Tfh细胞来源exosomes调控B细胞的机制及其在小鼠胶原诱导性关节炎发病中的作用

Follicular helper T cells (Tfh) have been identified as a new subset of effector helper CD4+T cells that are essential in regulating the development of B cell immunity. Tfh and B cells interactions are critical for germinal center (GC) reaction. Usually, most of Tfh-B contacts in GC are even less than 5 min, but it is not clear how Tfh cells provide signaling to B cells in this transient process, and whether other intercellular communication was involved in Tfh-B contacts. Exosomes are small membrane-bound vesicles (30-100nm) that are secreted in extracellular medium by hemopoietic and nonhemopoietic cells. Exosomes have important roles in intercellular communication by delivering bioactive molecules, including proteins, lipid and RNAs to the target cells. Exosomes are unable to change their phenotypes following purification and are very stable. The production of Tfh cells derived exosomes (Tfh-EXO) is enhanced by stimulation signal. In vitro, Tfh-EXO promoted the activation and Ig production of B cells by CD40L molecule. Tfh-EXO were able exacerbates the onset of murine collagen-induced arthritis (CIA) as well as increase severity of established arthritis. In this study, we will test how B cells regulate production of Tfh-EXO by ICOSL molecule. We will clarify the function and mechanism of CD40L from Tfh-EXO in regulating B cells, and find other functionally active molecules. More importantly, we will observe that Tfh-EXO exacerbates murine CIA by regulating GC reaction. This project will reveal a novel intercellular communication between Tfh and B cells, and provide an in-depth knowledge of humoral immunity and autoimmune diseases.

Tfh-B细胞的相互作用是生发中心反应的关键步骤，然而两者直接接触的过程非常短暂（通常<5分钟），Tfh细胞如何在短时间内完成其对B细胞的信号刺激，以及通过哪些方式发挥细胞间信息传递，值得我们进一步研究。Exosomes是细胞主动分泌到胞外的一种传递细胞间信息的膜性囊泡，且性质稳定，已有文献报道T细胞可通过分泌exosomes发挥重要的生物学功能。多种刺激可促进Tfh细胞分泌exosomes（Tfh-EXO），Tfh-EXO可促进B细胞活化并产生IgG，且这种作用可能与CD40L有关；Tfh-EXO可明显促进小鼠胶原诱导性关节炎（CIA）的发病。本项目拟进一步研究B细胞如何通过ICOSL对Tfh分泌exosomes发挥作用，明确Tfh-EXO携带的CD40L对B细胞的调控作用，并寻找其它可能发挥重要作用的分子；观察Tfh-EXO如何通过调控生发中心反应影响CIA的发病。这将揭示一种新的Tfh-B细胞互作模式，为了解机体体液免疫应答及自身免疫性疾病提供新思路。

类风湿关节炎（RA）是一种慢性自身免疫性疾病，伴有关节和全身炎症，通常产生大量的自身抗体，包括类风湿因子和抗CCP抗体。滤泡辅助性T细胞（Tfh）是辅助B细胞的最重要CD4+T细胞亚群。虽然Tfh细胞对于B细胞产生高亲和力抗体至关重要，但Tfh细胞的过度活化与许多自身免疫性疾病（如RA）的发病有关。细胞外小囊泡（sEV）是由母细胞释放的纳米级球形膜颗粒，通过向靶细胞传递蛋白质、脂质和RNA等生物活性分子，在细胞间通讯中发挥重要作用。在本项目中，我们通过体内外实验发现Tfh细胞来源的sEV促进B细胞分化和抗体产生，Tfh细胞来源sEV主要通过CD40L发挥调控作用，SNX27参与了sEV中CD40L的表达；ICOSL/ICOS信号是促进Tfh细胞分泌sEV的重要因素。在此基础上，我们发现CIA小鼠Tfh细胞分泌的sEV明显增加，并可通过sEV促进CIA发病；类风湿性关节炎（RA）患者血浆中CD4+CXCR5+ sEVs上CD40L的表达量与疾病活动度呈现正相关。本研究揭示了Tfh细胞调控B细胞生物学功能的新机制，为自身免疫性疾病的治疗提供新思路。