ROLE OF TNF FAMILY MEMBERS IN MEMORY B CELL DEVELOPMENT

近年来，相当多的注意力集中在参与调节幼稚B细胞向记忆B细胞区室分化的因素上。 不幸的是，记忆B细胞发育的研究一直很困难，因为这些B细胞的频率极低。 我们已经开发了一种体内模型系统，其中B细胞分化的事件以高频率发生，因此将允许对参与建立记忆的必要因素进行透彻的评价。 该系统的新特征包括利用IG转基因B细胞（称为QM B细胞），所述细胞是抗原特异性的，并且具有同种型转换和经历体细胞突变的能力。 待检验的基本假设是TNF家族成员（TNF α、LT α、OX40 L、CD40 L）差异性地调节记忆B细胞的产生和维持。 具体目的有两个：1）表征响应于T细胞依赖性抗原而被诱导分化为记忆B细胞的QM B细胞，和2）使用TNF家族成员的阻断剂来评估QM B细胞分化被破坏的阶段。 这些将通过进行过继转移实验来实现，表型分析将通过流式细胞术和多色共聚焦成像来评估。 将通过二次转移实验和QM B细胞产生高亲和力、同种型转换、体细胞突变抗体的能力来检查QM B细胞向记忆B细胞池的分化。这些研究将阐明参与记忆B细胞分化的因素，并有助于疫苗的开发以及自身免疫性疾病的免疫治疗。

In recent years, considerable attention has been focused on the factors involved in regulating the differentiation of naive B cells to the memory B cell compartment. Unfortunately, the study of memory B cell development has been difficult due to the extremely low frequency of these B cells. We have developed an in vivo model system in which the events of B cell differentiation occur at a high frequency, and thus will allow for the incisive evaluation of the requisite factors involved in establishing memory. The novel features of this system include the utilization of Ig transgenic B cells (referred to as QM B cells) which are antigen-specific, and have the ability to isotype switch and undergo somatic mutation. The underlying hypothesis to be tested is that TNF family members (TNFalpha, LTalpha, OX40L, CD40L) differentially regulate the generation and maintenance of memory B cells. The specific aims are two-fold: 1) to characterize QM B cells induced to differentiate into memory B cells in response to a T cell-dependent antigen, and 2) use blocking reagents to TNF family members to assess the stage(s) in which QM B cell differentiation is disrupted. These will be achieved by performing adoptive transfer experiments and phenotypic analysis will be assessed by flow cytometry and multi-color confocal imaging. QM B cell differentiation into the memory B cell pool will be examined by secondary transfer experiments and the capacity of QM B cells to generate high affinity, isotype switched, somatically mutated antibodies. These studies will elucidate factors involved in memory B cell differentiation, and contribute to vaccine development as well as immunotherapy for autoimmune disease.