Novel Functions of S1P in Neuroinflammation

描述（由申请人提供）：白细胞介素1 （IL-1）是一种促炎细胞因子，对人类多发性硬化症（MS）和小鼠实验性自身免疫性脑脊髓炎（EAE）的发生和进展至关重要。尽管IL-1通过激活核因子κB诱导许多细胞因子的表达，但诱导几种对MS和EAE重要的趋化因子，如RANTES和IP-10，需要激活另一个转录因子，干扰素调节因子-1 （IRF-1）。值得注意的是，IRF-1在MS患者的大脑中被激活，IRF-1敲除小鼠免受EAE的影响，EAE受IRF-1在中枢神经系统（CNS）中表达的调控。我们已经证明IL-1刺激并上调鞘氨醇激酶1 (SphK1)， SphK1是生成鞘氨醇-1-磷酸（S1P）的两种同工酶之一，S1P是一种调节多种对炎症重要的细胞过程的脂质介质。MS患者脑脊液中S1P升高，诱导星形胶质细胞增生。FTY720是一种被SphK2磷酸化为S1P类似物（FTY720- p）的前药，分泌后靶向S1P 5种受体中的4种（除S1P2外），目前处于III期临床试验，可有效降低MS的频率和严重程度。FTY720减轻MS症状的部分原因是通过下调淋巴细胞上的S1P1，从而阻止其依赖S1P的淋巴细胞从淋巴结排出。然而，FTY720也在大脑中积累，在那里它可以减少星形胶质细胞增生；然而，其对中枢神经系统的影响尚不完全清楚。我们得到了两个有趣的观察结果：1)S1P直接结合细胞内的细胞凋亡抑制剂2 (cIAP2)，刺激cIAP2介导的K63调节IRF-1的泛素化及其激活。2)与细胞内S1P不同，细胞外S1P通过S1P2抑制星形胶质细胞中IRF-1的核易位，并诱导RANTES和IP-10的表达。因此，我们假设“S1P通过调节IRF-1功能的新的细胞内和细胞外机制调节脑炎症”。这一假设对于理解S1P、FTY720和IRFs在星形胶质细胞介导的神经炎症和ms中的作用机制具有重要意义。我们提出以下具体目标：分析细胞内S1P作为ciap2介导的IRF-1泛素化的辅助因子的作用。目标2。确定细胞外S1P影响IRF-1激活的机制。目标3。明确体内S1P调控IRF-1的作用。这一建议将加强对星形胶质细胞中IL-1， S1P及其受体之间复杂的相互作用如何调节炎症的理解，并为设计更有效的MS治疗方法铺平道路。

DESCRIPTION (provided by applicant): Interleukin 1 (IL-1) is a proinflammatory cytokine that is critical for the development and progression of multiple sclerosis (MS) in humans and experimental autoimmune encephalomyelitis (EAE) in mice. Although IL-1 induces expression of many cytokines via the activation of nuclear factor κB, induction of several chemokines important for MS and EAE, such as RANTES and IP-10, requires the activation of an additional transcription factor, interferon regulatory factor-1 (IRF-1). Significantly, IRF-1 is activated in the brains of MS patients, IRF-1 knockout mice are protected from EAE, and EAE is regulated by IRF-1 expressed in the central nervous system (CNS). We have shown that IL-1 stimulates and upregulates sphingosine kinase 1 (SphK1), one of the two isoenzymes that generate sphingosine-1-phosphate (S1P), a lipid mediator that regulates diverse cellular processes important for inflammation. S1P is increased in the CSF of MS patients and induces astrogliosis. FTY720, a pro-drug that is phosphorylated by SphK2 to an S1P mimetic (FTY720-P) and after secretion targets four of the five S1P receptors, except S1P2, is currently in phase III clinical trials, and effectively reduces the frequency and severity of MS. FTY720 reduces MS symptoms, in part, by downregulating S1P1 on lymphocytes thus preventing their S1P-dependent egress from the lymph nodes. However, FTY720 also accumulates in the brain where it reduces astrogliosis; nevertheless, its effects in the CNS are incompletely understood. We made two intriguing observations: 1) S1P directly binds to cellular inhibitor of apoptosis 2 (cIAP2) inside cells and stimulates cIAP2-mediated K63 regulatory ubiquitination of IRF-1 and its activation. 2) In contrast to intracellular S1P, extracellular S1P suppresses nuclear translocation of IRF-1 and induction of RANTES and IP-10 expression via S1P2 in astrocytes. Thus, we hypothesize that "S1P modulates brain inflammation by novel intracellular and extracellular mechanisms that regulate IRF-1 functions". This hypothesis has significant implications for understanding mechanisms of S1P, FTY720, and IRFs actions in astrocyte-mediated neuroinflammation and MS. We propose the following specific aims: Aim 1. Analyze the role of intracellular S1P as a cofactor for cIAP2-mediated ubiquitination of IRF-1. Aim 2. Identify the mechanisms by which extracellular S1P affects IRF-1 activation. Aim 3. Define the roles of S1P regulation of IRF-1 in vivo. This proposal will enhance understanding of how the intricate interplay between IL-1, S1P and its receptors in astrocytes regulates inflammation and could pave the way for the design of even more effective MS therapies.