Functional and developmental T cell subsets

功能性和发育性 T 细胞亚群

• 批准号: 8552889
• 负责人: Jonathan Ashwell
• 金额: $ 40.03万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8552889
• 关键词: 1-Phosphatidylinositol 3-Kinase; Accounting; Acute; Animals; Antigen Receptors; Antigen-Presenting Cells; Antigens; Apoptosis; B-Lymphocytes; CD3 Antigens; CD44 gene; CD8B1 gene; Cell Cycle; Cell Cycle Progression; Cell Death; Cell Maintenance; Cell Nucleus; Cell Survival; Cell surface; Cells; Characteristics; Clonal Expansion; Complementary DNA; DNA; Data; Defect; Dendritic Cells; Development; Drug or chemical Tissue Distribution; Exposure to; G1 Phase; Generations; Genes; Haploidy; Homing; Human; ITGAX gene; Immune; Immune response; Immune system; Immunity; Individual; Infection; Influenza; Interferon Type II; Interleukin-12; Interphase Cell; Kinetics; Life; Ligation; Lymphocyte; Lymphocyte Homing Receptors; Lymphocytic choriomeningitis virus; MHC Class II Genes; Mediating; Memory; Methods; Microarray Analysis; Mus; Natural Immunity; Organism; Peptide/MHC Complex; Phenotype; Phosphorylation; Play; Population; Process; Proteins; RNA; Readiness; Role; SELL gene; Signal Transduction; Spleen; Surface; T memory cell; T-Cell Receptor; T-Lymphocyte; T-Lymphocyte Subsets; Techniques; Testing; Thymus Gland; Tissues; Translations; Tumor Necrosis Factor Receptor; Vaccination; Vesicular stomatitis Indiana virus; Viral; Virus; Work; alpha-beta T-Cell Receptor; arm; base; cDNA Arrays; cytokine; cytotoxicity; density; in vivo; lymph nodes; mTOR protein; member; neoplastic; pathogen; peripheral blood; prevent; receptor; research study; response; transcription factor

The immune system consists of two major arms: innate and adaptive. Innate immune cells, in particular dendritic cells (DC), are critical in early responses to control pathogens, and play an essential role in activating the adaptive response. The later is mediated by lymphocytes, and is important in dealing with acute threats as was as secondary re-exposures (e.g. a second infection with the same organism). Upon primary exposure to antigen, nave T cells bearing complementary antigen receptors (TCRs) undergo rapid activation and clonal expansion, leading to the generation of effector T cells. The vast majority (90-95%) of antigen-specific effector T cells that participate in the primary immune response undergo apoptosis (cell death) after antigen clearance. A small subset, however, survives and gives rise to long-lived memory T cells. Upon antigen re-encounter, memory T cells respond swiftly and robustly to eliminate the pathogen. Based on their tissue distribution, cell surface markers, and effector functions, memory T cells have been divided into two major subsets. Memory T cells expressing receptors such as CD62L and CCR7, which allow efficient homing to lymph nodes (LN), are termed central memory (TCM) cells; memory T cells lacking LN-homing receptors and preferentially residing in non-lymphoid tissues are termed effector memory (TEM) cells. Notably, both memory subsets display high levels of the marker CD44. Once established CD4 and CD8 memory T cell populations can be maintained for many years in vivo. Although early studies suggested that continued antigenic stimulation is required for maintaining T cell memory, neither cognate antigen nor MHC-encoded molecules appear to be required for the long-term survival of CD4 or CD8 memory T cells. Members of the tumor necrosis factor receptor (TNFR) family are involved in T cell costimulation and play a role in T cell survival. Mice deficient in OX40, CD27, or 4-1BB show greater defects in secondary compared to primary responses when infected with pathogens such as lymphocytic choriomeningitis virus (LCMV), vesicular stomatitis virus, and influenza.We developed a method of separating naive from memory T cells based upon their density. Using this