Project 3: Anti-Inflammatory Mechanisms of Inhaled Hypertonic Saline

创伤、出血和血液复苏通过诱导趋化因子、细胞因子和黏附蛋白而产生急性肺炎症。上调机制依赖于将基因调节、转录因子(TF)等转录因子(TF)转移到细胞核中的系统信号。这些基因直接表达白细胞的黏附、迁移和肺泡环境中的表型发育。高渗(HOsm)似乎可以阻止炎症基因的表达，但其机制和范围尚不清楚。在创伤复苏期间输注治疗性高渗盐水，或像治疗囊性纤维化那样吸入高渗盐水，是很好的耐受性。我们假设，高渗透压通过促进新的复合体来改变炎症刺激诱导的特定转录因子的核移位。在这个建议期(2005-2010)，我们将首先研究控制被HOsm修饰的典型肺泡细胞因子和黏附蛋白的TF的流量，并研究这些TF的复合体，包括那些与Importins形成的复合体。 1.用IP-MS蛋白质组学方法研究HOsm对核因子-KB和IKK家族成员易位的影响及其对细胞质复合体的影响，并用IP-MS蛋白质组学方法检测HOsm促进的交替复合体。 2.由于HOsm不抑制IL-1诱导的p65-Rel A易位，但抑制某些细胞因子，我们将研究IRF的其他重要启动子C/EBP的易位。和R/Flat家庭，并确定他们的HOsm改变的伴侣。 接下来，为了将调查扩展到已知的细胞因子之外，我们将执行以下操作： 3.基因阵列分析，以指导潜在TF的生物信息学查询。目标是逐步建立在HOSm期间未能成功移位的转录因子的文库，并确定它们的隔离伙伴(从NF-kB和IRF家族成员开始)。 最后，我们将转换这些台架实验，以评估雾化吸入HOsm在休克动物和创伤患者中的耐受性或益处。 4.观察创伤动物和吸入HOsm患者的肺部炎症反应和组织因子转位。来自AIMS 1-3的信息将用于解释治疗后的分子表型和Tf的再分布。

Trauma, hemorrhage and blood resuscitation produce acute lung inflammation by inducing chemokine, cytokine and, and adhesive proteins. Up-regulating mechanisms depend on systemic signals that translocate gene-regulating, transcription factors (TFs), such as NF-kB into the nucleus. The genes expressed direct the adhesion, transmigration of leukocytes and phenotypic developments in the alveolar milieu. Hyperosmolarity (HOsm), appears to prevent inflammatory gene expression but the mechanisms and scope are unclear. Therapeutic hypertonic saline infusions during trauma-resuscitation, or inhaled as for cystic fibrosis, are well tolerated. We hypothesize that hyperosmolarity alters the nuclear translocation of selected transcription factors induced by inflammatory stimuli by promoting novel complexes. In this proposal period (2005-2010) we will first examine the traffic of TFs controlling prototypical alveolar cytokines and adhesive proteins that are modified by HOsm, and investigate complexes of these TFs, including those formed with importins. 1. Investigate HOsm altered translocation of the NF-KB and IKK family members, their HOsm altered cytoplasmic complexes and examine alternate complexes promoted by HOsm with IP-MS proteomics. 2. Since HOsm does not prohibit p65 Rel A translocation induced by IL-1 yet suppresses certain cytokines, we will examine translocation of other essential promoters of the IRF, C/EBP. and R/FLAT families and identify their HOsm altered partners. Next, to expand the inquiry beyond the known cytokines we will execute: 3. Gene array analyses, to guide bioinformatics queries for potential TFs. The goal is to gradually build up a library of TFs that do not translocate successfully during HOSm and identify their sequestering partners, (starting with NF-kB and IRF family members). Lastly, we will translate these bench experiments to asses the tolerability or benefit of nebulized HOsm in shocked animals and trauma patients. 4. Evaluate lung inflammation and TF translocation in traumatized Animal or Patients treated with inhaled HOsm. The information from Aims 1-3 will be used to interpret the molecular phenotypes and TF redistributions after treatments.