急性冠状动脉综合征患者外周血CD4+T细胞及CD28null/CD28+亚型活化后Kv1.3钾通道的表达和意义

目的 研究急性冠状动脉综合征(ACS)患者外周血中CD4+T细胞及CD28null/CD28+亚型活化前后Kv1.3钾通道数目的 变化以及Kv1.3钾通道阻滞剂对CD4+T细胞活化表达的影响,探讨Kv1.3钾通道在不稳定斑块中的意义.方法 用免疫磁珠法分离出27例ACS患者外周血中的CD4+T细胞,其中12例进一步分出亚型CD4+CD28null和CD4+CD28+T细胞,采用全细胞膜片钳技术记录细胞活化前及经CD3抗体活化72 h后的Kv1.3钾电流.CD4+T细胞活化时分别加入终浓度为0.1、1、10 nmol/L特异性Kv1.3钾通道阻滞剂rMargatoxin(rMgTX),共同培养72 h后用反转录-PER法检测干扰素-γ、肿瘤坏死因子(TNF)-α及颗粒酶B mRNA的表达.结果 活化后CD4+、CD4+CD28null、CD4+CD28+T细胞的Kv1.3钾通道的峰电流均明显增加,细胞平均通道数分别增加约90%、60%、80%[活化前后每个细胞的通道数分别为(402±88)个比(752±275)个、(553±328)个比(874±400)个、(392±133)个比(716±251)个,均P<0.05].活化前CD4+CD28nullT细胞Kv1.3钾通道的平均数目比CD4+CD28+T细胞多约40%(P<0.05),活化后两者差异无统计学意义(P=0.102).不同浓度的rMgTX均下调CD4+T细胞活化后干扰素-γ、TNF-α、颗粒酶B mRNA的表达,各浓度组间干扰素-γ、TNF-α、颗粒酶B mRNA的表达差异均有统计学意义(均P<0.01),浓度越高,各mRNA表达越低.结论 ACS患者外周血CD4+T细胞及CD28null/CD28+亚型活化后Kv1.3钾通道表达增加,特异性Kv1.3通道阻滞剂rMgTX呈浓度依赖性地抑制CD4+T细胞活化时干扰素-γ、TNF-α及颗粒酶B mRNA的表达,提示CD4+T细胞特别是CD4+CD28nullT细胞的Kv1.3钾通道可作为预防动脉粥样斑块不稳定的潜在治疗靶点.     

高血压患者外周血CD4+T细胞Kv1.3钾通道的变化

目的:探讨高血压患者外周血CD4+T淋巴细胞上的电压门控性钾通道(Kv1.3)电流的变化。方法:收集24例高血压患者和24例健康志愿者外周血,免疫磁珠法分选出CD4+T淋巴细胞,分别给予0、2nmol/L血管紧张素Ⅱ(AngⅡ)培养48h,采用全细胞膜片钳技术记录淋巴细胞Kv1.3钾通道电流,并用RT-PCR检测Kv1.3钾通道和血管紧张素受体(AT1)蛋白mRNA的表达,ELISA试剂盒检测培养上清IFN-γ浓度。结果:①高血压患者CD4+T细胞Kv1.3钾通道电流峰值(307±117)pA,明显高于正常人(233±95)pA(P0.05)。②给予AngⅡ干预后,正常人CD4+T细胞Kv1.3钾通道电流峰值明显增高(325±99)pA(P0.05);而高血压患者CD4+T细胞Kv1.3钾通道电流峰值增高不明显。正常加药组CD4+T细胞Kv1.3钾通道mRNA表达也增高。③淋巴细胞存在AT1受体,且高血压患者CD4+T细胞AT1受体mRNA表达明显高于正常人;AngⅡ干预后高血压患者CD4+T细胞培养上清IFN-γ浓度较正常人增加更明显。结论:高血压患者淋巴细胞Kv1.3钾通道表达增多,AngⅡ能增加Kv1.3钾通道表达,AngⅡ促进CD4+T细胞活化和IFN-γ分泌的效应可能由AT1受体介导。本研究提示炎症参与高血压的发病及其靶器官损害,而CD4+T细胞Kv1.3钾通道在高血压炎症的发生发展中发挥重要作用。     

