COMPARATIVE ONTOGENETIC APPROACH TO VACCINE STRATEGY

该项目集中在B细胞生产的两个主要方面， 在人类中的功能：处女和记忆B细胞的产生。的 造血祖细胞必须经历的分化步骤， 将分析产生克隆多样性的原始B淋巴细胞， 这些实验着眼于B可能的临床操作 细胞生产建议进行研究，以剖析（一）的精确顺序 B系分化事件，（ii）转录活性的顺序 在这一进程中重要的基因和（iii）组装，表达和 作为分化阶段的函数的抗原受体单位的功能 和年龄。新的线索表明骨髓中的B细胞 可以通过非抗原特异性的 受体（IL-7，IFN α/β和CDI 9）将在一系列研究中进行探索。 实验 假设B细胞可以由一种 替代分化途径，涉及轻链基因 在常规重链基因重排之前重排， 会得到考验在这些实验中，⑴“替代”重链（MC） 将κ轻链与Igalpha/β信号转导偶联 将分离和表征两个B系细胞系上的单位，（ii） 克隆、测序和表达替代RC基因，（iii）单克隆 针对替代HC产生的抗体，以及（iv）使用的这些试剂 作为探针来识别这一通路中的骨髓细胞。研究将 进行追踪初步数据，表明抗体库 可能在通过该替代途径产生的B细胞中不同。的 记忆B细胞的表型和功能特征将被 作为年龄的函数进行检查。在这些实验中，IgA 1和IgA 2 B细胞 将从成人和脐带血样本中纯化，用于分析其 (i)细胞表面抗原谱和活化状态，（ii）机制 同种型开关，（iii）抗体库，（iv） 发育调控基因，和（v）功能能力。这些 原始IgM B细胞及其IgA 1和IgA 2记忆B细胞的研究 子代可为疫苗的设计提供有价值的信息 不同年龄段的人的策略。

This project focuses on two major aspects of B cell production and function in humans: virgin and memory B cell generation. The differentiation steps that hemopoietic progenitor cells must undergo to give rise to clonally diverse virgin B lymphocytes will be analyzed in these experiments with an eye toward possible clinical manipulation of B cell production. Studies are proposed to dissect the (i) precise order of B-lineage differentiation events, (ii) order of transcriptional activity of genes important in this progression and (iii) assembly, expression and function of antigen receptor units as a function of differentiation stage and age. Fresh leads suggesting that B cell production in the bone marrow can be regulated both positively and negatively via non-antigen-specific receptors (IL-7, IFNalpha/Beta and CDI9) will be explored in one series of experiments. The hypothesis that B cells can be generated by an alternative differentiation pathway, involving light chain gene rearrangement before the rearrangement of conventional heavy chain genes, will be tested. In these experiments, (i) 'surrogate' heavy chains (MC) that couple kappa light chains to the Igalpha/beta signal transduction units on two B-lineage cell lines will be isolated and characterized, (ii) the surrogate RC gene cloned, sequenced, and expressed, (iii) monoclonal antibodies produced against the surrogate HC, and (iv) these reagents used as probes to identify bone marrow cells in this pathway. Studies will be conducted to pursue preliminary data suggesting the antibody repertoire may differ in B cells generated via this alternative pathway. The phenotypic and functional characteristics of memory B cells will be examined as a function of age. In these experiments IgA1 and IgA2 B cells will be purified from adult and cord blood samples for analysis of their (i) cell surface antigenic profile and activation status, (ii) mechanism of the isotype switch, (iii) antibody repertoire, (iv) expression of developmentally-regulated genes, and (v) functional capabilities. These studies of virgin IgM B cells and their IgA1 and IgA2 memory B cell progeny should provide valuable information for the design of vaccine strategy in individuals of various ages.