Immunoregulation by TLR-activated TIM-1+ ProB Cells in Transplantation

TLR 激活的 TIM-1 ProB 细胞在移植中的免疫调节

• 批准号: 10455069
• 负责人: DAVID M ROTHSTEIN
• 金额: $ 43.75万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10455069
• 关键词: Address; Adult; Allograft Tolerance; Allografting; Antigens; B cell differentiation; B-Lymphocytes; Bone Marrow; Cell Count; Cell Lineage; Cells; Cues; Dependence; Engraftment; Environment; Genetic Transcription; Immune response; Impairment; Inflammatory; Interleukin-10; Knockout Mice; Lead; Ligation; MS4A1 gene; Maintenance; Mediating; Methods; Modeling; Monoclonal Antibodies; Nature; Pathway interactions; Peripheral; Phenotype; Population; Proliferating; Property; Protocols documentation; Receptor Signaling; Regulation; Regulator Genes; Regulatory T-Lymphocyte; Rest; Role; Signal Pathway; Site; Therapeutic; Time; Translating; Transplantation; Transplantation Tolerance; base; heart allograft; immunoregulation; islet allograft; knockout gene; member; mouse model; nano-string; single-cell RNA sequencing; transcriptome sequencing

Abstract (Project 2) Rather than use passive transfer of well differentiated adult immunoregulatory cells as a means to create transplant tolerance, we have discovered that a population CpG activated pro-B cells are extraordinarily potent, far more potent on a per cell basis than Tregs and far easier to prepare. Transfer of only 40,000, but not 90,000, induces permanent engraftment and probably tolerance in MHC incompatible islet and cardiac allograft mouse models. We have discovered that CpG induces expression TIM-1 a potential master switch for expression of an immunoregulatory module in mature Bregs (see Project 1/ Kuchroo). We also learned that ligation of TIM-1 by anti-TIM-1 mAb induces expression of IL-10, a member of the regulatory model discovered by Kuchroo. The stepwise contributions of CpG and activation of the TIM-1 pathway are examined through RNAseq and NanoString based analysis. To more precisely characterize the fate and phenotype, proliferative and differentiation of CpG activated pro-B cells, lineage-tracking methods are utilized throughout. We know that CpG activated pro-B cells proliferate and differentiate after transfer when introduced in optimal, not excessive, cell number. Depending upon environmental cues CpG activated pro-B cells can differentiate into a variety Bregs but CpG activated pro-B cells from RAG KO mice do not generate Bregs. In this project we will analyze the cellular basis of tolerance believing that CpG activated B regs and their progeny create an environment conducive to activation, expansion and retention of donor specific Tregs. The specific but multifaceted requirements for B cell differentiation, including the nature of the niche impairing expansion and differentiation of CpG-ProBs, in the acquisition of tolerance will be analyzed through use of a panel of carefully selected gene knockout mice. The role of T and B regs in the induction and maintenance of tolerance will be determined through time course protocols that selectively destroy Tregs or the mature progeny of CpG activate pro cells. The diversity of CpG-ProB progeny and their expression of a regulatory module that is IL-10 inclusive will be examined by single cell RNAseq of CpG-ProBs and their progeny at the transplant.

摘要（项目2） 而不是使用被动转移分化良好的成体免疫调节细胞作为一种手段， 移植耐受性，我们已经发现CpG激活的pro-B细胞群非常有效， 在每个细胞的基础上比Tclad更有效，而且更容易制备。转账仅4万，但不 90，000，诱导MHC不相容的胰岛和心脏移植物的永久植入和可能的耐受 小鼠模型。我们已经发现CpG诱导TIM-1表达，TIM-1是一个潜在的主开关， 在成熟的Bacteria中表达免疫调节模块（参见项目1/ Kuchroo）。我们还了解到 通过抗TIM-1 mAb连接TIM-1可诱导IL-10的表达，IL-10是发现的调控模型的成员 关于Kuchroo CpG的逐步贡献和TIM-1通路的激活通过以下方式进行检查： 基于RNAseq和NanoString的分析。为了更准确地描述命运和表型，增殖性 和CpG激活的前B细胞的分化，自始至终使用谱系追踪方法。我们知道 CpG激活的前B细胞在转移后增殖和分化，当引入最佳时， 细胞数量过多。根据环境因素，CpG激活的pro-B细胞可以分化为 但是来自RAG KO小鼠的CpG活化的pro-B细胞不产生BclB。在这个项目中，我们将 分析耐受性的细胞基础，认为CpG激活了B受体及其后代产生了一种免疫耐受性。 环境有利于激活、扩增和保留供体特异性TCLs。具体但 B细胞分化的多方面要求，包括损害扩增的小生境的性质， CpG-ProB在获得耐受性中的差异将通过使用一组仔细的 选择基因敲除小鼠。T和B受体在诱导和维持耐受性中的作用将在 通过选择性地破坏TdR或CpG激活的成熟后代的时程方案来确定 亲细胞。CpG-ProB子代的多样性及其包含IL-10的调节模块的表达 将通过CpG-ProB及其后代的单细胞RNAseq在移植时进行检查。