myelin basic protein-specific CD4+ T-cell lines in multiple sclerosis

Myelin basic protein (MBP)-specific T-cell lines from patients with multiple sclerosis (MS) and healthy controls were analyzed for the expression of CD45 isoforms and adhesion molecules. In the multiple sclerosis group, 22 of 24 MBP-specific T-cell lines were CD4+. Two distinct patterns were observed with regard to CD45 isoform expression. Pattern I showed dual expression of CD45 isoforms (CD4+CD45RA+CD45RO+CD29+) and Pattern II included cells with a single CD45 isoform (CD4+CD45RA–CD45RO+CD29+). All 10 cell lines from healthy controls were CD4+ and displayed Pattern II (CD4+CD45RA–CD45RO+CD29+). The dual expression of CD45 isoform in T-cell lines from MS was stable, did not represent a transition stage from CD45RA to CD45RO, and was cell-cycle independent. All cell lines from MS and controls expressed increased levels of LFA-1 (CD11a), LFA-2 (CD2), LFA-3 (CD58), ICAM-1 (CD54), and VLA-4 (CDw49d). These data show the presence of unique MBP-specific T cells (CD4+CD45RA+CD45RO+CD29+) that might play a role in the pathogenesis of MS.     

Phenotypic switch from CD45RA+ to CD45RA- by normal blood T cells is associated with increased HLA-ABC expression for CD4+ and CD8+ populations but not for the NK-associated CD4-CD8dim+ or CD4-CD8- fractions.

Using the combined techniques of immunomagnetic depletion and multiple colour flow cytometry, the expression of HLA-ABC (W6/32) by normal T-cell subpopulations, defined by 2H4 (CD45RA) expression, was examined. It is thought that a CD45RA+CD45RO- phenotype defines the 'virgin' T-cell fraction, whereas a CD45RA-CD45RO+ phenotype defines the 'primed' or memory T-cell population. In addition, an intermediate phenotype (CD45RA+CD45RO+) appears to correspond to a transitional stage of development. In this study, these three phenotypic stages were represented by distinct levels of 2H4 staining defined as 2H4+, 2H4int and 2H4-, respectively. The results of this current investigation are of importance in two main areas. Firstly, when compared to the 2H4+ component, the HLA-ABC expression of 2H4- cells was significantly higher. This was true for both CD4+CD8- and CD4-CD8+ lymphocytes, but was not the case for CD4-CD8dim+, CD3+CD4-CD8- and CD3-CD4-CD8- fractions. Additionally, when HLA-ABC expression was examined as a function of 2H4 staining intensity, it was found that, for the CD4+ fraction, the greatest increase in HLA-ABC expression occurred between the 2H4int and 2H4- stages. In contrast, the increase in HLA-ABC expression by CD8+ lymphocytes was associated with transition from 2H4+ to 2H4int status, which suggests that increased HLA-ABC expression occurs at an earlier stage in the acquisition of CD45RO in CD8+ cells than for CD4+ cells. Secondly, for each individual blood examined, a close and highly significant correlation (P = 0.002) for membrane HLA-ABC expression was found between (i) CD4+2H4+ and CD8+2H4+ and (ii) CD4+2H4- and CD8+2H4- subpopulations. This suggests that modulation of HLA-ABC expression in CD4+ and CD8+ cells is subject to common control mechanisms and remains proportionate for these lymphocyte fractions in any given individual.     

CD45 isoform expression in autoimmune myasthenia gravis

In myasthenia gravis (MG), humoral and cellular immune mechanisms are involved in the autoimmune pathogenesis. In this study, we investigated the role of the CD45 molecule in MG, having recently reported an association in multiple sclerosis. CD45, a protein-tyrosine phophatase receptor type C (PTPRC), is essential for both thymic selection and peripheral activation of T and B cells. Our aims were to determine (a) the prevalence of a functional mutation in the CD45 gene (exon 4 77C --> G; prevalence analysis), and (b) the distribution of memory (CD45RO+) and naive (CD45RA+) T cells in the peripheral blood (subset analysis). T cells from 78 patients with generalised MG were stained with monoclonal antibodies against CD45RO, CD45RA, CD4 and CD8 and quantified by four-colour flow cytometry. The control panel for the prevalence analysis (a) consisted of 303 healthy individuals. (b) From those, 67 age- and sex-matched probands were randomly selected as controls for the subset analysis. Patients were stratified according to their MG onset age, thymic pathology and immunosuppressive treatment. Statistical analysis was performed by Fisher's exact test, asymptotic chi2 test, the two-sided Mann-Whitney test and Spearman's correlation coefficient. As a result, the 77C --> G mutation in exon 4 of the CD45 gene was found in 1 of 78 patients versus none of the 303 controls. Thus, no association was detected with this single nucleotide polymorphism in MG patients overall. Surprisingly, however, ratios of CD45RO+ to CD45RA+ T cells were lower among CD8+ T cells from patients with late-onset MG (P = 0.023). Thymoma patients also showed a similar trend among CD4+ and CD8+ T-cells, as expected. These differences were not related to immunosuppressive drug treatment or thymectomy (in the 67 informative patients). Since there is no other evidence for increased thymopoiesis in late-onset MG, we propose an altered subset balance in the circulation.     

