Deciphering the metabolic signature of CD4+ T helper cells that express dual master transcription factors.

破译表达双主转录因子的 CD4 T 辅助细胞的代谢特征。

• 批准号: 1953078
• 金额: --
• 依托单位: Newcastle University
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2017
• 资助国家: 英国
• 起止时间: 2017 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-1953078
• 关键词: Deciphering; metabolic; signature; CD4+; helper

Importance:Immunotherapy has shown success in recent years in the field of both cancer and autoimmunity. In particular, clinical trials using T cells have revolutionised treatment of tumour (Th1 cells) and autoimmune juvenile diabetes (Treg cells). The major pitfall of these trials remains in the area of in-vivo T cell persistence. Recent reports have attributed the lack of persistence to the bioenergetics of T cells utilised in these trials. Hence there is an immediate need for understanding the bioenergetics profile of inflammatory versus regulatory T cells for improving clinical care.Background: CD4+ T cells are important mediators of adaptive immunity and are classified into various subtypes based on their transcription factor profile. CD4+ T helper cells that express Tbet transcription factor are called Th1 cells, Th2 (GATA3+), Th17 (RORgt+) or Tregs (FoxP3+) cells. Each subset has a unique function as determined by the respective transcription factor with Tbet driving the production of IFNg cytokine and hence promoting inflammatory responses and FoxP3 instills suppressive function in Tregulatory cells. Activation of the T cell receptor (TCR) along with co-stimulatory signalling is also critical for T helper cell differentiation, which results in metabolic remodelling and acquiring aerobic glycolysis. Differentiation of CD4+ T helper subsets with diverse effector functions is accompanied by additional changes in metabolism to meet their bioenergetic demands. Hence, Th1 cells have a unique metabolic signature, mitochondrial energetics and glycolytic profile (strongly glycolytic) as compared to Tregs that prefer oxidation of lipids. We and others have shown that Tbet+Th1 cells can also express FoxP3 and acquire a regulatory phenotype but maintain Tbet expression. Providing co-inhibitory signals through PD-1/PDL-1 (programmed death 1pathway) in Th1 cells stabilizes FoxP3 expression and Tbet expression. The metabolic signature of these cells that express dual opposing transcription factors is unknown.In this proposal we aim to elucidate the metabolic signature of Tbet+FoxP3+ cells and identify how metabolism co-relates to their function in mouse models of inflammatory diseases.Methodology: A unique resource that is available to us is a fate mapping reporter murine strain that drives both Tbet and FoxP3 and hence allow isolating Tbet+FoxP3+ T cells based on their transcription factor expression. We aim to isolate these cells from WT mice to analyze their metabolic profile by using the Seahorse XF Analyzer to evaluate real time cellular respiration of the cells and response to metabolic modulators. This analysis takes simultaneous repeated measurements of oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) that are representative of mitochondrial oxidative phosphorylation (OXPHOS) and glycolysis respectively (measured by changes in oxygen and proton (pH) concentration respectively). In addition, CD4+ T helper cells that are Tbet+, will be expanded in vitro in the presence/absence of PDL-1 to become Tbet+FoxP3+ cells. As before we will compare the metabolic profile and in vitro effector phenotype of these expanded cells to day 0 WT Tbet+FoxP3+ sorted cells. The in-vivo function of Tbet+FoxP3+ cells will be tested to determine if function co-relates to specific metabolic signature in this subset.Training opportunities: The students involved in this project will be trained in cross-disciplinary fields of biochemistry and immunology which include murine models of disease and fate mapping reporters, in vitro cellular immunological and biochemical assays and use of multi-parameter flow cytometry. The strong inter-disciplinary aspect of this PhD will significantly enhance the student's future career prospects. As well as taking part in the weekly specialist subject research meetings, journal clubs and institute research programs, the student will engage with the Graduate School's Skills Dev Prog.

