CD4＋T淋巴细胞对肺结核患者血清白蛋白的影响

目的 探讨外周血CD4⁺ T淋巴细胞对肺结核患者血清白蛋白的影响。 方法 对119例初治肺结核患者进行淋巴细胞亚群和血清白蛋白测定,探讨细胞免疫功能与血清白蛋白相关性;同时,根据其CD4⁺ T淋巴细胞亚群水平,分为免疫低下组62例和正常组57例,探讨细胞免疫功能低下对低蛋白血症的影响。 结果 （1）所有纳入患者外周血CD4⁺ T淋巴细胞［（516.6±266.1）×10⁹/L］与CD3⁺ T淋巴细胞［（841.6±398.8）×10⁹/L］、CD8⁺ T淋巴细胞［（261.0±142.6）×10⁹/L］、CD4⁺/CD8⁺ T淋巴细胞比值（2.33±1.40）呈正相关（分别为r=0.883,P=0.000;r=0.579,P=0.000;r=0.365,P=0.000）。（2）外周血CD4⁺T淋巴细胞与血清白蛋白呈正相关（r=0.116,P=0.033）。（3）在免疫低下组中,肺部病灶范围达4个肺野以上者43例（占69.4%）,明显高于正常组的22例（占38.6%）,差异有统计学意义（χ²=11.335,P=0.001）;免疫低下组治疗前血清白蛋白［（33.9±5.5）g/L］明显低于正常组［（36.1±5.7）g/L］（t=2.187,P=0.031）。 结论 在肺结核患者中CD4⁺ T淋巴细胞与治疗前血清白蛋白水平呈正相关。CD4⁺ T淋巴细胞低下肺结核患者,其病灶范围较广,容易并发低蛋白血症。     

肺结核及肺外结核患者外周血T淋巴细胞亚群的变化及其临床意义

目的 探讨肺外结核患者外周血CD3 ⁺、CD4 ⁺、CD8 ⁺T淋巴细胞亚群及CD4 ⁺/CD8 ⁺比值的变化及其临床意义方法 搜集2017年11月至2019年1月在成都市公共卫生临床医疗中心结核病区住院的147例患者(包括63例单纯肺结核、60例肺结核并发肺外结核和24例单纯肺外结核患者,分别为肺结核组、肺结核并发肺外结核组和肺外结核组)作为研究对象。采用流式细胞仪及四色淋巴细胞亚群检测试剂盒检测纳入的各组患者外周血CD3 ⁺、CD4 ⁺、CD8 ⁺T淋巴细胞的绝对计数及CD4 ⁺/CD8 ⁺比值,分析上述各组患者外周血T淋巴细胞亚群的差异。结果 肺结核并发肺外结核患者外周血CD3 ⁺和CD4 ⁺T淋巴细胞计数分别为596.000(377.500,823.250)个/μl和347.500(214.000,479.250)个/μl,均低于单纯肺结核患者的698.000(572.000,904.000)个/μl和409.000(311.000,545.000)个/μl,差异均有统计学意义(Z=-2.507,P=0.012;Z=-2.431,P=0.015)。肺结核并发肺外结核患者的CD8 ⁺T淋巴细胞计数与CD4 ⁺/CD8 ⁺比值分别为195.500(137.250,278.750)个/μl和1.670(1.258,2.273),较肺结核患者的244.000(154.000,317.000)个/μl和1.770(1.290,2.350)降低,但差异均无统计学意义(Z=-1.892,P=0.058;Z=-0.546,P=0.585)。单纯肺外结核患者外周血CD3 ⁺和CD4 ⁺ T淋巴细胞计数分别为551.500(283.750,949.000)个/μl和295.500(134.250,461.750)个/μl,低于单纯肺结核患者的698.000(572.000,904.000)个/μl和409.000(311.000,545.000)个/μl,差异均有统计学意义(Z=-2.089,P=0.037;Z=-2.460,P=0.014);单纯肺外结核患者外周血的CD8 ⁺T淋巴细胞计数[185.000(92.000,366.250)个/μl]及CD4 ⁺/CD8 ⁺比值[1.455(1.018,2.128)]与单纯肺结核患者[分别为244.000(154.000,317.000)个/μl和1.770(1.290,2.350)]相比,差异均无统计学意义(Z=-1.315,P=0.188;Z=-1.429,P=0.153)。结论 随着肺结核患者外周血CD3 ⁺、CD4 ⁺ T淋巴细胞计数的逐步降低,结核分枝杆菌可能更易播散至肺外;外周血CD8 ⁺T淋巴细胞计数及CD4 ⁺/CD8 ⁺比值在肺外结核发生机制中的作用尚需进一步探讨。     

