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 包括骨内稳态的过程。然而，广泛的体外和体内研究表明， 评估NO在骨生物学中的作用常常产生相反的结果。这至少部分是 由于一氧化氮合酶（NOS）的药理学抑制或 NOS缺陷受NOS同种型冗余的限制，不能解决细胞- 精氨酸琥珀酸裂解酶（ASL）是一种尿素循环酶，不仅是必需的 用于NOS底物精氨酸的从头合成，但也用于维持结构 完整的NO合成复合物含有NOS，乙酰氨基琥珀酸合酶（ASS 1），精氨酸 CAT-1和HSP 90。ASL的丢失导致非冗余和小区自主的 一氧化氮合酶依赖的一氧化氮生成和ASL缺乏症（ASLD）是一种人类NO遗传性疾病 生产 这项提案的总体目标是研究NO在骨转换、密度和骨代谢中的作用。 一氧化氮缺乏症的人体模型的结构，并了解其机制基础， NO影响骨代谢。通过利用这种罕见遗传疾病的正在进行的试验，我们将 解决这些具体问题：1）ASLD患者是否存在骨转换异常， 非NOS依赖性NO补充是否影响这些终点？2)耐心点- 诱导多能干细胞（iPSC）显示沿着成骨细胞的分化缺陷， 这是如何影响骨细胞分化的？3)从病人身上提取的成骨细胞- 由于小窝蛋白依赖性负调控占主导地位，iPSC表现出NO产生失调 调节NOS复合体？ 这些研究可能会对骨生物学的基本理解产生重大影响， 促进利用不依赖NOS的NO补充剂作为治疗剂的转化研究 对更常见的疾病如骨质疏松症进行干预。

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.