GRANULOMA REGULATION THROUGH SOMATOSTATIN RECEPTOR SSTR2

肉芽肿性炎症是许多疾病的特征， 克罗恩病、结节病和原发性胆汁性肝硬化。肉芽肿 是炎性细胞的集中聚集体，受T 细胞反过来，其他类型的炎症细胞调节肉芽肿T 细胞当这种交叉被破坏时， 调节电路这种调节回路的大部分通过 神经因子和其他淋巴因子的释放。 生长抑素（SOM）是一种 这类神经因子改变淋巴细胞的反应。肉芽肿巨噬细胞 分泌SOM。小鼠肉芽肿T细胞显示SOM受体一样， 在人类肉芽肿中。克隆了五种不同的SOM受体（SSTR）， 测序和鉴定。我们目前的证据表明，SSTR 2 是肉芽肿T细胞上表达的唯一SOM受体。此外，本发明还提供了一种方法， 肉芽肿细胞表达SSTR 2的两种同种型。SOM抑制干扰素- γ-干扰素（IFN-γ）从肉芽肿T细胞释放。SOM可能被证明是一个 IFN-γ是慢性炎症的重要调节因子，因为IFN-γ是 调节多种炎症细胞类型的细胞因子。因此 为了进一步阐明SSTR 2在炎症中的重要性，该项目 我将继续研究这四个假设：（1）。SOM调节IFN-γ释放 通过与SSTR 2相互作用从肉芽肿T细胞中分离，2.）SSTR2 表达受T细胞活化的调节，3）表达的两种同种型 SSTR 2抑制IFN-γ释放的能力不同，4）SOM对照 通过与SSTR 2的相互作用在体内引起肉芽肿反应。鼠 血吸虫病提供了一个定义明确的自然模型系统来研究 肠道和肝脏的肉芽肿小鼠肉芽肿 血吸虫表达SOM、SSTR 2等方面的SOM回路。 因此，我们将使用从这些肉芽肿中分离的细胞来实现 目的1：确定SOM是否直接与T细胞相互作用 SSTR 2调节IFN-γ释放而不介入参与 使用受体阻断抗体，特异性激动剂， 流式细胞术和具有废除的SSTR 2展示的克隆T细胞。目标二： 确定T细胞活化是否影响SSTR 2 mRNA和蛋白质合成， 和SSTR 2磷酸化，并使用定量RT-PCR， 分析、流式细胞术、蛋白质印迹和免疫沉淀。目的 3：确定SSTR 2的两种亚型是否受到差异调节 并具有不同的抑制T细胞IFN-γ释放的能力 同种型特异性定量RT-PCR和免疫沉淀，和 同种型的选择性转染。目的4：确定是否为病灶内 SOM与SSTR 2相互作用以使用SSTR 2调节体内肉芽肿 阻断抗体与选择性受损T细胞的转基因小鼠 SSTR 2显示。我们希望这些发现能带来新的 治疗IBD。

Granulomatous inflammation is characteristic of many diseases like Crohn's disease, sarcoidosis and primary biliary cirrhosis. Granulomas are focal collections of inflammatory cells, tightly regulated by T cells. In turn, other inflammatory cells types regulate granuloma T cells. Diseases like IBD develop when there is disruption of this cross regulatory circuitry. Much of this regulatory circuitry proceeds through release of neurokines and other lymphokines. Somatostatin (SOM) is one such neurokine that alters lymphocyte responses. Granuloma macrophages secrete SOM. Murine granuloma T cells display SOM receptors as do cells in human granulomas. Five distinct SOM receptors (SSTR) are cloned, sequenced, and characterized. Our current evidence indicates that SSTR2 is the only SOM receptor expressed on granuloma T cells. In addition, granuloma cells express both isoforms of SSTR2. SOM inhibits interferon- gamma (IFN-gamma) release from granuloma T cells. SOM may prove to be an important regulator of chronic inflammation, since IFN-gamma is a pivotal cytokine that regulates many inflammatory cell types. Therefore, to further elucidate the importance of SSTR2 in inflammation, this project will pursue these four hypotheses: 1.) SOM regulates IFN-gamma release from granuloma T cells through interaction with SSTR2, 2.) SSTR2 expression is regulated by T cell activation, 3) The two isoforms of SSTR2 differ in ability to inhibit IFN-gamma release, 4) SOM controls granulomatous responses in vivo by interaction with SSTR2. Murine schistosomiasis provides a well defined natural model system to study granulomas in the intestines and liver. Granulomas in murine schistosomiasis express SOM, SSTR2 and other aspects of the SOM circuit. Thus, we will use cells isolated from these granulomas to achieve the following aims: Aim 1: Determine if SOM interacts directly with T cell SSTR2 to regulate IFN-gamma release without the interposed participation of another cell type using receptor blocking antibody, specific agonists, flow cytometry and cloned T cells with abrogated SSTR2 display. Aim 2: Determine if T cell activation affects SSTR2 mRNA and protein synthesis, and SSTR2 phosphorylation, and using quantitative RT-PCR, nuclear run-off analysis, flow cytometry, western blotting and immunoprecipitation. Aim 3: Determine if the two isoforms of SSTR2 are differentially regulated and have different capacity to inhibit T cell IFN-gamma release using isoform specific quantitative RT-PCR and immunoprecipitation, and selective transfection of the isoforms. Aim 4: Determine if intralesional SOM interacts with SSTR2 to regulate granulomas in vivo using SSTR2 blocking antibodies and transgenic mice with selectively impaired T cell SSTR2 display. Our hope is that these discoveries will lead to new therapies for IBD.