NLRP3 Inflammasome Activation, T Follicular Helper Cells and Autoimmunity

摘要 系统性红斑狼疮（SLE）是一种多器官系统性自身免疫性疾病， 处于生育年龄的女性增生性肾小球肾炎（GN）患者通常进展至终末期， 肾脏疾病（ESRD）。50多年来，FDA一直没有批准LN的新疗法。 更好地了解LN的发病机制，以制定更有效的治疗策略。我们 实验室已经表明，免疫复合物（IC）本身不足以引起ESRD， 器官（肾脏）对损伤的抵抗力在决定LN的临床结果中是重要的。三种GN模型 已经建立。通过一种新的技术，我们已经在慢性肾小球肾炎中发现了独特的 CD 11b +IA-F4/80-浸润的肾小球内巨噬细胞是产生TNFα、足细胞的优势细胞。 系膜细胞是产生IL-1β/IL-18的优势细胞，系膜细胞是产生IL-6的优势细胞。提出了一个新的假设 TNFα、IL-1β、IL-18和IL-6是由患病肾小球中的不同细胞（区室）产生的， 这些细胞因子与其受体之间的相互作用促进肾小球内持续的炎症 导致足细胞消失和肾衰竭。我们发表的和初步的数据显示NLRP 3的激活 足细胞中的炎性小体。此外，NLRP 3激活发生在Tfh细胞亚群中，这些Tfh细胞是有效的 来激活B细胞。除了抑制足细胞中的NLRP 3活化外，特异性激酶MCC 950也可抑制足细胞中的NLRP 3活化。 NLRP 3抑制剂抑制自身抗体产生和生发中心（GC）形成。我们建议审问 足细胞和Tfh细胞中NLRP 3活化在狼疮性肾炎发病机制中的不同作用。三 提出了具体目标：具体目标1：利用具有CD 4特异性基因缺失的突变小鼠， NLRP 3活化途径，以确定CD 4 T细胞中这些基因的失活对Tfh细胞的影响 B6背景下小鼠的发育、GC形成、Ab产生。将进行类似的实验 在NZM 2328背景下的小鼠中，增加了自身抗体产生试验和 LN;具体目的2：确定足细胞缺失NLRP 3活化途径中基因的影响 在自身抗体缺乏的情况下，严重蛋白尿和早期死亡率的发展;具体目标3： 为了证明狼疮株在胱天蛋白酶-1激活的Tfh细胞中固有地升高， 具体目标4：探究MCC 950抑制NLRP 3的机制 活化、Tfh细胞活化、GC形成和自身抗体产生以及严重蛋白尿的减轻 在肾毒血清（NTS）诱导的肾炎中，IFNγ可加速LN和自发性LN的发生。的结果予以 拟议的实验将提供令人信服的数据，以支持靶向NLRP 3作为辅助治疗 LN的治疗方法他们将提供关于NLRPs激活和IL-1β在 和/或IL-18在Tfh细胞活化和终末器官损伤中的作用。因此，该建议具有重要意义 与我们对免疫反应和转化潜能的理解有关的基础科学。

Abstract Systemic lupus erythematosus (SLE) is a multi-organ systemic autoimmune disorder affecting mainly females in their reproductive ages. Patients with proliferative glomerulonephritis (GN) often progress to end stages of renal disease (ESRD). No new therapy for LN has been approved by the FDA for more than 50 years. A better understanding of the pathogenesis of LN is needed to devise more effective therapeutic strategies. Our laboratories have shown that immune complexes (IC) by themselves are not sufficient to cause ESRD, and end organ (kidney) resistance to damage is important in determining the clinical outcome of LN. Three GN models have been established. With a novel technique, we have identified in chronic glomerulonephritis that unique CD11b+IA-F4/80- infiltrating intraglomerular macrophages are the dominant cells that make TNFα, podocytes are the dominant cells that made IL-1β/IL-18 and mesangial cells make IL-6. A novel hypothesis is proposed that TNFα, IL-1β, IL-18 and IL-6 are made by different cells (compartments) in the diseased glomeruli and that the interactions among these cytokines with their receptors fuel the on-going inflammation within glomeruli leading to podocyte effacement and renal failure. Our published and preliminary data show activation of NLRP3 inflammasome in podocytes. In addition, NLRP3 activation occurs in a subpopulation of Tfh cells that are potent to activate B cells. In addition to the inhibition of NLRP3 activation in podocytes, MCC950, the specific kinase inhibitor of NLRP3 inhibits auto-Ab production and germinal center (GC) formation. We propose to interrogate the differential roles of NLRP3 activation in podocytes and Tfh cells in the pathogenesis of lupus GN. Three specific aims are proposed: Specific Aim 1: To utilize mutant mice with CD4 specific deletion of genes in the NLRP3 activation pathway to determine the effect of inactivation of these gene in CD4 T cells on Tfh cell development, GC formation, Ab-production in mice on B6 background. Similar experiments will be carried out in mice on NZM2328 background with the addition of assays for auto-Ab production and the development of LN; Specific Aim 2: To determine the effects of podocyte deletion of genes in the NLRP3 activation pathway on the development of severe proteinuria and early mortality in absence of auto-Ab reduction; Specific Aim 3: To demonstrate that lupus strains are intrinsically elevated in caspase-1 activated Tfh cells and hyper-responsive to immunization and Specific Aim 4: To interrogate the mechanism by which MCC950 inhibits NLRP3 activation, Tfh cell activation, GC formation and auto-Ab production and the attenuation of severe proteinuria in nephrotoxic serum (NTS) induced nephritis, IFN𝛼 accelerated LN and spontaneous LN. The results of these proposed experiments will provide convincing data to support targeting NLRP3 as an adjunct therapeutic approach in treating LN. They will provide definitive data regarding the roles of NLRPs activation and IL-1β and/or IL-18 in Tfh cell activation and end organ damage in autoimmunity. Thus the proposal has significance in basic science relating to our understanding of immunological responses and translational potentials.