Metabolic Studies in Disorders of Branch Chain Amino Acid Metabolism

描述（由申请人提供）：本研究的总体目标是研究 IVA、MMA 和 PA 中间代谢紊乱所涉及的分子和代谢机制。越来越多的证据表明线粒体功能障碍在这些疾病的病理学中发挥着重要作用。由于这些疾病的标准治疗是通过饮食和辅因子补充来纠正中间代谢的缺陷，因此我们认为更好地了解这些线粒体紊乱将有助于改进治疗。我们对 IVA、MMA 和 PA 患者血浆中柠檬酸循环中间体的新评估揭示了独特的疾病特异性代谢物模式。我们的目标是优化我们的代谢组学方法并引入新的基因组学方法，以进一步定义异常线粒体代谢所涉及的代谢和分子途径。我们将应用质谱 (MS) 技术来创建 MMA、PA 和 IVA 患者血浆样本的“非靶向”定性代谢组学谱。然后，我们将使用包括稳定同位素稀释 MS 在内的靶向技术来量化最感兴趣的代谢物。我们期望这些代谢物谱将有助于定义这些疾病的病理生理学基础的生物化学的复杂畸变。这些代谢物谱也有可能帮助识别治疗干预的新靶点。接下来，我们将使用非靶向和靶向代谢组学方法来检查病人和健康患者中间代谢的差异。这些差异将有助于 了解导致急性疾病的生物化学变化，以及确定并发疾病期间治疗干预的新目标。 然后，我们将通过血浆特异性代谢谱来探索 MMA 小鼠模型中能量代谢受损的潜在生化基础。 MMA 小鼠血浆中的代谢谱将揭示物种特异性对疾病的生化反应的相似性和差异。在研究的后期阶段，我们将使用 mRNA-seq 转录组分析来确定受 MMA 影响的小鼠肝组织中中间线粒体代谢的分子介质。这些研究将补充我们的代谢组学工作，并且了解这些基因编码的蛋白质的功能将为了解生物化学紊乱的机制提供见解。

DESCRIPTION (provided by applicant): The overall objective of this research is to investigate the molecular and metabolic mechanisms involved in disturbed intermediary metabolism in IVA, MMA, and PA. There is mounting evidence that mitochondrial dysfunction plays a role in the pathology of these disorders. Since standard treatment for these disorders is to corect the deficiencies in intermediary metabolism through dietary and cofactor supplementation, and we submit that a better understanding of these mitochondrial disturbances will allow for improved treatments. Our novel assessment of citric acid cycle intermediates in plasma from patients with IVA, MMA, and PA has revealed distinct and disease specific patterns of metabolites. Our goal is to optimize our metabolomic approaches and introduce new genomic approaches in order to further define the metabolic and molecular pathways involved in aberrant mitochondrial metabolism We wil apply mass spectrometry (MS) techniques to create "non-targeted" qualitative metabolomic profiles of plasma samples from patients with MMA, PA and IVA. We will then use targeted techniques including stable isotope dilution MS to quantify metabolites of greatest interest. We expect that these metabolite profiles will help to define the complex aberrations in biochemistry underlying the pathophysiology in these diseases. These metabolite profiles also have the potential to aid in the identification of novel targets for therapeutic intervention. We will next use non-targeted and targeted metabolomic approaches to examine the differences in intermediary metabolism in sick and well patients. These differences will aid in the understanding of the changes in biochemistry leading to acute illness, as wel as the identification of novel targets for therapeutic intervention during intercurrent illness. We then ill explore the underlying biochemical basis of impaired energy metabolism in mouse models of MMA via plasma specific metabolic profiling. Metabolic profiling in the plasma of mice with MMA will reveal similarities and differences in species specific biochemical response to disease. In the later phase of investigation we will determine the molecular mediators of intermediary mitochondrial metabolism in liver tissue derived from mice affected with MMA using mRNA-seq transcriptome analysis. These studies will complement our metabolomic work, and understanding of the functions of proteins encoded by these genes will provide insights into the mechanisms underlying disturbed biochemistry.