The immunological mechanism of PGRNs anti-inflammatory effect

肿瘤坏死因子-α TNFα）由于其位于肿瘤的顶端而受到最大的关注。 促炎细胞因子级联反应及其在炎症发病机制中的主导地位。抗tnf疗法 已被公认为治疗炎症性关节炎的有效方法。在绑定的全局屏幕中， 我们发现PGRN与TNFR 1和TNFR 2结合。后续 研究表明PGRN保护调节性T细胞（Treg）免受TNFα的负调节; PGRN信号传导的缺失使得B6小鼠对胶原诱导的关节炎（CIA）高度易感，而重组PGRN信号传导的缺失使得B6小鼠对胶原诱导的关节炎（CIA）高度易感。 PGRN预防CIA模型中的炎症性关节炎;并且在体内用PGRN刺激的 Treg细胞中的IL-10产生和IL-10信号传导对于体外PGRN刺激的Treg功能是重要的。 这些研究导致了中心假设，即PGRN通过其相互作用发挥其保护作用 与TNFR和刺激IL-10产生的Treg细胞在炎症性关节炎的过程中。 本研究的具体目的是：（1）探讨PGRN在原发性肝癌发病中的作用及其机制。 炎性关节炎我们将确定PGRN的诱导性敲除是否能减轻关节炎症状， 存在炎症性关节炎的小鼠。我们假设PGRN缺陷型中的易感表型 可能是由于Treg细胞的频率低。我们将研究PGRN的免疫调节作用 在CIA发病机制中的作用，重点关注PGRN的作用和Treg发育的潜在机制。 (2)阐明TNFR在炎症性关节炎发病机制中PGRN介导的信号传导中的作用。 我们将首先使用串联亲和纯化比较TNFα/TNFR和PGRN/TNFR的受体复合物， 随后进行质谱分析。我们还将比较PGRN和TNFα的关键细胞内事件， 包括人Treg细胞中的信号传导、靶基因表达谱和NFκB通路的激活 分离自正常人和炎性关节炎患者。此外，我们将阐明TNFR是 这对于通过使用TNFR 1-/-和TNFR 2-/-小鼠介导PGRN在CIA中的保护作用是重要的。(3)来定义 IL-10在炎症性关节炎发病机制中介导PGRN作用的作用。为了深入了解 为了研究已知的PGRN治疗机制，我们将使用IL-10-GFP报告基因老虎小鼠和Foxp 3 RFP 报告基因FIR小鼠，以鉴定IL-10表达非Treg细胞和响应于IL-10获得或丧失的Treg细胞。 PGRN在CIA中的作用，这将提供更多关于为什么PGRN缺陷的B6小鼠变得高度易感的见解。 以及IL-10是否是CIA中PGRN的靶点。为了检验PGRN通过IL-10介导细胞凋亡的假设， 观察对CIA的治疗效果，我们将确定是否过继转移PGRN诱导的IL-10表达 TMN可以抑制CIA，阻断IL-10信号通路是否抑制PGRN对CIA的作用。拟议 这些研究不仅将为CIA的发病机制提供新的见解， 炎症性关节炎和其他TNFR介导的病理和病症的新治疗策略。

Tumor necrosis factor-α TNFα) has received the greatest attention because of its position at the apex of the pro-inflammatory cytokine cascade, and its dominance in the pathogenesis of inflammation. Anti-TNF therapies have been accepted as the effective approach to treating inflammatory arthritis. In a global screen for the binding proteins of progranulin (PGRN) growth factor, we found that PGRN bound to TNFR1 and TNFR2. Subsequent studies demonstrated that PGRN protected regulatory T cells (Treg) from negative regulation by TNFα; Loss of PGRN signaling rendered B6 mice highly susceptible to collagen-induced arthritis (CIA), whereas recombinant PGRN prevented inflammation arthritis in CIA models; and in vivo treatment with PGRN stimulated IL-10 production in Treg cells and IL-10 signaling was important for PGRN-stimulated Treg function in vitro. These studies have led to the central hypothesis, that PGRN exerts its protective effects through its interaction with TNFR and stimulation of IL-10 producing Treg cells in the course of inflammatory arthritis. The Specific Aims are: (1) To determine the roles of PGRN and underlying mechanisms in the pathogenesis of inflammatory arthritis. We will determine whether inducible knockout of PGRN worsens arthritic symptoms in mice with existing inflammatory arthritis. We hypothesize that the susceptible phenotype seen in PGRN deficient mice may be due to the low frequency of Treg cells. We will study the immune regulatory roles of PGRN in the pathogenesis of CIA, focusing on the role of PGRN and the underlying mechanisms in Treg development. (2) To elucidate TNFR’s role in PGRN-mediated signaling in the pathogenesis of inflammatory arthritis. We will first compare the receptor complexes of TNFα/TNFR and PGRN/TNFR using tandem affinity purification, followed by mass spectrometry. We will also compare the key intracellular events of PGRN and TNFα, including signaling, target gene expression profiling, and activation of the NFκB pathway in human Treg cells isolated from normal and the patients with inflammatory arthritis. Furthermore, we will elucidate which TNFR is important for mediating PGRN’s protective role in CIA by using TNFR1-/- and TNFR2-/- mice. (3) To define the role of IL-10 in mediating PGRN action in the pathogenesis of inflammatory arthritis. To gain insight into the mechanisms of already known therapies of PGRN, we will use IL-10-GFP reporter tiger mice and Foxp3 RFP reporter FIR mice to identify the IL-10 expressing non-Treg cells and Treg cells in response to gain- or loss-of- PGRN in CIA, which would provide more insight on why PGRN-deficient B6 mice become highly susceptible to CIA, and whether IL-10 is the target of PGRN in CIA. To test the hypothesis that PGRN via IL-10 mediates the observed therapeutic effect upon CIA, we will determine if adoptive transfer of PGRN-induced IL-10-expressing Tregs can suppress CIA, and whether blockage of IL-10 signaling inhibits PGRN’s effects on CIA. The proposed studies will not only provide new insights into the pathogenesis of CIA, but also lead to the development of novel therapeutic strategies for inflammatory arthritis and other TNFR-mediated pathologies and conditions.