Nitric Oxide and Bone Homeostasis in Patients with Argininosuccinate Lyase Deficiency

项目摘要 一氧化氮(NO)是一种普遍存在的信号分子，对大多数生理学 包括骨骼动态平衡在内的各种过程。然而，广泛的体外和体内研究已经 评估一氧化氮在骨生物学中的作用往往会产生截然不同的结果。这至少是部分原因 由于一氧化氮合酶(NOS)的药物抑制或遗传模型 一氧化氮合酶缺乏症由一氧化氮合酶亚型的冗余所限制，不能解决细胞- NO的自主作用。精氨酸琥珀酸裂解酶(ASL)，是尿素循环中唯一必需的酶 对于精氨酸的从头合成，底物为一氧化氮合酶，也是维持结构的 含有精氨酸琥珀酸合成酶(ASS1)和精氨酸的NO合成复合体的完整性 转运蛋白CAT-1和HSP90。ASL的丢失导致非冗余和细胞自主的丢失 一氧化氮合酶依赖的一氧化氮的产生和ASL缺乏症(ASLD)是一种人类的NO遗传性疾病 制作。 这项建议的总体目标是研究一氧化氮在骨转换、密度和骨密度中的作用。 建筑在一个没有缺陷的人类模型中，并理解其机械基础 一氧化氮影响骨代谢。通过利用正在进行的这种罕见遗传疾病的试验，我们将 解决这些具体问题：1)ASLD患者是否存在骨转换异常和 骨量和不依赖一氧化氮合酶的NO补充是否会影响这些终点？2)患者- 来源的诱导多能干细胞(IPSC)沿成骨细胞方向显示分化缺陷 世系以及这对破骨细胞分化有何影响？3)患者的成骨细胞- IPSC表现出NO产生的失调，这是由于依赖小窝蛋白的负性基因占优势 一氧化氮合酶复合体？ 这些研究可能会对骨生物学的基本理解和 促进利用一氧化氮合酶非依赖性一氧化氮补充作为治疗手段的翻译研究 对骨质疏松症等更常见疾病的干预。

Project Summary Nitric oxide (NO), a ubiquitous signaling molecule, is important for most physiological processes including bone homeostasis. However, extensive in vitro and in vivo studies that have assessed the role of NO in bone biology have often yielded contrasting results. This is at least in part due to the fact that pharmacologic inhibition of nitric oxide synthases (NOS) or genetic models of NOS deficiency are limited by the redundancies of the NOS isoforms and cannot address the cell- autonomous roles of NO. Argininosuccinate lyase (ASL), is a urea cycle enzyme is not only required for the de novo synthesis of arginine, the substrate for NOS, but also to maintain the structural integrity of a NO-synthesis complex containing NOS, argininosuccinate synthase (ASS1), the arginine transporter CAT-1, and HSP90. Loss of ASL leads to non-redundant and cell-autonomous loss of NOS-dependent NO production and thus ASL deficiency (ASLD) is a human genetic disorder of NO production. The overall goals of this proposal are to study the role of NO in bone turnover, density, and architecture in a human model of NO deficiency and to understand the mechanistic basis by which NO affects bone metabolism. By leveraging an ongoing trial in this rare genetic disorder, we will address these specific questions: 1) Do patients with ASLD have abnormalities in bone turnover and bone mass and does NOS-independent NO supplementation affect these endpoints? 2) Do patient- derived induced pluripotent stem cells (iPSC) show differentiation defects along the osteoblastic lineage and how does this impact osteoclastic differentiation? 3) Do osteoblasts derived from patient- iPSC exhibit dysregulation of NO production due to dominance of a caveolin-dependent negative regulatory NOS complex? These studies could have a significant impact on the basic understanding of bone biology and foster translational studies in utilizing NOS-independent NO supplementation as a therapeutic intervention in the more common disorders like osteoporosis.