Nitric oxide regulation of glycolysis in osteoblasts

项目摘要/摘要--项目3 一氧化氮(NO)是一种重要的信号分子，影响着广泛的生物过程。 包括骨骼代谢。最近，我们利用精氨酸琥珀酸裂解酶缺乏症(ASLD)作为模型来研究 细胞自主的一氧化氮合酶(NOS)依赖的NO缺乏症。精氨酸琥珀酸裂解酶(ASL)是 只有哺乳动物的酶能够合成精氨酸，精氨酸是一氧化氮合酶依赖的一氧化氮合成的唯一前体。 此外，ASL还需要将细胞外精氨酸输送到一氧化氮合酶以产生NO。使用ASLD作为 在一个模型中，我们报告了NO促进骨形成与刺激糖酵解有关 成骨细胞分化。越来越多的证据表明，成骨细胞通过 糖酵解是它们的主要能量来源。重要的是，一些用于增加骨量的合成代谢剂， 例如Wnt蛋白也被证明在成骨细胞分化过程中刺激糖酵解，因此 提示糖酵解在骨合成代谢中起着重要作用。该项目的总体目标是确定 一氧化氮在生理条件下和机械负荷时刺激骨形成的机制。 我们还将检查是否没有糖酵解影响对骨量的影响。我们报道了 NO缺乏的细胞下调了编码糖酵解途径组成部分的mRNAs，这表明 NO可能调节转录因子，这些转录因子调节糖酵解途径中基因的表达。 此外，NO诱导的糖酵解刺激可能参与了负荷诱导的骨形成，这是一种 大量的工作表明，NO是一个关键的调解人。这个项目的中心假设是没有 成骨细胞系细胞通过刺激促进成骨细胞分化和骨形成 糖酵解。我们将通过三个具体目标来检验这一假设。目标1将确定一氧化氮是否促进骨骼 通过激活cAMP反应元件结合蛋白(CREB)在成骨细胞中形成和糖酵解。目标2 将决定糖酵解的基因上调是否足以刺激骨形成或拯救 成骨细胞特异性As1基因敲除小鼠的骨量不足。目标3将确定是否从 成骨细胞和骨细胞是负荷诱导合成代谢所必需的。成功完成这个项目应该 深入了解一氧化氮对骨骼影响的分子机制。此外，它可能会 为骨合成代谢的治疗靶点提供新的见解。

PROJECT SUMMARY/ABSTRACT – PROJECT 3 Nitric oxide (NO) is an important signaling molecule that influences a wide range of biological processes, including bone metabolism. Recently, we utilized argininosuccinate lyase deficiency (ASLD) as a model to study cell-autonomous, nitric oxide synthase (NOS)-dependent NO deficiency. Argininosuccinate lyase (ASL) is the only mammalian enzyme capable of synthesizing arginine, the sole precursor for NOS-dependent NO synthesis. Moreover, ASL is also required for channeling extracellular arginine to NOS for NO production. Using ASLD as a model, we reported that NO promotion of bone formation is associated with stimulation of glycolysis during osteoblast differentiation. Accumulating evidence suggests that osteoblasts utilize glucose metabolism via glycolysis as their main energy source. Importantly, some of the anabolic agents used to increase bone mass, such as Wnt proteins have also been shown to stimulate glycolysis during osteoblast differentiation, thus suggesting an essential role of glycolysis in bone anabolism. The overall goal of this project is to identify the mechanisms by which NO stimulates bone formation in physiological conditions and during mechanical loading. We also will examine whether NO effects on glycolysis are responsible for effects on bone mass. We reported that NO-deficient cells had downregulated mRNAs encoding components of the glycolysis pathway, suggesting that NO may modulate transcription factors that regulate expression of genes in the glycolysis pathway. Moreover, NO-induced stimulation of glycolysis may be involved in load-induced bone formation, a process in which abundant work has implicated NO as a critical mediator. The central hypothesis of this project is that NO production by osteoblast-lineage cells promotes osteoblast differentiation and bone formation by stimulating glycolysis. We will test this hypothesis with three specific aims. Aim 1 will determine whether NO promotes bone formation and glycolysis in osteoblasts by activating cAMP-responsive element binding protein (CREB). Aim 2 will determine whether genetic upregulation of glycolysis is sufficient to stimulate bone formation or rescue the bone mass deficiency in osteoblast-specific Asl knockout mice. Aim 3 will determine whether NO produced from osteoblasts and osteocytes is required for load-induced anabolism. Successful completion this project should advance knowledge of the molecular mechanisms underlying the effects of NO on bone. Furthermore, it may provide new insights into therapeutic targets for bone anabolism.