肺结核患者疗程中外周血辅助性和调节性T细胞的动态研究

目的 探讨辅助性T细胞17（Th17）和CD4⁺CD25⁺CD127 ^low调节性T细胞与结核病的发病及抗结核治疗转归的关系。 方法 纳入对象包括32例活动性肺结核患者、25例Mtb潜伏感染者、45例健康对照者。流式细胞术检测外周血Th17细胞分泌细胞因子IL-17和CD4⁺CD25⁺CD127 ^low调节性T细胞表达强度。结果以x±s表示,所有数据均使用Prism 4.0统计软件进行分析,两组间比较采用非配对t检验,多组间的比较采用ANOVA方差分析,以P<0.05为差异有统计学意义。 结果 肺结核患者组治疗前IL-17表达为（3.25±1.68）%,明显低于健康对照组［（4.62±1.46）%］（F=6.633,P<0.0001）。比较活动性肺结核患者组IL-17表达在治疗前、治疗3个月［（4.17±2.27）%］、治疗6个月［（5.58±1.66）%］时的检测结果,发现治疗3个月时高于治疗前,但差异无统计学意义（F=12.244,P=0.057）;而治疗6个月时明显高于治疗前（F=12.244,P<0.0001）和治疗3个月时（F=12.244,P=0.004）。健康对照组CD4⁺CD25⁺CD127 ^low调节性T细胞表达为（4.97±1.60）%,与Mtb潜伏感染组［（5.00±1.08）%］比较,差异无统计学意义（F=11.986,P=0.937）;活动性肺结核患者组治疗前表达为（6.59±1.73）%,显著高于健康对照组及Mtb潜伏感染组(F=11.986,P<0.0001)。比较活动性肺结核患者组治疗前、治疗3个月［（8.28±2.04）%］、治疗6个月［（7.46±1.87）%］时的CD4⁺CD25⁺CD127 ^low调节性T细胞表达,发现治疗3个月时的表达明显高于治疗前（F=6.458,P=0.001）;治疗6个月时的表达低于治疗3个月时（F=6.458, P=0.085）,但差异无统计学意义;治疗6个月与治疗前的表达相比较,差异无统计学意义（F=6.458,P=0.068）;治疗6个月CD4⁺CD25⁺CD127 ^low调节性T细胞表达明显高于健康对照组（t=6.255,P<0.0001）。 结论 结核病患者外周血Th17细胞明显减少,经有效抗结核治疗后,Th17细胞逐渐增加,表明Th17细胞在抗结核免疫中起保护作用。结核病患者外周血CD4⁺CD25⁺CD127 ^low调节性T细胞明显增多,经抗结核治疗后CD4⁺CD25⁺CD127 ^low调节性T细胞逐渐减少,进一步说明CD4⁺CD25⁺CD127 ^low调节性T细胞在抗结核免疫中起抑制作用。     

CD4+CD25+调节性T细胞对肺结核患者特异细胞免疫的调节作用

目的　探讨调节性T细胞对结核患者特异性细胞免疫的调节作用及其在结核病发生中的意义。方法 采用免疫磁珠从健康对照和结核患者外周血单个核细胞中分离CD4⁺CD25⁺调节性T细胞,观察其对结核患者外周血免疫反应,包括细胞增殖反应和细胞因子IFN-γ及IL-10分泌的影响。结果 体外消除CD4⁺CD25⁺调节性T细胞没有显著影响健康对照PBMC对BCG抗原的增殖反应,但可以显著增强结核患者PBMC对BCG抗原的细胞增殖反应、细胞因子IFN-γ及IL-10的分泌。分离的CD4⁺CD25⁺调节性T细胞能显著抑制结核患者CD4⁺CD25^-T细胞对BCG抗原及抗-CD3的细胞增殖和细胞因子IFN-γ分泌;CD4⁺CD25⁺调节性T细胞也抑制BCG刺激的CD4⁺CD25^-T细胞IL-10的分泌,但不影响抗-CD3刺激的IL-10分泌。结论 CD4⁺CD25⁺调节性T细胞可能通过抑制结核患者特异性细胞免疫应答促进肺结核病的发生发展。     

