Core--Chimerism and Immunologic Recovery

该核心的目的是提供一个集中的实验室资源，以监测GVHD预防对供体/宿主嵌合体程度和移植后免疫重建动力学的影响。 该核心还负责冷冻保存DFCI/BWH接受异基因造血干细胞移植的所有患者的外周血细胞、骨髓细胞和血清或血浆。 移植前，将从所有患者和正常供体中获得外周血单核细胞（PBMC）和血清。异基因造血干细胞移植（HSCT）后，将定期冷冻保存PBMC、骨髓和血清。通过VNTR或STR的PCR分析以及单核苷酸多态性（SNP）的快速焦磷酸测序，评估再生骨髓和循环淋巴细胞的嵌合性。将通过流式细胞术定量测量具有确定表型的循环淋巴细胞来评估HSCT后的免疫重建。此外，T细胞功能和分化的几种新的测定法已被用于检查T细胞重建。 具体而言，将通过TCR V β谱分析方法检查T细胞受体库的重建。 将通过定量PCR测量PBMC中的T细胞受体切除环（TREC）来检查胸腺功能的重建。这将提供一种新的测量T细胞新生。该核心也将能够使用ELISPOT或HLA-肽四聚体提供对特异性抗原的T细胞应答的评估。基于SNP的嵌合分析还将提供已知次要组织相容性抗原差异的基因分型。 这些测定的结果将可用于与免疫功能的其他参数以及与临床结果的相关性。冻存的活细胞和血清样品也将提供给本计划中每个研究项目的研究人员。

The purpose of this core is to provide a centralized laboratory resource to monitor the effect of GVHD prophylaxis on the degree of donor/host chimerism and the kinetics of immune reconstitution post transplant. This core is also responsible for cryopreservation of peripheral blood cells, marrow cells and serum or plasma from all patients undergoing allogeneic hematopoietic stem cell transplantation at DFCI/BWH. Prior to transplantation, peripheral blood mononuclear cells (PBMC) and serum will be obtained from all patients and normal donors. Following allogeneic hematopoietic stem cell transplantation (HSCT), PBMC, bone marrow and serum will be cryopreserved at regular intervals. The regenerating marrow and circulating lymphoid cells will be assessed for chimerism by PCR analysis of VNTR or STR as well as by rapid pyrosequencing of single nucleotide polymorphisms (SNP). Immune reconstitution after HSCT will be assessed by quantitative measurement of circulating lymphocytes with defined phenotype by flow cytometry. In addition, several new assays of T cell function and differentiation have been adapted to examine T cell reconstitution. Specifically, reconstitution of T cell receptor repertoire will be examined by the method of TCR Vbeta spectratyping. Reconstitution of thymic function will be examined by quantitative PCR measurement of T-cell receptor excision circles (TREC) in PBMC. This will provide a novel measurement of T cell neogenesis. This core will also be able to provide assessment of T cell responses to specific antigens using either ELISPOT or HLA-peptide tetamers. SNP-based chimerism analysis will also provide genotyping for known minor histocompatibility antigen disparity. Results of these assays will be available for correlation with other parameters of immune function as well as with clinical outcomes. Cryopreserved samples of viable cells and serum will also be available to investigators in each of the research projects within this program.