急性冠状动脉综合征患者外周血T淋巴细胞钾通道表达的差异

目的:探讨急性冠状动脉综合征(ACS)患者外周血CD_4~+CD28~(null)T细胞和CD_4~+CD28~+T细胞上电压依赖性钾离子通道(Kv1.3)和钙依赖激活的钾离子通道(IKCa1)的表达.方法:入选对象共75例,分为3组:其中ACS组27例,稳定型心绞痛组20例,对照组28例.分离外周血单个核细胞(PBMCs),用ELISA法测定PBMCs中NF-κB的活性,流式细胞仪计数CD_4~+CD28~(null)T细胞占CD4+T细胞的比例.免疫荧光染色标记结合膜片钳技术记录ACS组CD_4~+CD28~(null)T细胞和CD_4~+CD28~+T细胞上Kv1.3和IKCa1的电流,计算单细胞上通道的数目.结果:ACS组NF-κB的活性和CD_4~+CD28~(null)T细胞的比例显著高于稳定型心绞痛组和对照组(P<0.05).NF-κB的活性和CD4+CD28null T细胞的比例具有正相关关系(r=0.369,P<0.05).CD_4~+CD28~(null)T细胞上Kv1.3的数目和电流密度显著高于CD_4~+CD28~+T细胞(P<0.05).CD_4~+CD28~(null)T细胞、CD_4~+CD28~+T细胞上IKCa1的数目差异无统计学意义(P>0.05).结论:Kv1.3通道对CD_4~+CD28~(null)T细胞的功能具有重要作用,Kv1.3通道有可能成为AS治疗的新靶点.     

急性冠状动脉综合征患者外周血CD4~+及CD4~+ CD28~(null) T细胞活化后IKCa1通道的表达及意义

目的:研究急性冠状动脉综合征(ACS)患者外周血中CD4+T细胞及CD4+CD28null亚型活化前后IKCa1钾通道数目的变化以及IKCa1钾通道阻滞剂对活化CD4+T细胞效应分子表达的影响,探讨IKCa1钾通道对不稳定斑块的意义.方法:用免疫磁珠法分离出24例ACS患者外周血中的CD4+T细胞,其中12例进一步分出亚型CD4+CD28nullT细胞,采用全细胞膜片钳技术记录细胞活化前及活化3 d后的IKCa1钾电流.CD4+T细胞活化3 d后分别加入终浓度为0 2、1、5 μmol/L特异性IKCa1钾通道阻滞剂TRAM-34,再继续活化3 d,用反转录-PCR法检测干扰素-γ及颗粒酶B mRNA的表达.结果:活化后CD4+、CD4+CD28nullT细胞的IKCa1通道数目分别增加了9倍和8倍[活化前后2种细胞的通道数分别为(45±3)∶(439±33), (56±4)∶(497±45),均P<0 01].2种细胞的通道密度也分别增加了约3倍(P<0 01).活化前及活化后2种细胞的通道数目及通道密度均无差别.不同浓度的TRAM-34均下调CD4+T细胞活化后干扰素-γ、颗粒酶B mRNA的表达,各浓度组间干扰素-γ、颗粒酶B mRNA的表达差异均有统计学意义(P<0 01),浓度越高,各mRNA表达越低.结论:ACS患者外周血CD4+及CD4+CD28nullT细胞活化后IKCa1的通道数目及通道密度均明显增加,特异性IKCa1通道阻滞剂TRAM-34呈浓度依赖性地抑制CD4+T细胞活化时干扰素-γ及颗粒酶B mRNA的表达,提示CD4+T细胞的IKCa1钾通道可作为预防动脉粥样斑块不稳定的潜在治疗靶点.     