重症肺结核患者T细胞免疫球蛋白黏蛋白分子的表达及其预后意义

目的 探讨重症肺结核患者T淋巴细胞亚群、T细胞免疫球蛋白黏蛋白分子(TIM-1,TIM-3)的表达及其预后意义。方法 回顾性分析2019年1月至2021年6月在我院收治的1 056例重症肺结核患者为研究对象,并将其定义为重症肺结核组,另选同期在我院进行治疗的1 000名轻症肺结核患者作为轻症肺结核组。比较两组患者TIM-1、TIM-3以及外周血T细胞亚群(CD4⁺、CD8⁺)水平;检测并比较外周血单个核细胞中TIM-1、TIM-3的mRNA水平;分析重症肺结核患者外周血T细胞亚群、TIM-1、TIM-3与患者预后的关系。结果 重症肺结核组CD8⁺、TIM-1和TIM-3水平均高于轻症肺结核组,CD4⁺水平低于轻症肺结核组(均P<0.001)。重症肺结核组TIM-1 mRNA和TIM-3 mRNA水平均高于轻症肺结核组(均P<0.001)。死亡组CD8⁺、TIM-1和TIM-3水平均高于存活组,CD4⁺水平低于存活组(均P<0.001)。相关性分析发现:存活组患者TIM-1、TIM-3与CD4⁺均呈负相关(r值分别为-0.114和-0.211,P<0.05),而与CD8⁺均呈正相关(r值分别为0.571和0.680,P<0.05)。结论 重症肺结核患者免疫功能紊乱可能与其体内外周血T细胞亚群水平异常以及TIM-1、TIM-3 表达水平升高有关。     

肺结核患者临床特征与外周血流式细胞亚群的相关性分析

目的 了解肺结核患者临床特征与外周血流式细胞亚群[T淋巴细胞亚群及自然杀伤 (NK)细胞]的相关性。方法 连续性收集2019年1月1日至5月25日期间同济大学附属上海市肺科医院肺结核住院患者1000例的临床资料进行回顾性分析。将患者各临床特征数据与外周血流式细胞亚群的检测值采用Eviews 8.0软件分别建立多元线性逐步回归模型,以明确各临床特征与外周血流式细胞亚群的检测值之间的相关性。结果 (1)肺结核患者的肺部病灶范围及呼吸道标本抗酸染色涂片结果与CD3⁺T细胞表达有关:肺部病灶范围越大、呼吸道标本抗酸染色涂片阳性程度越高,CD3⁺ T细胞数量越低(回归系数分别为-0.255、-0.499, P值分别为0.021、0.027)。(2)患者的年龄、性别、结核分子生物学检测结果与CD4⁺ T细胞表达有关: 0~<20岁年龄段患者CD4⁺ T细胞数量低于其他年龄段患者(回归系数-4.710,P=0.031);男性CD4⁺ T细胞数量低于女性患者(回归系数-2.150, P=0.001);分子生物学检测阳性的患者CD4⁺ T细胞数量高于检测阴性的患者(回归系数1.433, P=0.030)。(3)初治患者CD8⁺ T细胞数量高于复治患者(回归系数1.247, P=0.029);呼吸道标本抗酸染色涂片阳性程度越高CD8⁺ T细胞数量越高(回归系数0.442, P=0.033)。(4)并发肺外结核者的NK细胞低于未并发肺外结核者(回归系数0.375, P=0.030)。结论 肺结核患者的肺部病灶累及范围、呼吸道标本抗酸染色阳性与外周血流式细胞亚群CD3⁺ T细胞具有相关性;年龄、性别、结核分子生物学检测结果与CD4⁺ T细胞具有相关性;是否初治、呼吸道标本抗酸染色阳性与CD8⁺ T细胞具有相关性;是否并发肺外结核与NK细胞表达水平具有相关性。     

Correlation analysis between clinical characteristics and flow cytometry cells subsets in peripheral blood of patients with pulmonary tuberculosis