technique, we made the following findings (especially with regard to CD8 T cell memory, which is important in anti-viral and anti-neoplastic responses):1. Memory T cells exist in the G1 phase (high RNA, haploid DNA) of the cell cycle, whereas naive T cells exist in G0 (low RNA, haploid DNA). Stimulation of the former results in rapid release of high amounts of cytokines such as interferon gamma (IFNgamma).2. Culture of purified memory T cells in the absence of other cells (resting) allows them to revert to G0. When they do, they respond to stimulation to the same degree and with the same kinetics as naive T cells. Culture of memory T cells with purified dendritic cells (DC) prevents their reversion to G0 and maintains their characteristic potent effector responses. Blockade of two TNFR molecules, CD70 and 4-1BB reverses the "protection" provided by DC. Furthermore, direct stimulation of the CD70 receptor on T cells, CD27, also maintains T cells in the memory state in the absence of any other cells. These results were verified with memory T cells generated against lymphocytic choriomeningitis virus (LCMV) and identified by MHC-peptide "tetramers", which can identify individual antigen-specific cells.3. CD27-deficient mice, memory T cells responsive to vesicular stomatitis virus (VSV) have difficulty maintaining their G1 state.4. Ligation of CD27 activated phosphatidylinositol-3 kinase (PI3K) and Akt. This in turn resulted in phosphorylation of the quiescence transcription factor FOXO1 and its export from the nucleus (deactivation). Notably, whereas freshly isolated naive T cells expressed a substantial amount of FOXO1, levels were markedly reduced in memory T cells. These results account for the requirement for CD27 signaling inmemory T cell maintenance. PI3K/Akt activate mammalian target of rapamycin (mTOR), which plays a vital role in protein translation and cell cycle progression. FOXO1 is important for maintaining cells in a quiescent state (i.e. in G0).Based upon this work, we have generated CD70-deficient mice. We have found that these animals have a blunted primary response to LCMV and fail to clear the virus normally. In contrast, the generation of memory T cells appears to be relatively normal. We are performing experiments now to test the function of such memory T cells. The data thus far suggest that CD70 (and by extension its receptor, CD27), has a substantial role in primary immune responses, but does not play a non-redundant role in generation of T cell memory.In parallel studies with wild type animals, we repeatedly observed a small population of cells in the lymph nodes and spleen that expressed an alpha/beta T cell receptor (TCR), CD11c, and MHC class II (the latter two being hallmarks of conventional DC). These cells, which we call TDC, have been characterized extensively at the cDNA and protein level. TDC, which can be either CD4+ or CD8+, but not both, require a thymus to develop like conventional T cells. However, like DC, they require FLT3L to develop. cDNA microarray analysis found that TDC express many genes heretofore found uniquely in T cells (TCR and CD3 genes) or DC (PU.1 and Zbtb46), but not both, as well as a distinctive signature of cytotoxicity-related genes. Unlike other T cell subsets characterized as innate, TDC had a polyclonal TCR repertoire and responded to cognate antigen. However, they differed from conventional T cells in that they did not require help from antigen-presenting cells. Strikingly, TDC responded to TLR-mediated stimulation by producing IL-12 and processed and presented antigen to MHCII-restricted T cells. TDC, as defined by surface phenotype and the ability to produce IL-12 when stimulated via TLRs, were identified in human peripheral blood as well. TDC, therefore, represent a heretofore unappreciated cell subset that combines key features of both innate and adaptive immune cells.

