CONTROL OF PERSISTENT ANTIBODY RESPONSE BY FC RECEPTORS

抗原特异性抗体的长期存在和高度特异性记忆B细胞的产生是体液免疫反应的公认特征。生发中心(GC)的特殊微环境可能是这些途径的关键，特别是滤泡树突状细胞(FDC)以免疫复合体(IC)的形式长期保留免疫原性抗原的能力。已经提出，这种形式的抗原在初始免疫反应中既驱动高亲和力的B细胞受体体细胞突变体的选择，又在此后的较长时间内维持记忆B细胞室的稳定性。缺乏激活的Fc受体FcGammaRI和FcGammaRIII的小鼠表现出显著增加IC在FDCs上的沉积和滞留。然而，这不会导致初级免疫反应的扰动。在目标1中，我们提出了一种假设，即这种增强的IC沉积和在FDCs上的滞留将导致骨髓中持久血清抗体、记忆B细胞和抗体形成细胞(AFC)水平的升高。我们还将确定ICs在FDC上的滞留是否对第二种抗原的后续免疫稳定，以及当FDC上的同源抗原水平较高时，持久血清抗体的亲和力是否较低。在记忆的产生和维持过程中，B细胞的活性也可能通过抑制FCR FcGammaRIIB与抗原特异性B细胞直接相互作用来调节。缺乏这种FCR的小鼠在免疫后表现出较高的血清抗体和AFC水平，这与这一观点和过去对这种FCR功能的体外研究一致。然而，由于FcGammaRIIB是在许多髓系和FDC上发现的最广泛表达的FCR，这一结论仍然是初步的。在目标2中，我们建议测试FcGammaRIIB中观察到的抗体产生的异常表达的假设。缺陷小鼠是由于B细胞上这种FCR的缺失。最后，最近的数据表明，BCR信号抑制BCR-FcGammaRIIB共连接的机制是通过在FcGammaRIIB细胞质尾部的ITIM基序上募集肌醇多聚磷酸酶而介导的。突变形式的FcGammaRIIB在ITIM基序中Y-大于F的变化，排除了SHIP结合所需的Y磷酸化，不介导对BCR信号的抑制。在目标3中，我们建议使用在B细胞室表达FcGammaRIIB Y-大于F突变体的小鼠来验证这一假设，即ITIM-SHIP途径是唯一FcGammaRIIB启动的途径，是体内抑制B细胞活性所必需的。

Long-term persistence of antigen-specific antibody and the generation of highly specific memory B cells are well recognized features of the humoral immune response. The specialized microenvironment of the germinal center (GC) may be key to these pathways, in particular, the ability of the follicular dendritic cells (FDC) to retain immunogenic antigen in the form of immune complexes (ICs) for extended periods. It has been proposed that this form of antigen both drives the selection of high affinity B cell receptor somatic mutants during the primary immune response, and maintains the stability of the memory B cell compartment over extended periods thereafter. Mice deficient in the activation Fc receptors FcgammaRI and FcgammaRIII display dramatically enhanced IC deposition and retention on FDCs. However, this does not lead to perturbation of the primary immune response. In Aim 1, we propose to test the hypothesis that this enhanced IC deposition and retention on FDCs will result in elevated levels of persistent serum antibody, memory B cells, and antibody forming cells (AFCs) in the bone marrow. We will also determine whether retention of ICs on FDCs is stable to subsequent immunization with a second antigen, and whether the affinity of persistent serum antibody is lower when cognate antigen levels on FDC are higher. B cell activity during the generation and maintenance of memory may also be regulated by direct interaction of ICs with antigen specific B cells via the inhibitory FcR, FcgammaRIIB. Mice deficient in this FcR display elevated serum antibody and AFC levels after immunization, consistent with this idea and with past in vitro studies on the functioning of this FcR. However, since FcgammaRIIB is the most widely expressed FcR, found on many myeloid lineages and FDCs, this conclusion remains tentative. In Aim 2 we propose to test the hypothesis that the dysregulated expression of antibody production observed in FcgammaRIIB. deficient mice is due to absence of this FcR on B cells. Finally, recent data have shown that the mechanism of inhibition of BCR signaling upon BCR-FcgammaRIIB co-ligation is mediated by recruitment of the inositol polyphosphate phosphatase SHIP to the ITIM motif in the cytoplasmic tail of FcgammaRIIB. A mutant form of FcgammaRIIB with a Y-greater than F change in the ITIM motif, precluding the Y phosphorylation necessary for SHIP binding, does not mediate inhibition of BCR signaling in transfected cell lines. In Aim 3, we propose to test the hypothesis that the ITIM-SHIP pathway is the only FcgammaRIIB initiated pathway necessary for inhibition of B cell activity in vivo, using mice engineered to express the FcgammaRIIB Y-greater than F mutant in the B cell compartment.