Cabral Joana, Hanley Shirley A, Gerlach Jared Q, O'Leary Neil, Cunningham Stephen, Ritter Thomas, Ceredig Rhodri, Joshi Lokesh, Griffin Matthew D
Regenerative Medicine Institute (REMEDI) at CÚRAM Centre for Research in Medical Devices, School of Medicine, College of Medicine, Nursing and Health Sciences, National University of Ireland, Galway, Ireland.
Flow Cytometry Core Facility, National Centre for Biomedical Engineering Sciences (NCBES), National University of Ireland, Galway, Ireland.
Front Immunol. 2017 Aug 21;8:987. doi: 10.3389/fimmu.2017.00987. eCollection 2017.
Regulatory T-cells (Treg) are essential for maintaining immune homeostasis and tolerance. Surface glycosylation is ubiquitous on mammalian cells and regulates diverse biological processes. While it is currently well accepted that surface glycan expression influences multiple aspects of T-cell function, little is known about the relevance of glycosylation to Treg biology. This study aimed to profile the surface glycosylation characteristics of Treg in various lymphoid compartments of mouse and in human peripheral blood with comparison to non-regulatory, conventional CD4 T-cells (Tconv). It also sought to determine the relationship between the surface glycosylation characteristics and suppressive potency of Treg. Lectin-based flow cytometric profiling demonstrated that Treg surface glycosylation differs significantly from that of Tconv in the resting state and is further modified by activation stimuli. In mouse, the surface glycosylation profiles of FoxP3 Treg from spleen and lymph nodes were closely comparable but greater variability was observed for Treg in thymus, bone marrow, and blood. Surface levels of tri/tetra-antennary glycans correlated with expression of proteins known to be involved in Treg suppressive functions, including GITR, PD-1, PD-L1, CD73, CTLA-4, and ICOS. In coculture experiments involving purified Treg subpopulations and CD4 or CD8 Tconv, higher surface tri/tetra-antennary glycans was associated with greater Treg suppressive potency. Enzymatic manipulation of mouse Treg surface glycosylation resulting in a temporary reduction of surface -glycans significantly reduced Treg capacity to suppress Tconv activation through contact-dependent mechanisms. Overall, these findings demonstrate that Treg have distinctive surface glycan characteristics that show variability across anatomical locations and are modulated by activation events. They also provide evidence of an important role for surface glycosylation in determining Treg phenotype and suppressive potency. These insights may prove relevant to the analysis of Treg in disease settings and to the further development of Treg-based immunotherapies.
调节性T细胞（Treg）对于维持免疫稳态和耐受性至关重要。表面糖基化在哺乳动物细胞中普遍存在，并调节多种生物学过程。虽然目前人们普遍认为表面聚糖表达会影响T细胞功能的多个方面，但关于糖基化与Treg生物学的相关性却知之甚少。本研究旨在剖析小鼠不同淋巴器官以及人类外周血中Treg的表面糖基化特征，并与非调节性的传统CD4 T细胞（Tconv）进行比较。该研究还试图确定Treg表面糖基化特征与其抑制能力之间的关系。基于凝集素的流式细胞术分析表明，在静息状态下，Treg的表面糖基化与Tconv有显著差异，并且会因激活刺激而进一步改变。在小鼠中，脾脏和淋巴结中的FoxP3 Treg表面糖基化谱密切可比，但胸腺、骨髓和血液中的Treg糖基化谱变异性更大。三/四天线聚糖的表面水平与已知参与Treg抑制功能的蛋白质表达相关，包括糖皮质激素诱导的肿瘤坏死因子受体（GITR）、程序性死亡受体1（PD-1）、程序性死亡配体1（PD-L1）、5'-核苷酸酶（CD73）、细胞毒性T淋巴细胞相关蛋白4（CTLA-4）和诱导共刺激分子（ICOS）。在涉及纯化的Treg亚群和CD4或CD8 Tconv的共培养实验中，较高的表面三/四天线聚糖与更强的Treg抑制能力相关。对小鼠Treg表面糖基化进行酶处理导致表面聚糖暂时减少，这显著降低了Treg通过接触依赖性机制抑制Tconv激活的能力。总体而言，这些发现表明Treg具有独特的表面聚糖特征，这些特征在不同解剖位置存在变异性，并受激活事件调节。它们还提供了证据，证明表面糖基化在决定Treg表型和抑制能力方面起着重要作用。这些见解可能与疾病背景下Treg的分析以及基于Treg的免疫疗法的进一步发展相关。
