Gene expression profiling of effector and regulatory T-cells from peripheral blood of rheumatoid arthritis (RA) patients and healthy volunteers.

Objective: Conflicting evidence exists regarding the suppressive capacity of Tregs from the peripheral blood (PB) of patients with rheumatoid arthritis (RA). Our aim was to determine whether Tregs are intrinsically defective in RA using a wide range of read-out assays. Methods: CD3+CD4+CD25+CD127low Tregs from CD45RO+ and CD45RA+ compartments of PB from patients with RA and healthy controls (HC) were analysed for phenotype, cytokine expression profile (ex vivo and after in vitro stimulation), suppression of effector T-cell proliferation and cytokine production, suppression of monocyte-derived cytokine/chemokine production, and gene expression profiling. Results: No differences were observed between patients with RA and HC regarding Treg frequency, ex vivo phenotype (CD4, CD25, CD127, CD39, CD161) or pro-inflammatory cytokine profile (IL-17, IFN-gamma, TNF-alpha). FOXP3 expression was increased in Tregs from RA blood. The ability of Tregs to suppress T-cell proliferation or cytokine (IFN-gamma, TNF-alpha) production upon co-culture with autologous CD45RO+ effector T-cells and monocytes was not significantly different between patients with RA and HC. CD45RO+ Tregs from RA blood showed a slightly impaired ability to suppress production of some cytokines/chemokines by autologous LPS-activated monocytes (IL-1-beta, IL-1Ra, IL-7, CCL3, CCL4), but this was not true for all patients and other cytokines/chemokines (TNF-alpha, IL-6, IL-8, IL-12, IL-15, CCL5) were suppressed in the majority of patients similarly to HC. Finally, gene expression profiling of CD45RA+ or CD45RO+ Tregs from PB revealed no statistically significant differences between patients with RA and HC. Conclusions: Our findings suggest that Tregs isolated from PB of patients with RA are not intrinsically defective. Sorted regulatory and effector T-cells were isolated from peripheral blood (6 healthy controls and 6 patients with rheumatoid arthritis). Total RNA was isolated and profiled using Affy HG-U133_Plus_2 chips.

研究目的：目前针对类风湿关节炎（rheumatoid arthritis, RA）患者外周血（peripheral blood, PB）来源调节性T细胞（Tregs）的抑制功能，已有研究结论存在相互矛盾之处。本研究旨在通过多种检测分析手段，明确RA患者体内的Tregs是否存在固有功能缺陷。研究方法：本研究分别从RA患者与健康对照（healthy controls, HC）的外周血中，分选CD45RO+与CD45RA+亚群的CD3+CD4+CD25+CD127low Tregs，对其表型、细胞因子表达谱（离体状态及体外刺激后）、效应T细胞增殖与细胞因子产生抑制能力、单核细胞来源细胞因子/趋化因子产生抑制能力，以及基因表达谱进行分析。研究结果：RA患者与健康对照的Tregs频率、离体表型（CD4、CD25、CD127、CD39、CD161）及促炎细胞因子谱（白细胞介素17、干扰素γ、肿瘤坏死因子α）均无显著差异。RA患者外周血Tregs的FOXP3表达水平升高。在与自体CD45RO+效应T细胞及单核细胞共培养时，Tregs对T细胞增殖或细胞因子（干扰素γ、肿瘤坏死因子α）产生的抑制能力，在RA患者与健康对照间无显著差异。RA患者外周血的CD45RO+ Tregs在抑制自体脂多糖（lipopolysaccharide, LPS）活化单核细胞产生部分细胞因子/趋化因子（白细胞介素1β、白细胞介素1受体拮抗剂、白细胞介素7、趋化因子配体3、趋化因子配体4）时功能略有受损，但该现象并非存在于所有患者中；且多数患者的Tregs对其余细胞因子/趋化因子（肿瘤坏死因子α、白细胞介素6、白细胞介素8、白细胞介素12、白细胞介素15、趋化因子配体5）的抑制能力与健康对照无显著差异。最后，对外周血CD45RA+或CD45RO+ Tregs的基因表达谱分析显示，RA患者与健康对照间无统计学意义的显著差异。研究结论：本研究结果表明，从RA患者外周血中分离得到的Tregs并不存在固有功能缺陷。本研究从外周血中分离得到调节性T细胞与效应T细胞，共纳入6名健康对照与6名RA患者。提取总RNA后，使用Affy HG-U133_Plus_2芯片进行基因表达谱分析。