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: 10214480
• 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 2023-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10214480
• 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） 而不是使用良好分化的成人免疫调节细胞的被动转移作为创造的一种手段 移植耐受性，我们发现种群CpG活化的Pro-B细胞非常有效， 比Tregs比Tregs更有效，并且更容易准备。仅转移40,000，但不转移 90,000，诱导永久性植入，可能在MHC不兼容的胰岛和心脏同种异体移植物中耐受性 鼠标模型。我们发现CPG诱导表达Tim-1一个潜在的主开关 成熟的布雷格人中免疫调节模块的表达（请参阅项目1/ kuchroo）。我们还了解到 通过抗TIM-1 MAB的结扎TIM-1诱导IL-10的表达，IL-10是发现的调节模型的成员 由川起。 CPG的逐步贡献和TIM-1途径的激活将通过 基于RNASEQ和纳米源分析。更精确地表征命运和表型，增殖性 和CpG激活的Pro-B细胞的分化，整个过程中都使用了谱系跟踪方法。我们知道 CpG激活的Pro-B细胞在最佳中引入时转移后扩散并分化，而不是 过多的细胞数。根据环境提示，CpG激活的pro-B细胞可以区分 Bregs，但CPG激活了来自RAG KO小鼠的Pro-B细胞，不会产生Bregs。在这个项目中，我们将 分析耐受性的细胞基础，认为CpG激活B regs及其后代会产生一个 有利于特定于供体特雷格的激活，扩展和保留的环境。具体但 B细胞分化的多方面要求，包括利基市场的性质损害扩展和 CPG探针的差异化，在获得耐受性时将通过使用一组小组进行分析 选定的基因敲除小鼠。 T和B规则在耐受性的诱导和维持中的作用将是 通过时间课程协议确定，这些方案有选择地破坏Treg或CpG的成熟后代激活 Pro细胞。 CpG-prob后代的多样性及其对IL-10包含的调节模块的表达 将通过CpG-probs的单细胞RNASEQ及其在移植时的后代进行检查。