Co-stimuli for Human Marginal Zone B Cell Activation

描述(由申请人提供)：边缘带(MZ)B细胞与小鼠的SLE样自身免疫有关，因此可能在人类SLE的诱导中起重要作用。这一建议是基于这样一个假设，即MZ B细胞的几个方面(即C3dg结合CD21的水平升高，它们对BAFF的接近和敏感性，以及它们对微生物的Toll样受体(TLR)结合DNA的接近和可能的敏感性)可能会增强MZ B细胞对弱BCR刺激的反应能力。重要的是，后者是自身反应性B细胞与自身抗原(Ag)相互作用的特征，C3dg包被的凋亡细胞以表面抗原的形式表达细胞内分子，很可能在MZ内普遍存在。该应用的长期目标是阐明MZ环境中的刺激和人类MZ B细胞表面受体表达有助于增强MZ中发现的自身反应性B细胞的激活的机制。使用分离的人脾LGM+LGD-CD27+MZ B细胞和LGM+LGD+CD27-卵泡(FO)B细胞，我们将：(A)比较IL-4和BAFF在促进MZ和FO B细胞活性和防止BCR触发的凋亡方面的有效性；(B)比较MZ和FO B细胞BAFF和IL-4的受体密度；(C)与一系列亲和力不同的抗BCR：葡聚糖结合物和CD21结合部位，确定BCR与CD21：CD19共刺激复合体结合的程度：(I)在限制BCR参与的条件下，促进依赖BAFF的MZ和FO B细胞的增殖；(Ii)与BAFF和TLR9结合的CpG DNA基序协同促进MZ或FO B细胞的分化；(D)探讨同型CD27：CD7O相互作用在上述刺激激活时促进MZ B细胞分化的作用；以及(E)检测在BAFF和微生物DNA基序存在的情况下，正常人MZ中的自身反应性B细胞是否被触发分泌自身抗体。上述具有明确体外刺激的功能研究应该阐明，在MZ环境中预期的刺激，特别是在微生物感染期间，是否可以在限制BCR参与的条件下协同促进人B细胞的激活。这样的见解应该建议采用免疫干预疗法来阻断SLE中自身反应性B细胞的激活。

DESCRIPTION (provided by applicant): Marginal zone (MZ) B cells have been implicated in SLE-like autoimmunity in mice and thus may be important in the induction of SLE in man. This proposal is based on the hypothesis that several facets of MZ B cells (i.e. heightened levels of C3dg-binding CD21, their proximity and sensitivity to BAFF, and their proximity and possible sensitivity to Toll-like receptor (TLR)-binding DNA from micro-organisms) may augment the capacity of MZ B cells to respond to weak BCR stimuli. Importantly, the latter characterizes the interaction of autoreactive B cells with self antigen (Ag), and C3dg-coated apoptotic cells expressing intracellular molecules as surface Ag are likely prevalent within the MZ. The application's long-term objectives are to illuminate the mechanisms by which stimuli within the MZ environment and human MZ B cell surface receptor expression contribute to the enhanced activation of autoreactive B cells which are found within the MZ. Using isolated lgM+lgD-CD27+ MZ B cells and lgM+lgD+CD27- follicular (FO) B cells from human spleens, we will: (a) compare the efficacy of IL-4 and BAFF at promoting MZ and FO B cell viability and preventing BCR-triggered apoptosis, (b) compare MZ and FO B cells for density of receptors for BAFF and IL-4, (c) with a series of affinity-diverse anti-BCR:dextran conjugates ¿ CD21 binding sites, determine the degree to which BCR ligation with the CD21 :CD19 costimulatory complex (i) augments BAFF-dependent MZ and FO B cell proliferation under conditions of limiting BCR engagement and (ii) collaborates with BAFF and TLR9-binding CpG DNA motifs to enhance MZ or FO B cell differentiation, (d) explore the role of homotypic CD27:CD7O interactions in promoting the differentiation of MZ B cells upon activation by the above stimuli, and (e) examine whether autoreactive B cells in the MZ of normal individuals are triggered to secrete autoantibody when cultured with C3dg-coated apoptotic cells in the presence of BAFF and microbial DNA motifs. The above functional studies with defined in vitro stimuli should elucidate whether stimuli expected in the MZ environment, particularly during microbial infection, can collaborate in promoting the activation of human B cells under conditions of limiting BCR engagement. Such insights should suggest immunologic intervention therapies for blocking the activation of autoreactive B cells in SLE.