目的 了解肺结核患者临床特征与外周血流式细胞亚群[T淋巴细胞亚群及自然杀伤 (NK)细胞]的相关性。方法 连续性收集2019年1月1日至5月25日期间同济大学附属上海市肺科医院肺结核住院患者1000例的临床资料进行回顾性分析。将患者各临床特征数据与外周血流式细胞亚群的检测值采用Eviews 8.0软件分别建立多元线性逐步回归模型,以明确各临床特征与外周血流式细胞亚群的检测值之间的相关性。结果 (1)肺结核患者的肺部病灶范围及呼吸道标本抗酸染色涂片结果与CD3⁺T细胞表达有关:肺部病灶范围越大、呼吸道标本抗酸染色涂片阳性程度越高,CD3⁺ T细胞数量越低(回归系数分别为-0.255、-0.499, P值分别为0.021、0.027)。(2)患者的年龄、性别、结核分子生物学检测结果与CD4⁺ T细胞表达有关: 0~<20岁年龄段患者CD4⁺ T细胞数量低于其他年龄段患者(回归系数-4.710,P=0.031);男性CD4⁺ T细胞数量低于女性患者(回归系数-2.150, P=0.001);分子生物学检测阳性的患者CD4⁺ T细胞数量高于检测阴性的患者(回归系数1.433, P=0.030)。(3)初治患者CD8⁺ T细胞数量高于复治患者(回归系数1.247, P=0.029);呼吸道标本抗酸染色涂片阳性程度越高CD8⁺ T细胞数量越高(回归系数0.442, P=0.033)。(4)并发肺外结核者的NK细胞低于未并发肺外结核者(回归系数0.375, P=0.030)。结论 肺结核患者的肺部病灶累及范围、呼吸道标本抗酸染色阳性与外周血流式细胞亚群CD3⁺ T细胞具有相关性;年龄、性别、结核分子生物学检测结果与CD4⁺ T细胞具有相关性;是否初治、呼吸道标本抗酸染色阳性与CD8⁺ T细胞具有相关性;是否并发肺外结核与NK细胞表达水平具有相关性。     

PBC患者CD8~+效应T细胞的细微表型与功能特征

【据Hepatoatology 2011年11月报道】题:在人类原发性胆汁性肝硬化患者中,CD8+效应T细胞的细微表型与功能特征（作者Tsuda M等）原发性胆汁性肝硬化（PBC）患者对丙酮酸脱氢酶E2（PDC-E2）产生了多种特异反应,包括自身反应性CD4⁺、CD8⁺T细胞和自身抗体等。美国加利福尼亚大学戴维斯分校的研究者检测了132例研究对象（包括76例PBC患者和56例对照）的CD8⁺T细胞表型。研究发现PBC患者外周血单核细胞中一种效应记忆T细胞（TEM）出现频率较高,其细胞表型的特征是表达CD45RO^highCD57⁺CD8^high,同时也表达肠道归巢整合素、α4β7。这些CD8^high TEM细胞在TCR被激活后膜联蛋白V表达降低。CD45RO^highCD57⁺CD8^highT细胞较其他CD8^high T细胞表达更高水平的粒酶A、粒酶B、穿孔素、CCR5     

肺结核患者外周血免疫细胞水平与临床意义分析

目的 探讨肺结核患者外周血免疫细胞的表达水平及其临床意义。方法 回顾性分析2016年1月至2017年5月徐州市传染病医院收治的676例肺结核患者和体检中心的40例健康对照者。采用贝克曼库尔特公司FC500型流式细胞仪,以荧光微球为参照,分别检测外周血CD4⁺、CD8⁺、B和NK细胞的绝对数。结果 肺结核患者免疫细胞CD4⁺、CD8⁺、B和NK细胞数量低于健康对照者(t=6.25,5.12,5.05,4.37;P值均为0.00);痰菌阳性的肺结核患者免疫细胞CD4⁺、CD8⁺、B、NK细胞数量低于痰菌阴性的肺结核患者(t=4.07,3.70,2.75,2.71;P=0.00,0.00,0.01,0.01);老年肺结核患者免疫细胞CD4⁺、CD8⁺、B细胞数量低于年轻患者(t=4.04,2.60,5.25;P=0.00,0.01,0.00),554例初治肺结核患者经过2个月化疗后,其中510例单纯采用标准化抗结核治疗方案的患者CD4⁺、CD8⁺、B和NK细胞数量增加无统计学意义(t=1.19,1.04,0.39,1.82;P=0.23,0.30,0.70,0.07),44例合并免疫治疗的肺结核患者除NK细胞数量增加无统计学意义(t=0.80;P=0.42),CD4⁺、CD8⁺、B数量增加(t=2.15,2.17,3.12;P=0.03,0.03,0.00)。结论 肺结核患者免疫细胞的表达水平降低,痰菌阳性以及老年肺结核患者降低尤为明显,对于免疫细胞表达水平降低的肺结核患者,在化疗的基础上配合免疫治疗更有利于患者的康复。     