Upregulated voltage-gated potassium channel Kv1.3 on CD4+CD28null T lymphocytes from patients with acute coronary syndrome

Objective The purpose of our study is to observe the voltage-gated potassium channel Kv1.3 expressed on CD4+ CD28null T cells from the peripheral blood of acute coronary syndrome (ACS) patients by the patch clamp technique. Methods Kvl.3 potassium channels expression from 17 patients with ACS and 11 healthy age-match controls was detected in single cell(CD4+CD28null T cells and CD4+CD28+T cells) by fluorescence microscopy and patch clamp. Results The percentage of CD4+CD28null T cells was higher in the ACS patients [(6.97±2.05)%] than that in the controls [(1.38±0.84)%, P<0.05]. The concentration of hsCRP was directly correlated with the number of the CD4+CD28null T cells in the ACS patients (r=0.52, P<0.05). The conductance (6.89±1.17ns vs 3.36±0.66ns), dens (1.95±0.80 μm2 vs 1.13±0.57 μm2) and numbers (574.5±97.6 n/cell vs. 280.3±55.3 n/cell) of the Kvl.3 channels on the CD4+CD28null T cells were significantly higher than those on the CD4+CD28+T cells (all P<0.0l) in ACS patients, but were similar on CD4+CD28+T between ACS patients and controls. Conclusion The CD4+CD28null T cells and the numbers of Kv1.3 channels on the CD4+CD28null T cells from patients with ACS are significantly upregulated and might contribute to the pathogenesis of ACS.     

Upregulated voltage-gated potassium channel Kvl.3 on CD4＋CD28null T lymphocytes from patients with acute coronary syndrome

Objective The purpose of our study is to observe the voltage-gated potassium channel Kvl.3 expressed on CD4＋CD28~ T cells from the peripheral blood of acute coronary syndrome （ACS） patients by the patch clamp technique. Methods Kvl.3 potassium channels expression from 17 patients with ACS and 11 healthy age-match controls was detected in single cell（CD4＋CD28null T cells and CD4＋CD28＋T cells） by fluorescence mieroscopy and patch clamp. Results The percentage of CD4＋CD28mullT cells was higher in the ACS patients [（6.97±2.05）%] than that in the controls [（1.38±0.84）%, P〈0.05]. The concentration of hsCRP was directly correlated with the number of the CD4~CD28nul~ T cells in the ACS patients （r=0.52, P〈0,05）. The conductance （6.89±1.17ns vs 3.36±0.66ns）, dens （1.95±0.80 μm2 vs 1.13±0.57 μm2） and numbers （574.5±97.6 n/cell vs. 280.3±55.3 n/cell） of the Kv1.3 channels on the CIM＋CD28null T cells were significantly higher than those on the CD4＋CD28＋ T cells （all P〈0.01） in ACS patients, but were similar on CD4＋CD28＋T betweenACS patients and controls. Conclusion The CD4＋CD28nullT cells and the numbers of Kvl.3 channels on the CD4＋CD28nullT cells from patients with ACS are significantly upregulated and might contribute to the pathogenesis of ACS （d Geriatr Cardio12010; 7：40-46）.     

2型糖尿病动脉粥样硬化患者CD4~+CD28~(null)T细胞亚群的表达及意义

目的探讨CD4+CD28nullT细胞亚群在2型糖尿病动脉粥样硬化发生发展中的作用。方法根据彩色多普勒超声检查结果,将92例2型糖尿病患者分为颈动脉粥样硬化组与对照组,采用流式细胞仪检测分析颈动脉粥样硬化组和对照组外周血CD4+CD28nullT细胞亚群的表达情况,并分析其与年龄、C反应蛋白(CRP)、血糖、血脂、体质指数(BMI)、腰围、臀围、舒张压、收缩压的关系。结果颈动脉粥样硬化组CD4+CD28nullT细胞亚群的表达明显高于对照组,分别为(4.40±2.95)%和(3.04±2.20)%,差异有统计学意义(P0.05)。CD4+CD28nullT细胞亚群的表达与年龄、CRP、糖化血红蛋白(Hb A1c)相关(r=0.20~0.25,P0.05)。控制年龄和Hb A1c因素后,CD4+CD28nullT细胞亚群的表达与CRP仍相关(r=0.24,P0.05)。结论 2型糖尿病动脉粥样硬化患者具有更高的CD4+CD28null T细胞表达。CD4+CD28nullT细胞亚群可能通过介导炎症反应影响2型糖尿病患者动脉粥样硬化的发生和发展。     