免疫系统由两个主要的武器组成：先天和适应性。先天免疫细胞，尤其是树突状细胞（DC），在对控制病原体的早期反应中至关重要，并且在激活适应性反应中起着至关重要的作用。后者是由淋巴细胞介导的，对于处理急性威胁（例如次生再曝光）（例如，第二种感染同一生物体）很重要。一级暴露于抗原时，带有互补抗原受体（TCR）的中含Have T细胞会经历快速激活和克隆膨胀，从而导致效应T细胞的产生。抗原清除后，参与原发性免疫反应的绝大多数（90-95％）抗原特异性效应T细胞会经历凋亡（细胞死亡）。然而，一个小子集生存并产生长寿命的记忆T细胞。抗原重新探测后，记忆T细胞迅速而稳健地反应以消除病原体。根据其组织分布，细胞表面标记和效应子功能，记忆T细胞已分为两个主要子集。表达受体（例如CD62L和CCR7）的记忆T细胞，这些受体允许有效地归因于淋巴结（LN），称为中央记忆（TCM）细胞；缺乏LN的受体和优先驻留在非淋巴组织中的记忆T细胞称为效应子记忆（TEM）细胞。值得注意的是，两个内存子集都显示出高级别的标记CD44。一旦建立的CD4和CD8记忆T细胞种群可以在体内维持多年。尽管早期的研究表明，维持T细胞记忆需要持续的抗原刺激，但对于CD4或CD8记忆T细胞的长期存活似乎不需要同源抗原和MHC编码的分子。肿瘤坏死因子受体（TNFR）家族的成员参与T细胞共刺激，并在T细胞存活中起作用。与原发性反应相比，与原发性反应相比，缺乏OX40，CD27或4-1BB的小鼠在感染了病原体（例如淋巴细胞绒毛膜脑膜炎病毒病毒（LCMV）），小囊炎病毒和流感时的小鼠缺陷。使用此技术，我们做出了以下发现（尤其是关于CD8 T细胞记忆，这在抗病毒和抗塑性反应中很重要）：1。 记忆T细胞存在于细胞周期的G1相（高RNA，单倍体DNA）中，而天真的T细胞存在于G0（低RNA，单倍体DNA）中。对前者的刺激导致大量细胞因子（例如干扰素伽马（Ifngamma））的迅速释放。2。在没有其他细胞（静止）的情况下，纯化的记忆T细胞的培养使它们可以恢复为G0。当他们这样做时，他们对刺激的程度和动力学与幼稚T细胞相同。用纯化的树突状细胞（DC）的记忆T细胞培养可阻止其重新转换为G0，并保持其特征性的有效效应子响应。两个TNFR分子CD70和4-1BB的封锁逆转了DC提供的“保护”。此外，在没有任何其他细胞的情况下，在T细胞上直接刺激CD27上的CD27受体也将T细胞保持在记忆状态。用针对淋巴细胞脉络膜宿主性炎病毒（LCMV）产生的记忆T细胞验证了这些结果，并通过MHC肽“四聚体”鉴定，该细胞可以鉴定出可以鉴定单个抗原特异性细胞。3。 CD27缺乏小鼠，对囊泡气孔病毒（VSV）反应的记忆T细胞难以维持其G1状态。4。 CD27激活的磷脂酰肌醇-3激酶（PI3K）和Akt的连接。反过来，这导致了静脉转录因子FOXO1及其从细胞核出口（停用）的磷酸化。值得注意的是，尽管新鲜分离的幼稚T细胞表达了大量FOXO1，但在记忆T细胞中的水平明显降低。这些结果解释了CD27信号传导含量T细胞维护的需求。 PI3K/AKT激活雷帕霉素（MTOR）的哺乳动物靶标，该靶标在蛋白质翻译和细胞周期进程中起着至关重要的作用。 FOXO1对于将细胞保持在静止状态很重要（即在G0中）。基于这项工作，我们已经产生了CD70缺陷小鼠。我们发现，这些动物对LCMV的主要反应钝，无法正常清除病毒。相反，记忆T细胞的产生似乎相对正常。我们现在正在执行实验来测试此类内存T细胞的功能。 The data thus far suggest that CD70 (and by extension its receptor, CD27), has a substantial role in primary immune responses, but does not play a non-redundant role in generation of T cell memory.In parallel studies with wild type animals, we repeatedly observed a small population of cells in the lymph nodes and spleen that expressed an alpha/beta T cell receptor (TCR), CD11c, and MHC class II (the latter two being常规DC的标志）。这些称为TDC的细胞已在cDNA和蛋白质水平上进行了广泛的特征。可以是CD4+或CD8+的TDC，但并非两者都要求胸腺像常规T细胞一样发展。但是，像DC一样，它们需要FLT3L才能开发。 cDNA微阵列分析发现，TDC表达许多在T细胞（TCR和CD3基因）或DC（PU.1和ZBTB46）中发现的许多基因，但并非兼具与细胞毒性相关基因的独特特征。与其他特征为先天性的T细胞子集不同，TDC具有多克隆TCR曲目，并对同源抗原做出了反应。但是，它们与常规T细胞不同，因为它们不需要抗原呈递细胞的帮助。令人惊讶的是，TDC通过产生IL-12并对MHCII限制的T细胞进行加工并呈现抗原对TLR介导的刺激做出了反应。 TDC在通过TLR刺激时由表面表型和产生IL-12的能力定义，也在人的外周血中鉴定出来。因此，TDC代表了迄今未批准的细胞子集，该细胞子集结合了先天性和适应性免疫细胞的关键特征。