微波消融联合经肝动脉介入栓塞化疗和全身化疗治疗结直肠癌肝转移患者临床疗效比较*

目的 探讨采用微波消融联合经肝动脉栓塞化疗（TACE）治疗结直肠癌肝转移患者的临床疗效。方法 2012年6月~2015年6月我院收治的90例结直肠癌肝转移患者,45例对照组接受全身化疗治疗,而另45例观察组接受TACE联合微波消融治疗。比较两组患者应答情况及外周血淋巴细胞亚群的变化情况。结果 在治疗6个月末,观察组完全缓解（CR）、部分缓解（PR）、疾病稳定（SD）和疾病进展（PD）率分别为28.9%、40.0%、20.0%和11.1%,显著优于对照组的11.1%、28.9%、26.7%和33.3%（x²=7.571,P=0.006）;治疗前,两组患者外周血CD8⁺、CD4⁺和CD3⁺细胞百分比及CD4⁺/CD8⁺细胞比值比较,差异无统计学意义（P>0.05）,而在治疗后,观察组外周血CD3⁺和CD4⁺细胞百分比及CD4⁺/CD8⁺细胞比值改善情况显著优于对照组（t=7.256、t=5.916、t=7.701,P<0.01）;随访发现,对照组患者平均生存时间为（16.0±5.2）个月,显著短于观察组的（29.1±8.4）个月（t=8.895,P<0.001）。结论 采用微波消融术联合TACE治疗结直肠癌肝转移患者临床有效,能有效改善患者的免疫功能,延长生存期。     

Ex vivo analysis of human T lymphotropic virus type 1-specific CD4+ cells by use of a major histocompatibility complex class II tetramer composed of a neurological disease-susceptibility allele and its immunodominant peptide

HLA-DRB1*0101 is associated with susceptibility to human T lymphotropic virus type 1 (HTLV-1)-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Here, we used a synthetic tetramer of DRB1*0101 and its epitope peptide to analyze HTLV-1-specific CD4(+) T cells ex vivo. The frequency of tetramer(+)CD4(+) T cells was significantly greater in patients with HAM/TSP than in healthy HTLV-1 carriers (HCs) at a given proviral load and correlated with HTLV-1 tax messenger RNA expression in HCs but not in patients with HAM/TSP. These cells displayed an early to intermediate effector memory phenotype and were preferentially infected by HTLV-1. T cell receptor gene analyses of 2 unrelated DRB1*0101-positive patients with HAM/TSP showed similar Vbeta repertoires and amino acid motifs in complementarity-determining region 3. Our data suggest that efficient clonal expansion of virus-specific CD4(+) T cells in patients with HAM/TSP does not simply reflect higher viral burden but rather reflects a rapid turnover caused by preferential infection and/or in vivo stimulation by major histocompatibility complex-peptide complexes.     

Expanded cells in monoclonal TCR-alphabeta+/CD4+/NKa+/CD8-/+dim T-LGL lymphocytosis recognize hCMV antigens

Recent studies suggest the potential involvement of common antigenic stimuli on the ontogeny of monoclonal T-cell receptor (TCR)-alphabeta(+)/CD4(+)/NKa(+)/CD8(-/+dim) T-large granular lymphocyte (LGL) lymphocytosis. Because healthy persons show (oligo)clonal expansions of human cytomegalovirus (hCMV)-specific TCRVbeta(+)/CD4(+)/cytotoxic/memory T cells, we investigate the potential involvement of hCMV in the origin and/or expansion of monoclonal CD4(+) T-LGL. Peripheral blood samples from patients with monoclonal TCR-alphabeta(+)/CD4(+) T-LGL lymphocytosis and other T-chronic lymphoproliferative disorders were evaluated for the specific functional response against hCMV and hEBV whole lysates as well as the "MQLIPDDYSNTHSTRYVTVK" hCMV peptide, which is specifically loaded in HLA-DRB1*0701 molecules. A detailed characterization of those genes that underwent changes in T-LGL cells responding to hCMV was performed by microarray gene expression profile analysis. Patients with TCR-alphabeta(+)/CD4(+) T-LGL displayed a strong and characteristic hCMV-specific functional response, reproduced by the hCMV peptide in a subset of HLA-DRB1*0701(+) patients bearing TCRVbeta13.1(+) clonal T cells. Gene expression profile showed that the hCMV-induced response affects genes involved in inflammatory and immune responses, cell cycle progression, resistance to apoptosis, and genetic instability. This is the first study providing evidence for the involvement of hCMV in the ontogeny of CD4(+) T-LGL, emerging as a model disorder to determine the potential implications of quite a focused CD4(+)/cytotoxic immune response.     