初发过敏性紫癜患儿外周血CD4+CD28-和CD8+CD28-T细胞水平变化及意义

目的 观察初发过敏性紫癜(HSP)患儿外周血T淋巴细胞亚群(CD4+ CD28-T细胞和CD8+ CD28-T细胞)的表达变化,并探讨其临床意义.方法 采用流式细胞技术(FCM)检测89例初发HSP患儿(HSP组)和30例健康儿童(对照组)外周血T淋巴细胞免疫表型.结果 HSP组CD4+ CD28+、CD8+ CD28+、CD3+ CD4+T淋巴细胞表达水平及CD4+/CD8+比率较对照组显著降低(P均＜0.05);HSP组CD3+和CD3+ CD8+T淋巴细胞表达水平与对照组相比无显著性差异(P＞0.05);HSP组CD4+ CD28-T细胞和CD8+ CD28-T细胞较对照组明显增高(P均＜0.05),但在皮肤型、关节型、腹型、混合型和紫癜性肾炎五种临床类型间无明显差别(P均＞0.05).结论 HSP患儿存在T淋巴细胞亚群的紊乱,CD4+ CD28-T细胞及CD8+ CD22-T细胞的异常增高可能参与了其发病过程,但与HSP临床类型无关.     

原发性胆汁性肝硬化T淋巴细胞亚群分析

目的 探讨原发性胆汁性肝硬化(PBC)患者的T淋巴细胞亚群及共刺激信号表达的特点及临床意义.方法 以未经治疗的98例PBC患者为研究组,性别、年龄匹配的健康人30名作为对照组.以流式细胞仪技术检测外周血淋巴细胞亚群以及T淋巴细胞表面的共刺激信号CD28.结果 PBC与对照组T细胞亚群差异有统计学意义:PBC组CD4+T淋巴细胞升高,CD8+T淋巴细胞下降,CD4+/CD8+比值上升(P<0.05);CD4+CD28-T细胞和CD8+CD28-T细胞明显增加(P<0.05).结论 PBC存在免疫调节功能的异常,其中CD28的表达明显减少;CD8+CD28-的细胞群可能在PBC中有一定的免疫调节作用.     

间充质干细胞对系统性红斑狼疮CD4+Foxp3+T淋巴细胞的调节

目的 探讨同种异体骨髓间充质干细胞(MSC)体内外对系统性红斑狼疮(SLE)患者外周血CD4+Foxp3+T淋巴细胞及人脐带MSC移植对MRL/lpr鼠脾脏和淋巴结CD4+Foxp3+T淋巴细胞水平的影响.方法 血缘相关供者骨髓中分离培养MSC移植治疗5例SLE患者,采用流式细胞术检测移植前后外周血CD4+Foxp3+T淋巴细胞百分率.7例SLE患者外周血单个核细胞(PBMC)分别与SLE患者和正常人骨髓MSC按不同比例体外共培养72 h,检测共培养后PBMC中CD4+Foxp3+T淋巴细胞百分率.MRL/Ipr鼠输注脐带MSC后检测脾脏和淋巴结CD4+Foxp3+T淋巴细胞百分率.结果 SLE患者异基因骨髓MSC移植后1周外周血CD4+Foxp3+T淋巴细胞百分率(4.8±1.6)%和移植后3个月(6.0±2.6)%均较移植前(2.1±1.2)%明显升高(5例,P<0.05).正常骨髓MSC与SLE患者PBMC共培养后CD4+Foxp3+T淋巴细胞百分率明显升高(P<0.05).且存在剂量依赖性,狼疮MSC也可上调SLE患者CD4+Foxp3+T淋巴细胞水平,但作用较正常MSC弱(P<0.05);正常MSC培养上清也可上调SLE患者PBMC中CD4+Foxp3+T淋巴细胞水平,但作用弱于MSC:PBMC=1:1组(P<0.05).MRL/Ipr鼠经1次或3次脐带MSC移植后脾脏CD4+Foxp3+T淋巴细胞百分率均较对照组高(P<0.05),但淋巴结CD+Foxp3+T淋巴细胞百分率均较对照组低(p<0.01),1次和3次移植组间差异无统计学意义.结论 异基因甚至异种MSC移植可上调SLE患者或MRL/Ipr鼠CD4+Foxp3+T淋巴细胞水平,同时体外试验也得出相同结论,且体外上调作用呈一定剂量依赖性,CD4+Foxp3+T淋巴细胞水平上调可能是MSC移植治疗SLE有效的机制之一.     

