The role of Sphingosine Kinase 1 in a Mouse Model of Chronic Inflammation

描述（由申请人提供）：项目概述：类风湿性关节炎（RA）是一种影响全世界人民的自身免疫性疾病。其潜在机制尚不清楚，但患者的关节中往往有高水平的肿瘤坏死因子（TNF）。TNF有助于在压力下保护身体，但它也可以通过引起炎症导致有害蛋白质的产生和非炎症区域的破坏来破坏未感染的细胞和周围组织。尽管医学取得了进步，但目前可用的抗炎疗法并不完全有效，并且通常昂贵，因此需要新的疗法。鞘脂存在于细胞膜中，但也被证明参与促进炎症的细胞途径。一种特定的鞘脂，鞘氨醇-1-磷酸（S1 P），已在文献中显示可引起炎症介质的增加。添加S1 P导致前列腺素E2（PGE 2）和考克斯-2蛋白增加。TNF和S1 P同时刺激比单独S1 P更能增加PGE 2和考克斯-2。因此，假设S1 P形成减少导致炎症减少。S1 P是通过鞘氨醇激酶（SphK）1和2的作用形成的;然而，SphK 1是炎症途径中负责S1 P形成的主要酶。使用siRNA去除成纤维细胞中的SphK 1导致S1 P、考克斯-2和PGE 2形成的减少。在这些细胞中TNF刺激后观察到的效应随着SphK 1的消除而消除，表明TNF和SphK 1之间的关系。该项目的目标是研究SphK 1对过度表达人TNF的小鼠（如RA患者）炎症的影响，并且没有SphK 1的工作拷贝。在初步研究中，缺失SphK 1功能拷贝的小鼠关节炎评分降低，尽管TNF α以恒定水平表达。本研究的目的是：1）确定鞘氨醇激酶1在体内TNF α诱导的关节炎发病机制中的作用; 2）表征具有和不具有SphK 1功能拷贝的TNF α转基因小鼠关节中的基因表达; 3）使用FLS确定SphK 1去除影响炎症的机制。这些研究的主要方法将是利用TNF/SphK 1小鼠模型来确定SphK 1影响炎症的机制。相关性：该项目旨在确定TNF α和SphK 1在慢性炎症中的意义，可能导致发现慢性炎症性疾病（如RA）的新治疗靶点。

DESCRIPTION (provided by applicant): Project Summary: Rheumatoid Arthritis (RA) is an autoimmune disease that affects people world wide. The underlying mechanism is unknown, but patients tend to have high levels of tumor necrosis alpha (TNF) in their joints. TNF helps protect the body when under stress, but it can also be damaging to un-infected cells and the surrounding tissue by causing inflammation leading to production of harmful proteins and destruction of non-inflammed areas. Despite advances in medicine, anti-inflammatory therapies currently available are not totally effective and are often costly, thus, novel therapies are needed. Sphingolipids are found in the cell membrane but have also been shown to be involved in the cellular pathways that promote inflammation. One specific sphingolipid, sphingosine-1-phosphate (S1P), has been shown in the literature to cause an increase in inflammatory mediators. Addition of S1P leads to increased prostaglandin E2 (PGE2) and COX-2 protein. Simultaneous stimulation with TNF and S1P increases PGE2 and COX-2 more than S1P alone. Therefore, decreased formation of S1P has been hypothesized to lead to decreases in inflammation. S1P is formed through the action of sphingosine kinase (SphK) 1 and 2; however SphK1 is the major enzyme responsible for S1P formation in the inflammatory pathway. Using siRNA to remove of SphK1 in fibroblast cells lead to decreases in S1P, COX-2, and PGE2 formation. The effects seen after TNF stimulation in these cells were removed with the elimination of SphK1 suggesting a relationship between TNF and SphK1. The goal of this project is to study the effects of SphK1 on inflammation in mice that over express human TNF, like RA patients, and do not have a working copy of the SphK1. In preliminary investigations, mice that are missing functional copies of SphK1 have decreased arthritis score despite the fact that TNF alpha is being expressed at constant levels. The aims of the study are 1) determine the contribution of sphingosine kinase 1 in the pathogenesis of TNF alpha induced arthritis in vivo; 2) characterize gene expression in joints from TNF alpha transgenic mice with and without functioning copies of SphK1; 3) determine the mechanism in which removal of SphK1 affects inflammation using FLS. The primary methodology of these studies will be to utilize the TNF/SphK1 mouse model to determine the mechanism of SphK1 affects inflammation. Relevance: This project seeks to define the significance of TNF alpha and SphK1 in chronic inflammation, potentially leading to the discovery of a novel therapeutic target for chronic inflammatory diseases like RA.