Impact of hepatitis B virus basic core promoter mutations on T cell response to an immunodominant HBx-derived epitope

The hepatitis B X (HBx) protein is a crucial component in HBV infection in vivo and has been implicated in HCC. In this study, we aimed to detect and characterize peripheral HBx-specific T cells in chronically infected patients at the inactive carrier state of the disease. HBx-specific IFN-gamma-secreting T cells were found in 36 of 52 patients (69%), and 78% (28/36) of responding patients had T cells targeting epitopes in the carboxy-terminal part of HBx. IL-10 secretion after the stimulation of T cells with HBx-derived peptides was weak or undetectable. IFN-gamma-secreting T cells recognized a previously unknown immunodominant CD4+ T cell epitope, HBx 126-140 (EIRLKVFVLGGCRHK), in 86% (24 of 28) of patients. This peptide bound several HLA-DR molecules (HLA-DRB1*0101, HLA-DRB1*0401, HLA-DRB1*1301, and HLA-DRB5*0101). Its coding sequence overlaps a domain of the HBV genome encompassing the basic core promoter (BCP) region. Taking into account the selection of viral core promoter mutants during HBV infection, we found that HBV variants with BCP mutations were present in patient sera. We further demonstrated that these viral mutant sequences activated T cells specific for the immunodominant epitope only weakly, if at all. This is the first study linking BCP mutations and HBx-specific T cell responses. CONCLUSION: Wild-type and variant peptides may represent potential tools for monitoring the HBV-specific T cell responses involved in sequence evolution during disease progression. Finally, the degenerate HLA-DR binding of this promiscuous, immunodominant peptide would make it a valuable component of vaccines for protecting large and ethnically diverse patient populations.     

HLA class II-restricted antigen presentation of endogenous bcr-abl fusion protein by chronic myelogenous leukemia-derived dendritic cells to CD4(+) T lymphocytes

Bcr-abl fusion peptide-specific CD4+ T-lymphocyte clones have recently been shown to augment colony formation by chronic myelogenous leukemia (CML) cells in a bcr-abl type-specific and HLA class II-restricted manner without addition of exogenous antigen. These findings suggest that CML cells can naturally process and present endogenous bcr-abl fusion protein to CD4+ T lymphocytes in the context of HLA class II molecules. To verify this possibility, the ability of CML-derived dendritic cells (DCs) to present endogenous bcr-abl fusion protein to bcr-abl fusion peptide-specific CD4+ T-lymphocyte clones was investigated. The bcr-abl b3a2 peptide-specific and HLA-DRB1*0901-restricted CD4+ T-lymphocyte clones produced interferon-gamma in response to stimulation with monocyte-derived DCs from HLA-DRB1*0901+ patients with b3a2 type CML. In contrast, DCs from patients with HLA-DRB1*0901- or b2a2 type CML and those from healthy individuals did not exert stimulatory activity on bcr-abl-specific CD4+ T-lymphocyte clones. The response of CD4+ T-lymphocyte clones to CML-derived mature DCs was higher than that to immature DCs and was inhibited by anti-HLA-DR monoclonal antibody. These data suggest that CML-derived DCs can process and present endogenous bcr-abl fusion protein to CD4+ T lymphocytes.     

Premature telomeric loss in rheumatoid arthritis is genetically determined and involves both myeloid and lymphoid cell lineages

In rheumatoid arthritis, peripheral blood T cells have age-inappropriate telomeric erosion. We examined whether HLA-DRB1*04 alleles, the major susceptibility genes for this disease, confer risk for T cell senescence. In healthy individuals, HLA-DRB1*04 alleles were associated with excessive loss of telomeres in CD4+ T cells. Accelerated telomeric erosion occurred during the first two decades of life and was followed by reduced homeostatic T cell proliferation during adulthood. Premature telomeric loss also affected granulocytes, suggesting that the hematopoietic stem cell is the primary target. Telomeric repair mechanisms were intact in HLA-DRB1*04+ donors. We propose that HLA-DRB1*04 alleles or genes in linkage disequilibrium regulate stem cell replication and contribute to the accumulation of senescent and autoreactive T cells in rheumatoid arthritis.     