De novo expression of killer immunoglobulin-like receptors and signaling proteins regulates the cytotoxic function of CD4 T cells in acute coronary syndromes

The inflammatory infiltrate in atherosclerotic plaque is composed of T cells and macrophages. CD4+ T cells with a unique phenotype, CD4+CD28null, are preferentially recruited into culprit lesions. These T cells are distinct from classic CD4+CD28+ T cells in gene expression and function, including their ability to mediate cytolysis. In this study, we have investigated the regulation of CD4+CD28null T-cell cytolytic function. In patients with acute coronary syndromes (ACS), CD4+CD28null T cells express killer immunoglobulin-like receptors (KIRs). KIRs encompass a polymorphic family of receptors that recognize HLA class I molecules and have been implicated in self-tolerance. CD4+CD28null T-cell clones from patients with ACS and age-matched controls were compared for their KIR-expression profile. T-cell clones derived from the patients expressed a broader spectrum of KIRs (P<0.001) with preference for the stimulatory variant, CD158j. Additionally, CD4+ T-cell clones from patients but not those from controls acquired de novo expression of the DAP12 molecule, an adapter chain that transmits CD158j-derived signals. Cumulative expression of CD158j and DAP12 endowed cytolytic competence on CD4+CD28null T cells, allowing them to kill in the absence of T-cell receptor triggering. Our data demonstrate that CD4+CD28null T cells in ACS are characterized by a unique gene expression profile. Consequently, these T cells acquire cytolytic capability that can bypass the need for T-cell receptor triggering and, thus, impose a threat to self-tolerance.     

急性冠状动脉综合征患者淋巴细胞电压依赖性钾通道的变化

目的用全细胞膜片钳技术和Western印迹法,探讨急性冠状动脉综合征（ACS）患者外周血淋巴细胞电压依赖性钾通道（Kv）电流及Kv1．3蛋白表达的变化。方法收集12例ACS患者和10例健康志愿者外周血淋巴细胞,采用膜片钳全细胞电流记录方法,记录淋巴细胞膜Kv的电流密度。用Western印迹法检测外周血淋巴细胞Kv1．3蛋白的表达。结果ACS患者淋巴细胞Kv的电流密度为（2694-94）pA／pF,明显高于正常人[（1914-64）pA／pF,P〈0．01]。ACS组淋巴细胞膜电容为（2．3±40．7）pF,对照组为（2．2±0．5）pF,两组间差异无统计学意义。Western印迹结果显示ACS患者淋巴细胞Kv1．3通道表达高于正常人（P〈0．01）。结论ACS患者淋巴细胞Kv表达明显增多,说明Kv在调节ACS患者淋巴细胞的激活中起着关键性作用。     

强直性脊柱炎患者外周血CD8+CD28-T细胞的表达及意义

目的 探讨外周血CD8+CD28- T细胞在强直性脊柱炎(AS)发病中的作用.方法 应用流式细胞术检测50例AS患者和21名正常人外周血CD3+CD8+ T细胞表面CD28分子的表达,同时用免疫散射比浊法检测相应血清中的C反应蛋白(CRP).结果 AS患者外周血中的CD8+CD28- T细胞的表达明显高于正常人[(18±6)%对(14±5)%,P=0.020],患者外周血CD3+、CD8+CD28+ T细胞的表达明显低于正常人[(65±9)%对(69±8)%,P=0.039];[(15±5)%对(18±4)%,P=0.038],CD8+ T细胞的表达与正常人相比差异无统计学意义(P＞0.05).结论 AS患者外周血中的CD8+CD28- T细胞的表达增加,可能参与了AS的疾病过程.     

Unusual CD4+CD28null T lymphocytes and recurrence of acute coronary events.

