TRANSPLANT IMMUNOMODULATION BY ALLOCHIMERIC MOLECULES

描述(由申请人提供)： 我们试图开发一种临床适用的策略，以诱导使用同种异体I类MHC分子治疗的“真正的”移植耐受。我们在大鼠同种异体心脏移植模型上的新发现为这一提议提供了基础。我们假设：(I)异基因嵌合诱导的耐受适用于遗传多样性的同种异体移植受者；(Ii)异基因嵌合分子通过间接递呈调节免疫反应；(Iii)异基因嵌合分子产生具有独特功能和TCR同种特异性的独特调节性T细胞。我们建议：1.确定不同的AII化学分子诱导耐受的要求。在不同的菌株组合中，将构建多个同种嵌合分子，以定义I类MHC多态区域的关键免疫原性表位。在受体I类抗原上显示供体主导表位的融合分子将接受测试，以确定它们是否有能力诱导“真正的”慢性无排斥耐受。2.间接提呈异基因[Alpha1hI/u]-RT1Aa-I类MHC分子分析AIIo免疫调节。将对免疫原性/隐蔽自身表位进行精细定位，这些表位对同种异体移植耐受至关重要。T细胞反应的间接同种嵌合偏差将通过使用宿主或供体类型的树突状细胞来解决，这些树突状细胞已在体外与供体野生型或Alpha1h1/u]-RT1.Aa分子冲击。同种异体蛋白质的标记和检测将定位同种异体的加工和表达所涉及的部位和细胞类型。同种异体耐受诱导所需的“未成熟”和“成熟”肝脏DC将在体内通过功能刺激DC成熟和使用造血生长因子FIt3L的同种异体刺激能力来研究，以潜在地消除门静脉内异体移植后间接诱导的耐受。3.鉴定间接同种异体识别诱导的独特调节性T细胞群。我们将进行体内和体外调节性T细胞的表型和功能表征。在这个模型中，调节性T细胞的异体嵌合产生是一个可能的关键耐受机制，将通过研究未成熟的肝树突状细胞间接向CD4T细胞递送同种嵌合分子来在体外探索。第三，在体内分析慢性排斥反应，结合CDR3分型、免疫显微镜和同种异体诱导的克隆性限制性调节性T细胞的序列分析，将确定独特的T细胞功能特异性和同种异体嵌合序列对同种异体特异性TCR谱系的影响。

DESCRIPTION (provided by applicant): We seek to develop a clinically applicable strategy to induce "true" transplantation tolerance treated with allochimeric Class I MHC molecules. Our novel findings in a rat cardiac allograft model provide the foundation of this proposal. We hypothesize: (i) allochimeric-induced tolerance is applicable to genetically diverse allograft recipients; (ii) allochimeric molecules modulate the immune response by indirect presentation; (iii) aIlochimeric molecules generate unique regulatory T cells with distinctive functional and TcR allospecificities. We propose: 1. To Define the Requirements of Different AIIochimeric Molecules to Induce Tolerance. Multiple allochimeric molecules, in different strain combinations, will be constructed to define critical immunogenic epitopes in the polymorphic regions of class I MHC. AIIochimeric molecules that display donor-dominant epitopes on recipient class I antigens will be tested for their ability to induce "true" chronic rejection-free tolerance. 2. To Analyze AIIoimmune Modulation by Indirect Presentation of Allochimeric [alpha1h I/u]-RT1.Aa Class I MHC Molecules. Fine mapping of immunogenic/cryptic self-epitopes that are critical for allograft tolerance will be performed. Indirect allochimeric deviation of T cell responses will be addressed by employing host or donor-type dendritic cells that have been pulsed in vitro with donor wild-type or alpha1h 1/u]-RT1.Aa molecules. AIIochimeric protein labeling and detection will localize the site and cell-type involved in allochimeric processing and presentation. Requirement of 'immature vs mature' hepatic DC for allochimeric tolerance induction will be investigated in vivo by functional stimulation of DC maturity and allostimulatory capacity using FIt3L, a hematopoietic growth factor, to potentially abrogate indirectly induced tolerance following intra-portal allochimeric delivery. 3. To Characterize the Unique Population of Regulatory T Cells Induced by Indirect Allorecognition. We will perform in vivo and ex vivo phenotypic and functional characterization of regulatory T cells. Allochimeric generation of regulatory T cells, a possible key tolerogenic mechanism in this model, will be probed in vitro by studying immature hepatic dendritic cells that indirectly present allochimeric molecules to CD4+ T cells. Third, in vivo analysis of chronic rejection coupled with CDR3 spectrotyping, immunoscope and sequence analysis of allochimeric-induced clonally-restricted regulatory T cells will define unique T cell functional specificities and the influence of allochimeric sequence on the allospecific TcR repertoire.