Direct killing of Epstein-Barr virus (EBV)-infected B cells by CD4 T cells directed against the EBV lytic protein BHRF1

Due to their low frequency, CD4 T-cell responses to Epstein-Barr virus (EBV) lytic antigens are, so far, poorly characterized. Human peptide major histocompatibility complex (MHC) class II multimers provide a means to detect and characterize such rare T cells. Along a screening of T-cell responses to lytic or latent EBV antigens within peripheral blood leukocyte (PBL)- or synovial-derived CD4 T-cell lines, we identified an human leukocyte antigen-DR*0401 (HLA-DR*0401)-restricted epitope derived from BHRF1 (BamHI fragment H rightward open reading frame 1), a viral protein produced during the early stages of the lytic cycle. We show here that T-cell responses to this particular BHRF1 epitope are shared by most EBV-infected DR*0401(+) individuals, as BHRF1-specific CD4 T cells could be sorted out from all the DRB*0401 T-cell lines analyzed, using magnetic beads coated with recombinant BHRF1/DR*0401 complexes. Sorting with these peptide MHC class II multimers was very efficient, as the yield of recovery of BHRF1-specific T cells was nearly 100%. Functional analysis of a large number of clones responding to BHRF1/DR*0401 demonstrated their cytolytic action against autologous and allogeneic DR*0401(+) EBV-transformed B-lymphoblastoid cell lines (B-LCLs), with 40% to 80% killing efficiency and potent interferon gamma production, thus suggesting that this CD4 T-cell population contributes to the control of EBV replication. B-LCL lysis by these T-cell clones was DR*0401 dependent, EBV dependent, and was not merely due to bystander killing. Taken together, these data provide the first demonstration that a lytic antigen can induce a direct cytolytic response against EBV-infected cells.     

Familial aggregation of polymyalgia rheumatica and giant cell arteritis: genetic and T cell repertoire analysis.

OBJECTIVE: Several reports of familial aggregation of giant cell arteritis (GCA) and polymyalgia rheumatica (PMR) have been described although detailed genetic and immunological studies are scarce. Our aims were to investigate the influence of HLA-DRB1 alleles and to analyze the phenotype and T cell receptor (TCR) usage of circulating T lymphocytes in a familial case of GCA and PMR. METHODS: HLA-DRB1 typing was carried out using polymerase chain reaction amplification with specific primers. The study of the circulating T cell repertoire was performed by staining with specific monoclonal antibodies and flow cytometry analysis. RESULTS: Patient 1 developed GCA at the age of 71, four years prior to the diagnosis of PMR in her older brother. The HLA-DRB1 typing of Patient 1 was DRB1*04 (DRB1*0401)/DRB1*12 and in Patient 2 was DRB1*07/DRB1*12. In our patient population, GCA was associated with an increased frequency of HLA-DRB1*04 compared with PMR patients. Regarding T cell phenotype, the brother with active PMR had a higher expression of surface markers indicating activation in both T cell subsets (CD25 and HLA-DR). The sister with GCA showed a pronounced decrease of CD4+/CD45RA+ T cells with respect to her brother with PMR. Both patients carried a significant depletion of CD28 in both subsets, specially within the CD8+ T cell compartment. The BV gene usage differed from one patient to the other. T cell expansions were identified in both patients but the specificities were different. CONCLUSION: We describe an association of GCA and PMR between two first degree relatives with significant genetic and immunologic differences. Our results suggest that the pathogenic mechanisms leading to the development of GCA and PMR are probably multifactorial, and both genetic and environmental factors may contribute to the development of these diseases.     