OBJECTIVES: We hypothesized that the expansion of unusual T lymphocytes, CD4+CD28null T cells, might represent a key pathogenetic mechanism of recurrent instability. BACKGROUND: Clinical presentation of acute coronary syndromes (ACS) is variable. Some patients have recurrent episodes of instability, despite optimal treatment, whereas others have a single acute event in their life. The CD4+CD28null T cells, with a functional profile that favors vascular injury, have recently been found both in peripheral blood and in unstable coronary plaques of patients with ACS. METHODS: Peripheral blood T cells from 120 consecutive unstable angina (UA) patients were analyzed for the distribution of T-cell subsets by flow cytometry. Patients were subgrouped according to the occurrence of prior (during the 24 months before the study enrollment) and subsequent (during the 24 months of follow-up) acute coronary events. For 51 patients, the index event was the first ever (G1); 30 patients had prior events (G2); and 39 patients had further events at follow-up (death, myocardial infarction, or UA) or both before and after the index event (G3). RESULTS: The CD4+CD28null T-cell frequency was higher in G3 than in G2 and G1 (median 9.5% [range 2.4% to 48.0%] vs. 5.1% [range 0.4% to 27.8%] and 2.3% [range 0.2% to 22.8%], respectively; p < 0.001). The expansion of these unusual T lymphocytes was higher in patients with elevated C-reactive protein levels, and it was reduced by statin therapy. On multivariate logistic regression analysis, CD4+CD28null T-cell frequency was an independent predictor of future acute coronary events (odds ratio 3.01, 95% confidence interval 1.1 to 8.25; p = 0.023). CONCLUSIONS: A perturbation of T-cell repertoire is strongly associated with the recurrence of acute coronary events, conceivably playing a key pathogenetic role.     

合并糖尿病的急性冠脉综合征患者共刺激分子与炎症指标的研究

目的探讨CD4^＋CD28^nullT淋巴细胞及炎症因子C反应蛋白（CRP）在合并糖尿病的冠心病患者发病机制中的作用。方法51例经冠状动脉造影确认为急性冠脉综合征（ACS）的患者,根据1999年WHO标准分为2型糖尿病合并ACS组21例,单纯ACS组30例,对所有研究对象均通过流式细胞术测量外周血中CD4^＋CD28^nullT淋巴细胞的数量,对比分析两组患者的临床资料和冠状动脉造影结果。结果与单纯ACS患者相比较,合并糖尿病的ACS患者外周血CD4^＋CD28^nullT淋巴细胞的数量显著增多[（4．66±4．24）％VS（8．89±6．45）％,JP〈0．05],CRP浓度无明显统计学差异[（1．97±1．05）VS（2．22±1．14）mg／L,P〉0．05],冠状动脉造影显示完全闭塞、弥漫病变比例较高（7w11,P〈0．05;6YS12,P〈0．01）。结论糖尿病合并ACS患者冠状动脉病变累及范围广且程度重,共刺激分子可能与合并糖尿病的ACS患者的免疫调节异常有关。     