重要性：近年来，免疫疗法在癌症和自身免疫领域都取得了成功。特别是，使用T细胞的临床试验彻底改变了肿瘤(Th1细胞)和自身免疫性幼年糖尿病(Treg细胞)的治疗方法。这些试验的主要缺陷仍然是体内T细胞的持久性。最近的报告将缺乏持久性归因于这些试验中使用的T细胞的生物能量学。因此，迫切需要了解炎症性T细胞与调节性T细胞的生物能量学特征，以改善临床护理。背景：CD4+T细胞是适应性免疫的重要介质，根据其转录因子特征被分为不同的亚型。表达Tbet转录因子的CD4+T辅助细胞称为Th1细胞，Th2(GATA3+)，Th17(RORgt+)或Tregs(FoxP3+)细胞。每个亚群都有独特的功能，由各自的转录因子决定，Tbet驱动IFNG细胞因子的产生，从而促进炎症反应和FoxP3在T调节细胞中的抑制功能。T细胞受体(TCR)的激活和共刺激信号对T辅助细胞分化也是至关重要的，T辅助细胞分化导致代谢重塑和获得有氧糖酵解。具有不同效应器功能的CD4+T辅助细胞亚群的分化伴随着代谢的额外变化，以满足其生物能量需求。因此，与喜欢脂质氧化的Treg细胞相比，Th1细胞具有独特的代谢特征、线粒体能量和糖酵解(强烈的糖酵解)。我们和其他人已经证明，Tbet+Th1细胞也可以表达FoxP3，并获得调节表型，但保持Tbet的表达。在Th1细胞中通过PD-1/PDL-1(程序性死亡1通路)提供共抑制信号可以稳定FoxP3和Tbet的表达。这些细胞表达双重相反转录因子的代谢特征尚不清楚。在这个建议中，我们旨在阐明Tbet+FoxP3+细胞的代谢特征，并在炎症性疾病的小鼠模型中确定代谢与其功能的相关性。方法：我们可以获得的唯一资源是命运映射报告鼠株，它同时驱动Tbet和FoxP3，从而允许基于其转录因子的表达来分离Tbet+FoxP3+T细胞。我们的目标是从WT小鼠中分离这些细胞，通过使用SeaHorse XF Analyzer来评估细胞的实时细胞呼吸和对代谢调节剂的反应，以分析它们的代谢特征。该分析同时重复测量分别代表线粒体氧化磷酸化(OXPHOS)和糖酵解(分别通过氧气和质子(PH)浓度变化衡量)的耗氧率(OCR)和胞外酸化率(ECAR)。此外，在PDL-1存在或不存在的情况下，Tbet+的CD4+T辅助细胞将在体外扩增为Tbet+FoxP3+细胞。如前所述，我们将比较这些扩增细胞与第0天WT Tbet+FoxP3+分选细胞的代谢特征和体外效应表型。将测试Tbet+FoxP3+细胞的体内功能，以确定功能是否与这一亚群中的特定代谢特征相关。培训机会：参与该项目的学生将接受生物化学和免疫学交叉学科领域的培训，其中包括小鼠疾病模型和命运图谱报告、体外细胞免疫和生化分析以及多参数流式细胞术的使用。这一博士学位具有很强的跨学科特点，将显著提升学生未来的职业前景。除了参加每周的专家主题研究会议、期刊俱乐部和研究所的研究项目外，学生还将参与研究生院的技能开发项目。

项目成果

PD-1 Inhibitory Receptor Downregulates Asparaginyl Endopeptidase and Maintains Foxp3 Transcription Factor Stability in Induced Regulatory T Cells.

• DOI: 10.1016/j.immuni.2018.05.006
• 发表时间: 2018-08-21
• 期刊: Immunity
• 影响因子: 32.4
• 作者: Stathopoulou C;Gangaplara A;Mallett G;Flomerfelt FA;Liniany LP;Knight D;Samsel LA;Berlinguer-Palmini R;Yim JJ;Felizardo TC;Eckhaus MA;Edgington-Mitchell L;Martinez-Fabregas J;Zhu J;Fowler DH;van Kasteren SI;Laurence A;Bogyo M;Watts C;Shevach EM;Amarnath S
• 通讯作者: Amarnath S