Co-stimuli for Human Marginal Zone B Cell Activation

描述（由申请人提供）：边缘区（MZ） B细胞与小鼠SLE样自身免疫有关，因此可能在诱导人类SLE中起重要作用。这一建议是基于这样的假设，即MZ B细胞的几个方面（即c3d结合CD21水平的提高，它们对BAFF的接近性和敏感性，以及它们对来自微生物的toll样受体（TLR）结合DNA的接近性和可能的敏感性）可能增强MZ B细胞对弱BCR刺激的反应能力。重要的是，后者表征了自身反应性B细胞与自身抗原（Ag）的相互作用，c3d包被的凋亡细胞表达细胞内分子作为表面Ag可能在MZ中普遍存在。该应用程序的长期目标是阐明MZ环境中的刺激和人类MZ B细胞表面受体表达促进MZ内发现的自身反应性B细胞活化的机制。使用人脾脏分离的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细胞活力和防止bcr引发的细胞凋亡方面的功效，(B)比较MZ和FO B细胞中BAFF和IL-4受体的密度，(c)一系列亲和不同的抗bcr：葡聚糖结合物CD21结合位点。确定BCR与CD21:CD19共刺激复合物结合的程度(i)在限制BCR结合的条件下增强BAFF依赖的MZ和FO B细胞的增殖，（ii）与BAFF和tlr9结合的CpG DNA基序合作以增强MZ或FO B细胞的分化，(d)探索同型CD27: cd70相互作用在促进MZ B细胞在上述刺激激活后的分化中的作用。(e)检测在BAFF和微生物DNA基序存在的情况下，与c3d包被的凋亡细胞一起培养时，正常人MZ中的自身反应性B细胞是否会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阐明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。这些发现提示免疫干预疗法可以阻断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.