非酒精性脂肪性肝病患者外周血恒定自然杀伤T细胞和CD4+/CD8+T细胞活化差异研究*

目的 分析不同非酒精性脂肪性肝病(NAFLD)患者外周血恒定自然杀伤T细胞(iNKT)、CD4⁺和CD8⁺ T细胞活化标记物(CD69、CD25、HLA-DR和NKG2D)的表达差异。方法 2020年1月～2022年7月我院诊治的NAFLD患者64例和同期健康体检者50例,对NAFLD患者行肝穿刺活检,使用流式细胞仪检测外周血iNKT、CD4⁺和CD8⁺T细胞CD69、CD25、HLA-DR和NKG2D表达。结果 在64例NAFLD患者中,经组织病理学检查,诊断NAFL 37例和NASH 27例;健康对照者、NAFL和NASH患者健康对照者、NAFL和NASH患者外周血CD69⁺iNKT细胞百分比分别为(10.1±1.7)%、(6.1±1.3)%和(26.7±3.6)%(P<0.05),CD25⁺iNKT细胞百分比分别为(83.0±5.9)%、(94.1±8.0)%和(90.8±7.5)%(P<0.05),HLA-DR⁺iNKT细胞百分比分别为(15.3±1.7)%、(15.8±2.0)%和(22.3±2.0)%(P>0.05),NKG2D⁺iNKT细胞百分比分别为(44.5±3.5)%、(59.7±4.0)%和(71.3±6.0)%(P<0.05);外周血CD69⁺CD4⁺ T细胞百分比分别为(0.7±0.2)%、(0.4±0.1)%和(0.5±0.1)%(P>0.05),CD25⁺CD4⁺ T细胞百分比分别为(1.4±0.6)%、(3.0±1.3)%和(1.5±0.7)%(P>0.05),HLA-DR⁺CD4⁺ T细胞百分比分别为(2.7±0.7)%、(4.1±1.0)%和(3.9±1.0)%(P<0.05),NKG2D⁺CD4⁺ T细胞百分比分别为(1.6±0.5)%、(0.6±0.2)%和(0.9±0.2)%(P<0.05);外周血CD69⁺CD8⁺ T细胞百分比分别为(2.0±0.4)%、(1.6±0.3)%和(2.1±0.6)%(P>0.05),CD25⁺CD8⁺ T细胞百分比分别为(1.3±0.3)%、(1.1±0.2)%和(1.0±0.2)%(P>0.05),HLA-DR⁺CD8⁺ T细胞百分比分别为(5.0±0.7)%、(6.5±1.0)%和(9.6±1.4)%(P<0.05),NKG2D⁺CD8⁺ T细胞百分比分别为(0.6±0.1)%、(0.5±0.1)%和(0.9±0.2)%(P<0.05)。结论 本研究发现NAFL与NASH患者可能存在外周血iNKT细胞、CD4⁺和CD8⁺ T细胞活化的免疫表型差异,显示NASH患者CD69⁺iNK T细胞百分比增高,可能对诊断有帮助,值得进一步研究。     

慢性乙型肝炎患者浆样树突状细胞体外诱导CD4+CD25+调节性T细胞的研究

目的 观察慢性乙型肝炎患者,乙型肝炎恢复者和健康人浆样树突状细胞(pDCs)体外诱导CD4+CD25+调节性T细胞(CD4+CD25+Treg)能力的差异,为阐明HBV感染慢性化的机制奠定基础.方法 采用免疫磁珠分离法体外分离46例慢性乙型肝炎患者,10例乙型肝炎恢复者和25名健康人外周血单个核细胞中pDCs,并将其分别与健康人CD4+CD45RA+初始T细胞共培养.采用HBcAg或破伤风毒素对去除CD25+细胞的外周血单个核细胞进行增殖刺激后,使用流式细胞仪及RT-PCR对pDCs-T共培养细胞中CD4+CD25+Treg的数量、表型及FOXP3基因表达情况进行测定;采用酶联免疫吸附法对共培养细胞上清液中的白细胞介素-10和转化生长因子β1进行了进一步检测.两组数据比较采用Mann Whitney U-test.结果 当细胞增殖刺激物为HBcAg时,细胞增殖幅度慢性乙型肝炎患者组为(7999.36±374.74)cpm,乙型肝炎恢复者组为(11 282.56±1174.46)cpm和健康人组为(12 304.58±1462.81)cpm,慢性乙型肝炎患者组细胞增殖幅度明显小于乙型肝炎恢复者组和健康人组,U=0～22.0,P值均<0.05·乙型肝炎恢复者组和健康人组间增殖幅度差异无统计学意义.当增殖刺激物为破伤风毒素时,细胞增殖幅度与阳性对照组之间,差异无统计学意义.CD4+CD25+Treg比例慢性乙型肝炎患者组为5.99%±1.85%,乙型肝炎恢复者组为3.04%±0.79%,健康人组为3.01%±1.53%,慢性乙型肝炎患者组中韵CD4+CD25+Treg比例明显高于乙型肝炎恢复者组和健康人组,U=6.0～71.5,P值均<0.05.3组人群pDCs-T共培养细胞的CD4+CD25+T细胞均检测到Fox p3 RNA,而在CD4+CD25 T细胞中,均未检测到Fox p3RNA.3组人群pDCs-T共培养细胞实验组上清液的白细胞介素-10和转化生长因子β1含量均明显高于阳性对照组.结论 pDCs以诱导CD4+CD25+Treg形式参与了乙型肝炎的慢性化.