Microbial sensing and innate immune activation in B cells

B细胞主要以其产生抗体的能力而闻名。然而，越来越多的文献表明，它们也可能通过各种抗体非依赖性机制参与免疫应答，如细胞因子产生、共刺激和抗原呈递。B细胞可分为三种不同的亚群：滤泡B细胞、先天性样边缘区B细胞（MZ B）和B1 B细胞。滤泡B细胞是最主要的B细胞亚群，存在于二级和三级淋巴器官的淋巴滤泡中。MZB位于脾的边缘区，具有结合免疫复合物、向脾白色髓迁移以及将抗原转移至滤泡树突状细胞的能力。已知B1 B细胞分泌天然抗体。 我们最近报道，B细胞可以捕获原虫寄生虫杜氏利什曼原虫。一旦接触到寄生虫，B细胞就会形成簇，并将L. donovani在细胞外富含IgM的口袋里。这种密切的相互作用导致多克隆B细胞活化，并最终在24- 48小时后导致细胞死亡。有趣的是，寄生虫触发内体Toll样受体（TLR），尽管它在细胞外附着在B细胞的细胞表面上。内体TLR的激活是诱导IL-10和IFN-I表达和促进高丙种球蛋白血症所必需的。B细胞内体TLR激活的上游和下游途径以及这些途径对高丙种球蛋白血症诱导的贡献尚不清楚。这些步骤的定义将需要更深入地了解B细胞如何与寄生虫相互作用。 我们研究的总体目标是了解各种B细胞亚群如何识别微生物并对其做出反应，以及这种识别如何影响其功能。我们的目的是在这个应用程序中的特征之间的相互作用L。donovani和B细胞，以了解导致多克隆B细胞活化的途径。特别是，我们将研究i）寄生虫捕获和细胞表面之间的相互作用L。donovani和各种B细胞亚群; ii）内体toll样受体和其他微生物传感器（主要是胞质）的寄生虫识别以及它们激活的下游途径;和iii）细胞间通讯（例如隧道纳米管;受体/配体相互作用）以及这种通讯在诱导多克隆B细胞激活中的作用。 转基因寄生虫，基因敲除小鼠，共聚焦显微镜，流式细胞术，和几个生化技术将被用来解剖在体外实验中的各种激活和细胞通讯途径。 这里确定的途径可以揭示新的策略，也可以用于其他微生物激活B细胞和调节它们的功能。

B-cells are mostly known for their capacity to produce antibodies. However, an increasing body of literature has now shown that they may also participate in the immune response by various antibody-independent mechanisms, such as cytokine production, co-stimulation, and antigen presentation. B-cells can be divided in three different subpopulations: follicular B cells , innate-like marginal zone B-cells (MZB), and B1 B-cells. Follicular B cells are the most prominent B-cell subpopulation and reside in lymphoid follicles of secondary and tertiary lymphoid organs. MZB are located in the marginal zone of the spleen and are endowed with the capacity to bind immune complexes, migrate towards the splenic white pulp, and transfer antigen to follicular dendritic cells. B1 B-cells are known for their secretion of natural antibodies. We have recently reported that B cells can capture the protozoan parasite Leishmania donovani. Upon exposure to the parasite, B cells form clusters and hold L. donovani in extracellular IgM-rich pockets. This close interaction results in polyclonal B cell activation and eventually in cell death after 24-48h. Interestingly, the parasite triggers endosomal Toll-Like Receptors (TLRs), although it is attached extracellularly on the cell surface of the B cell. Activation of endosomal TLRs was required to induce IL-10 and IFN-I expression and to promote hypergammaglobulinemia. The pathways up- and downstream of endosomal TLR activation in B-cells and the contribution of these pathways to the induction of hypergammaglobulinemia are as yet unknown. Definition of these steps will require a deeper understanding of how B cells interact with the parasite. The overall goal of our research is to understand how various B cell subpopulations recognize and react to microorganisms, and how this recognition affects their function. Our objective in this application is to characterize the interaction between L. donovani and B cells in order to understand pathways leading to polyclonal B cell activation. Particularly, we will investigate i) parasite capture and cell surface interaction between L. donovani and various B cell subpopulations; ii) parasite recognition by endosomal toll-like receptors and other microbial sensors (mainly cytosolic) and the downstream pathways they activate; and iii) cell-to-cell communication (e.g. tunnelling nanotubules; receptor/ligand interactions) and role of this communication in the induction of polyclonal B cell activation. Transgenic parasites, knock-out mice, confocal microscopy, flow cytometry, and several biochemical techniques will be used to dissect the various activation and cellular communication pathways in in vitro experiments. The pathways identified here could uncover novel strategies that may also be used by other microorganisms to activate B cells and modulate their functions.