Project 1: Systems-based Analysis of Redox Activity in Aortic Valve Stenosis

项目摘要 氧化应激在纤维钙化性主动脉瓣狭窄(BESS)中起着关键但未知的作用。偏爱是一种疾病 在主动脉瓣(AV)，叶逐渐变硬并最终钙化，导致心脏功能障碍。 唯一的治疗方法是外科瓣膜置换术。氧化还原失衡导致的氧化应激增加 被认为是主动脉瓣(AV)叶终末钙化的启动因素，但处于氧化还原状态 期间收受的利益仍未定义。我们目前的数据显示，在非常年轻的人中，氧化应激增加 伴有细胞外基质重塑的成年小鼠主动脉瓣硬化模型，提示 氧化还原机制在BESS中的作用比之前描述的要早得多。我们很早就假设 氧化还原失衡与ECM紊乱有关，ECM紊乱导致瓣膜僵硬。描绘，描绘 氧化还原机制，目标1将定义大宗、整体和组织横截面的氧化还原状态 主动脉瓣顺序性硬化早期房室瓣叶的测量。生理学测量 心功能和房室的生物力学特性将被用来定义房室僵硬的水平 氧化还原状态。AIM 2将利用脱细胞瓣叶提取物的蛋白质组学技术来鉴定 氧化还原敏感的细胞通路和蛋白相互作用网络(PIN)在顺序的瓣膜硬化过程中。 生物信息学方法将用于评估在瓣膜硬化过程中调节的PIN，专门用于新的 最受欢迎的候选人。脱细胞瓣膜结构中的细胞外基质(ECM)将由 蛋白质组学和免疫印迹以报告与氧化应激增加相关的细胞外基质变化。 生物信息学工具和免疫印迹技术将用于报告氧化应激诱导的后遗症 翻译修饰语。目标3将描述蛋白质相互作用网络中的调控 未鉴定的正常和良性临床标本中氧化应激增加。一种组合 免疫组织化学、免疫印迹和蛋白质组学将用于确定正常AV的氧化还原状态 在细胞水平上与BESS进行比较，并评估对瓣膜结构表型的总体贡献。候选人 从早期瓣膜僵硬的小鼠模型将评估在临床标本中对BESS的贡献。 这项调查依赖于南卡罗来纳州医科大学科布雷在#年的独特资源 氧化剂、氧化还原平衡和胁迫信号，如果不使用这些是不可能实现的 资源。这项研究将使我们能够确定氧化还原平衡的作用和受影响的蛋白质表达在 瓣膜僵硬的早期阶段，识别可能抑制疾病的新治疗靶点 进步，提高生活质量，降低死亡率。

Project Summary Oxidative stress plays a key but unknown role in fibrocalcific aortic valve stenosis (FAVS). FAVS is a disease of aortic valve (AV), where leaflets progressively stiffen and eventually calcify, causing cardiac malfunction. The only treatment is surgical valve replacement. Increased oxidative stress due to redox imbalance is believed to be an initiating factor for end point calcification of the aortic valve (AV) leaflets, but redox status during FAVS remains undefined. Our current data shows that oxidative stress is increased in a very young adult mouse model of aortic valve stiffening, concomitant with extracellular matrix remodeling, implicating that redox mechanisms play a much earlier role in FAVS than previously described. We hypothesize that early redox imbalances are associated with the ECM disorganization that results in valvular stiffening. To delineate redox mechanisms, Aim 1 will define redox status in bulk, whole mount and histological cross sectional measurements of the AV leaflet during early sequential aortic valve stiffening. Physiological measurements of heart function and biomechanical properties of the AV will be used to define levels of AV stiffening parallel with redox status. Aim 2 will utilize proteomic techniques on extracts from decellularized valve leaflets to identify redox sensitive cellular pathways and protein interaction networks (PINs) during sequential valve stiffening. Bioinformatic approaches will be used to assess PINs regulated during valve stiffening, earmarking new candidates of FAVs. Extracellular matrix (ECM) from the decellularized valve structure will be interrogated by proteomics and immunoblotting to report on ECM changes correlating to increased oxidative stress. Bioinformatics tools and immunoblotting techniques will be used to report on oxidative stress induced post translational modifications. Aim 3 will delineate regulation in protein interaction networks coinciding with increased oxidative stress in de-identified normal and FAVS clinical specimens. A combination of immunohistochemistry, immunoblotting, and proteomics will be used to define the redox status of normal AV compared to FAVS at a cellular level and assess overall contribution to valve structure phenotype. Candidates from the murine model of early valve stiffening will be evaluated for contribution to FAVS in clinical specimens. This investigation relies on the unique resources of the Medical University of South Carolina's COBRE in Oxidants, Redox Balance and Stress Signaling and would not be accomplished without use of these resources. This study will allow us to define the role of redox balance and affected protein expression during the early stages of valve stiffening in FAVS, identifying new therapeutic targets that might inhibit disease progression, improve quality of life and decrease mortality.