Identification and in vitro expansion of CD4+ and CD8+ T cells specific for human neutrophil elastase

Human neutrophil elastase (HNE) and proteinase 3 (PRO3) are myeloid tissue-restricted serine proteases, aberrantly expressed by myeloid leukemia cells. PRO3 and HNE share the PR1 peptide sequence that induces HLA-A*0201-restricted cytotoxic T cells (CTLs) with antileukemia reactivity. We studied the entire HNE protein for its ability to induce CTLs. In an 18-hour culture, HNE-loaded monocytes stimulated significant intracellular interferon gamma (IFN-gamma) production by CD4+ and CD8+ T cells in 12 of 20 and 8 of 20 healthy individuals, respectively. Lymphocytes from 2 HNE responders were pulsed weekly for 4 weeks to generate HNE-specific CTLs. One of 2 HLA-A*0201-negative individuals inhibited the colony formation of HLA-identical chronic myelogenous leukemia progenitor cells (73% inhibition at 50:1 effector-target [E/T] ratio), indicating that peptides other than PR1 can induce leukemia-reactive CTLs. Repetitive stimulations with HNE in 2 of 5 HLA-A*0201+ individuals increased PR1 tetramer-positive CD8+ T-cell frequencies from 0.1% to 0.29% and 0.02% to 0.55%, respectively. These CTLs recognized PR1 peptide or killed HNE-loaded targets. These results indicate that exogenously processed HNE is a source of PR1 peptide as well as other peptide sequences capable of inducing leukemia-specific CD8+ and CD4+ T cells. HNE could, therefore, be used in an HLA-unrestricted manner to induce leukemia-reactive CTLs for adoptive immunotherapy.     

CD4+ T-cell epitopes associated with antibody responses after intravenously and subcutaneously applied human FVIII in humanized hemophilic E17 HLA-DRB1*1501 mice

Today it is generally accepted that B cells require cognate interactions with CD4(+) T cells to develop high-affinity antibodies against proteins. CD4(+) T cells recognize peptides (epitopes) presented by MHC class II molecules that are expressed on antigen-presenting cells. Structural features of both the MHC class II molecule and the peptide determine the specificity of CD4(+) T cells that can bind to the MHC class II-peptide complex. We used a new humanized hemophilic mouse model to identify FVIII peptides presented by HLA-DRB1*1501. This model carries a knockout of all murine MHC class II molecules and expresses a chimeric murine-human MHC class II complex that contains the peptide-binding sites of the human HLA-DRB1*1501. When mice were treated with human FVIII, the proportion of mice that developed antibodies depended on the application route of FVIII and the activation state of the innate immune system. We identified 8 FVIII peptide regions that contained CD4(+) T-cell epitopes presented by HLA-DRB1*1501 to CD4(+) T cells during immune responses against FVIII. CD4(+) T-cell responses after intravenous and subcutaneous application of FVIII involved the same immunodominant FVIII epitopes. Interestingly, most of the 8 peptide regions contained promiscuous epitopes that bound to several different HLA-DR proteins in in vitro binding assays.     

Identification of circulating antigen-specific CD4+ T lymphocytes with a CCR5+, cytotoxic phenotype in an HIV-1 long-term nonprogressor and in CMV infection

Antigen-specific CD4+ effector T cells primarily provide help for B-cell antibody responses and CD8+ cytotoxic T-lymphocyte (CTL) responses. We have found an expanded population of HIV-1 p24-specific, T-cell receptor V beta 17+, CD4+ T lymphocytes, defined by in vitro proliferative and interferon-gamma responses to a 15-mer Gag peptide, in the peripheral blood of an individual with long-term nonprogressive HIV-1 infection. Ex vivo, these cells were CCR5+ and CCR7-, consistent with an effector/memory function. Surprisingly, these cells highly expressed several proteins characteristic of cytotoxic lymphocytes, including TIA-1 (T-cell intracellular antigen 1; GMP-17/NKG7), granzymes A and B, CD161 (NKRP-1), and CD244 (C1.7/2B4). Following in vitro peptide stimulation, these cells produced interleukin 2 (IL-2) and intracellular CD40L, suggesting possible helper function, in addition to induction of perforin and cytotoxicity. A subset of cytomegalovirus (CMV)-specific CD4+ T cells in healthy adults similarly expressed these CTL markers and CCR5, ex vivo. Furthermore, this distinct subset of CD4+ T cells was significantly elevated in healthy CMV-seropositive adults, compared with CMV-seronegative individuals. These results suggest that CCR5+ CD4+ CTL may be a major effector mechanism of the immune response to viral infections in humans. Moreover, expression of CCR5 may render them particularly susceptible to cytopathic effects during progressive HIV-1 infection.