Distribution of Alloreactivity Amongst the CD45 Isoforms of Circulating CD4 and CD8 T Lymphocytes

The expression of different isoforms of the CD45 surface molecule allows lymphocytes to be divided into two nonoverlapping categories, CD45RA and CD45RO. Previous studies of CD4 T cells have shown that responses to soluble antigens are present predominantly in the RO subset and to mitogens in the RA, alloreactivity being present in both subsets. Responses of CD8 T cells have not been investigated in such detail, nor have responses been compared to CD4 cells. Here we report the alloreactive responses of both CD45RA and RO subsets amongst both CD4 and CD8 T lymphocytes. Following isolation of CD4 and CD8 cells with immunomagnetic beads, CD45 subsets were separated by negative depletion using specific monoclonal antibodies. CD45RA populations were greater than 97% pure and CD45RO cells greater than 91%. One-way primary mixed lymphocyte reactions were established using the purified responder cells with irradiated allogeneic peripheral blood mononuclear cells as stimulators; experiments were all repeated at least three times. In assays of CD4⁺ RA and RO subsets, reactivity was present in both isoforms, being consistently, but not significantly, greater amongst the RO subset. With CD8⁺ T cells, reactivity was also present in both isoforms, but was significantly greater in the CD45RA subset, with mean proliferation 2.5–3-fold that of the CD45RO cells ( P  < 0.05).     

CD45 Isoform Expression in Autoimmune Myasthenia Gravis

In myasthenia gravis (MG), humoral and cellular immune mechanisms are involved in the autoimmune pathogenesis. In this study, we investigated the role of the CD45 molecule in MG, having recently reported an association in multiple sclerosis. CD45, a protein-tyrosine phophatase receptor type C (PTPRC), is essential for both thymic selection and peripheral activation of T and B cells. Our aims were to determine (a) the prevalence of a functional mutation in the CD45 gene (exon 4 77C &#77 G; prevalence analysis), and (b) the distribution of memory (CD45RO+) and naïve (CD45RA+) T cells in the peripheral blood (subset analysis). T cells from 78 patients with generalised MG were stained with monoclonal antibodies against CD45RO, CD45RA, CD4 and CD8 and quantified by four-colour flow cytometry. The control panel for the prevalence analysis (a) consisted of 303 healthy individuals. (b) From those, 67 age- and sex-matched probands were randomly selected as controls for the subset analysis. Patients were stratified according to their MG onset age, thymic pathology and immunosuppressive treatment. Statistical analysis was performed by Fisher's exact test, asymptotic &#104 2 test, the two-sided Mann-Whitney test and Spearman's correlation coefficient. As a result, the 77C &#77 G mutation in exon 4 of the CD45 gene was found in 1 of 78 patients versus none of the 303 controls. Thus, no association was detected with this single nucleotide polymorphism in MG patients overall. Surprisingly, however, ratios of CD45RO+ to CD45RA+ T cells were lower among CD8+ T cells from patients with late-onset MG ( P =0.023). Thymoma patients also showed a similar trend among CD4+ and CD8+ T-cells, as expected. These differences were not related to immunosuppressive drug treatment or thymectomy (in the 67 informative patients). Since there is no other evidence for increased thymopoiesis in late-onset MG, we propose an altered subset balance in the circulation.     

类风湿关节炎患者Th17细胞与调节性T细胞失衡的研究

目的观察类风湿性关节炎（RA）患者外周血Th17细胞与CD4^＋CD25^＋FoxP3^＋调节性T细胞（Treg）平衡状态与疾病的关系,分析Th17／Treg细胞免疫失衡在RA发病机制中的作用。方法采用流式细胞仪四色荧光抗体标记法分别对47例RA患者和39名健康志愿者（HVs）进行CD3、CD8、IL-17与CD4、CD25、F0xP3标记,测定Th17与调节性T细胞的比例变化及相关细胞因子IL-6、IL-23和IL-17水平。结果RA组患者外周血中,CD3^＋CD8^＋IL-17^＋T细胞占CD3^＋T淋巴细胞的百分比为（1．12±0．38）％,明显高于对照组（0．68±0．29）％（t=1．83,P〈0．05）;CD4’CD25’FoxP3^＋细胞占CD4^＋T淋巴细胞的百分比为（2．74±0．71）％,明显低于对照组（4．69±1．23）％（t=-2．94,P〈0．05）。相关细胞因子测定结果：IL-6水平在RA组为（13．5±3．7）ng／L,正常人为（4．6±0．9）ng／L（t=6．24,P〈0．01）;IL-23水平在RA组为（71±19）ng／L,正常人为（25±6）ng／L（t=14．37,P〈0．01）;IL-17水平在RA组为（122±33）ng／L,正常人为（37±9）ng／L（t=19．01,P〈0．01）;RA患者血清IL-6、IL-23和IL-17水平均明显升高。结论RA患者外周血Th17与CD4^＋CD25^＋FoxP3^＋调节性T细胞数量的异常可能是RA发病的重要因素,IL-6和IL-23的升高是引起这些改变的可能原因。