CORE--PROTEOMICS

核心E的主要目标是鉴定新的生物标志物，旨在表征肺肿瘤的特征。 基于肺动脉高压（PAH）状态的严重肺动脉高压（PAH）血管和右心室重构 蛋白质组学技术核心E和核心B-D支持本SCCOR的项目1、3、4和5， 基于四个战略的基本实验服务。首先，核心，使用广泛的蛋白质组学 分析，将表征肺和右心室肌细胞蛋白质组的变化， 实验和人类PAH的开发（项目1、3、4和5以及核心C和D）。这 该战略将得到现有蛋白质组学基础设施和JHU内已建立的专业知识的支持， NHLBI蛋白质组学中心和JHU Bayview蛋白质组学中心。深入分析，使用多个 蛋白质组学平台，包括二维差异成像电泳，二维液相色谱， 色谱法，O 16/O 18同位素标记，相对和绝对的同量异位素标签和定量 （iTRAQTM）、MALDI-TOF和液相色谱/质谱将用于最大化蛋白质组 覆盖率，同时允许表征蛋白质亚型和翻译后修饰。第二、 核心将提供一个集中设施，用于分析、汇编和关联候选人的数量水平 生物标志物，包括发现平台产生的新标志物以及其他潜在的生物标志物。 硬皮病相关PAH的相关候选生物标志物（如VEGF、HIMF、MMP-9和颗粒酶B）。 Beckman A2多重免疫测定将用于评价血清样本，而化学InkJet 打印机（CHIP，Proteome Systems）将用于我们的靶向方法以确认表达差异 从使用肺和右心室组织的基因组研究和蛋白质组发现研究推断 样品（项目1和3，以及核心C和D）。第三，核心将侧重于解开以下因素的相互作用组： 新的趋化因子，HIMF（项目4），使用多种蛋白质组学策略进行最大检测。 免疫沉淀后进行1D凝胶电泳或iTRAQTM标记和质谱分析， 用于识别结合伙伴。由于磷酸化在细胞因子信号传导中的突出作用， 固定化金属亲和色谱与液相色谱/质谱联用， 用于富集和鉴定各种结合配偶体的特异性磷酸化氨基酸残基。 最后，核心将进行计算建模，以开发PAH对 基于蛋白质组学、生理学和基因组学数据的右心室心肌细胞（项目2， 4和5，以及核心C和D）。这项工作将在心血管生物信息学中心进行 导演：Raimond L.温斯洛博士

The major goals of Core E are to identify novel biomarkers aimed at the characterization of pulmonary vascular and right ventricle remodeling in severe pulmonary arterial hypertension (PAH) based on state of the art proteomic technology. Core E, with Cores B-D,supports Projects 1,3,4, and 5 of this SCCOR with essential experimental services based on four strategies. First, the Core, using broad-based proteomic analysis, will characterize the changes in lung and right ventricular myocyte proteomes that occur during the development of experimental and human PAH (Projects 1, 3, 4 and 5 and Cores C and D). This strategy will be supported by the existing proteomic infrastructure and established expertise within the JHU- NHLBI Proteomics Center and the JHU Bayview Proteomics Center. In-depth analysis using multiple proteomic platforms, including two-dimensional differential imaging electrophoresis, two-dimensional liquid chromatography, O16/O18 isotopic labeling, isobaric tag for relative and absolute and quantification (iTRAQTM), MALDI-TOF and liquid chromatography/mass spectrometry will be used to maximize proteome coverage, while allowing characterization of protein isoforms and post-translational modifications. Second, the Core will provide a centralized facility to analyze, compile and correlate quantitative levels of candidate biomarkers, including novel markers arising from the discovery platforms as well as additional potentially relevant candidate biomarkers (e.g.VEGF, HIMF, MMP-9 and Granzyme B) in scleroderma-relatedPAH. Beckman A2 multiplex immunoassay will be used to evaluate serum samples while the chemical InkJet printer (CHIP, Proteome Systems) will be used for our targeted approach to confirm expression differences deduced from genomic studies and proteomic discovery studies using lung and right ventricle tissue samples (Projects 1 and 3, and Cores C and D). Third, the Core will focus on unraveling the interactome of the novel chemokine, HIMF (Project 4), using multiple proteomic strategies for maximal detection. Immunoprecipitation followed by 1D gel electrophoresis or iTRAQTM labeling and mass spectrometry will be used to identify binding partners. Due to the prominent role of phosphorylation in cytokine signaling, immobilized metal affinity chromatography online with liquid chromatography/mass spectrometry will be used to enrich and identify the specific phosphorylated amino acid residues of the various binding partners. Finally, the Core will perform computational modeling to develop a working cellular model of PAH effects on the right ventricular cardiac myocyte based on the proteomic, physiological and genomics data (Projects 2, 4 and 5, and Cores C and D). This work will be performed at the Center for Cardiovascular Bioinformatics and Modeling, directed by Raimond L. Winslow, PhD.