Demystify Heparan Sulfate–Sclerostin Interactions in Bone Formation

揭秘硫酸乙酰肝素与硬化素在骨形成中的相互作用

• 批准号: 10593214
• 负责人: Miaomiao Li
• 金额: $ 15.95万
• 依托单位: STATE UNIVERSITY OF NEW YORK AT BUFFALO
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-02-01 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10593214
• 关键词: Address; Affect; Binding; Binding Sites; Biochemical; Biological; Biological Availability; Biological Process; Biology; Bone Diseases; Bone Matrix; Bone Resorption; Bone Surface; Cell Lineage; Cell surface; Complex; Data; Diffusion; Disease; Drug Targeting; Extracellular Matrix; Future; Glycosaminoglycans; Health; Heparitin Sulfate; Impairment; In Vitro; Knock-in Mouse; Knowledge; Mammalian Cell; Maps; Mission; Modeling; Molecular; Monoclonal Antibodies; Mouse Strains; Oligosaccharides; Osteoblasts; Osteocytes; Osteogenesis; Osteoporosis; Persons; Point Mutation; Public Health; Recombinants; Research; Role; Site-Directed Mutagenesis; Stains; Surface; System; Testing; United States National Institutes of Health; WNT Signaling Pathway; Work; aged; bone cell; bone loss; bone strength; cell growth regulation; experimental study; fracture risk; in vivo; inhibitor; mutant; new therapeutic target; novel; receptor; skeletal disorder; tool; tool development; translational impact

Project Summary Sclerostin is a potent inhibitor of bone formation and has been shown to be a valuable drug target for treating osteoporosis. Mechanistically, sclerostin functions by binding to LRP5/6 on the osteoblast lineage cells to antagonize canonical Wnt signaling, thus negatively regulates bone formation. Presumably, after sclerostin is secreted by osteocytes, they reach the target osteoblasts at the bone surfaces by diffusion. However, to date it remains unknown how secreted sclerostin is regulated on the cell surface and in the extracellular matrix. To address this significant gap in sclerostin biology, we focus our study on sclerostin–heparan sulfate (HS) interaction. HS, a universal glycosaminoglycan found at the cell surface and in the extracellular matrix, is known to bind sclerostin and might regulate the bioactivity and diffusion of sclerostin. Our central hypothesis is that HS can regulate the biological functions of sclerostin in bone formation. To test this hypothesis, our overall objective here is to elucidate how HS interacts with sclerostin and how HS–sclerostin interaction regulates bone formation. We plan to pursue the following two specific aims: Aim 1. Determine the biological significance of HS–sclerostin interactions in vitro. We postulate that HS helps concentrate sclerostin on the osteoblast cell surface and facilitates the binding of sclerostin to its receptor LRP5/6 by forming ternary complex. We also hypothesize that HS serves a storage depot of sclerostin on the cell surface of osteocytes after it is secreted, and protects it from proteolytic degradation. We will manipulate HS–sclerostin interactions biochemically at the surface of both osteoblasts and osteocytes to determine the mechanisms by which HS regulates the function of sclerostin in these cellular contexts. Aim 2. Determine the role of HS–sclerostin interaction in bone formation in vivo. Our working hypothesis is that dampening HS–sclerostin interactions impairs the inhibitory potency of sclerostin towards LRP5/6, which leads to enhanced bone formation. Using a novel sclerostin knock-in mouse strain, we will examine the consequence of altering HS–sclerostin interactions in bone formation in vivo. Our contribution will be significant because we will identify multiple molecular mechanisms by which HS regulates sclerostin and elucidate how such interactions regulate bone formation. Results from the proposed experiments will substantially advance our understanding of the cellular regulation of sclerostin on both osteoblasts and osteocytes by elucidating the role of HS in the system. Importantly, these results are expected to have positive translational impact because by identifying how HS regulates the bioavailability of sclerostin, we may be able to provide new perspective for manipulating sclerostin in bone diseases.

项目摘要 硬化蛋白是骨形成的潜在抑制剂，已被证明是有价值的药物 治疗骨质疏松症的靶标。从机械上讲，硬化素通过与LRP5/6结合在上的功能 成骨细胞谱系细胞与规范Wnt信号拮抗，从而负调节 骨形成。据推测，硬化蛋白被骨细胞分泌后，它们到达靶 通过扩散在骨表面的成骨细胞。但是，迄今为止，它仍然未知 分泌的硬化蛋白受到细胞表面和细胞外基质的调节。解决 在硬化蛋白生物学中，我们的研究将研究重点放在硬化素 - 硫酸盐（HS）上 相互作用。 HS，一种在细胞表面和细胞外发现的通用糖胺聚糖 已知基质结合硬化素，可能调节硬化素的生物活性和扩散。 我们的中心假设是HS可以调节硬化蛋白在骨中的生物学功能 形成。为了检验这一假设，我们的总体目标是阐明HS的相互作用 用硬化素以及HS – Sclerostin的相互作用如何调节骨形成。我们计划购买 以下两个具体目的：目标1。确定HS – Sclerostin的生物学性 体外相互作用。我们假设HS有助于将硬化素浓缩在成骨细胞上 表面并通过形成端子来促进硬化蛋白与其受体LRP5/6的结合 复杂的。我们还假设HS在细胞表面提供硬化蛋白的存储沉积物 骨细胞分泌后的骨细胞，并保护其免受蛋白水解降解。我们会操纵 在成骨细胞和成骨细胞的表面上，HS – Sclerostin相互作用在生化上 确定HS调节硬化蛋白功能的机制 上下文。 AIM 2。确定HS – Sclerostin相互作用在体内骨形成中的作用。我们的 工作假设是，该死的HS – Sclerostin相互作用会损害抑制效力 朝着LRP5/6的硬化蛋白的of，导致骨形成增强。使用新颖的硬化蛋白 敲入小鼠应变，我们将检查改变HS – Sclerostin相互作用的结果 在体内骨形成中。我们的贡献将是重要的，因为我们将确定多个 HS调节硬化症并阐明这种相互作用的分子机制 调节骨形成。提出的实验的结果将大大推动我们的 了解硬化蛋白在成骨细胞和骨细胞上的细胞调节 阐明HS在系统中的作用。重要的是，这些结果预计将具有积极 翻译影响是因为通过确定HS如何调节硬化蛋白的生物利用度，我们 也许能够为操纵骨骼疾病中的硬化蛋白提供新的观点。