Molecular basis of mechanotransduction in bone cells

描述(申请人提供)：骨质疏松症在美国是一个重要的健康问题，是一种骨形成和骨吸收过程之间不平衡的疾病。众所周知，机械加载会影响这些过程。我们的长期目标是阐明骨细胞对机械负荷的反应的分子机制。在过去的几年里，大量的证据表明，细胞外核苷酸，如ATP，通过P2嘌呤能受体传递信号，在骨行为的调节中发挥重要作用。我们和其他人证明，机械负荷诱导的液体流动导致骨细胞释放ATP，而ATP随后通过P2Y嘌呤能受体激活钙信号通路。P2Y受体是一种G蛋白偶联受体，在正常的生物学过程中起着重要的作用。然而，P2Y受体在骨生物学中的作用，特别是在骨力学转导中的作用，目前还不清楚。我们的初步数据表明，由ATP激活的P2Y受体的一个亚型--P2Y2参与了体液流动时基因表达的变化。此外，GPCRs的脱敏最近被证明是骨骼机械传感装置的一个重要组成部分。我们的结果表明，G蛋白偶联受体激酶2(GRK2)参与了液体流动引起的P2Y激活的脱敏。有趣的是，我们的初步结果表明，P2Y2缺陷小鼠表现出中等的骨表型。更重要的是，我们的数据表明，P2Y2缺陷小鼠对机械负荷的成骨反应受到抑制。因此，我们的中心假设是，生物物理信号，如液体流动，通过一种涉及P2Y嘌呤能受体的机制来调节骨细胞的行为，该受体被G蛋白偶联受体激酶脱敏。为了验证这一假设，我们将进行一系列体外和体内实验，以检测P2Y受体在骨细胞机械转导中的作用(AIM 1)，GRK(AIM 2)和甲状旁腺激素(PTH)(AIM 3)对P2Y受体的调节，以及对P2Y和GRK缺乏的小鼠完整骨的机械负荷效应(AIM 4)。公共卫生方面的依赖。骨质疏松症是一个严重的健康问题，影响着超过4400万美国人。该项目旨在确定通过P2Y受体在骨骼中进行机械转导的分子机制。该项目的完成将最终导致对骨质疏松等具有机械成分的骨骼疾病进行药物干预的新靶点。

DESCRIPTION (provided by applicant): Osteoporosis, a significant health problem in US, is a disease with imbalance between the processes of bone formation and bone resorption. It is known that mechanical loading can influence these processes. Our long-term goal is to elucidate the molecular mechanisms of bone cell response to mechanical loading. Over the past several years, substantial evidence has indicated that extracellular nucleotides, such as ATP, signaling through P2 purinergic receptors, play an important role in the regulation of bone behavior. We and others demonstrated that mechanical loading induced fluid flow causes ATP release in bone cells, and ATP subsequently activates calcium signaling pathways via P2Y purinergic receptors. P2Y receptor is a G protein-coupled receptor (GPCR), which is universally critical in normal biological processes. However, the role of P2Y receptors in bone biology, particularly in bone mechanotransduction, is unknown. Our preliminary data suggest that P2Y2, one subtype of P2Y receptors, activated by ATP is involved in changes in gene expression in response to fluid flow. In addition, desensitization of a GPCR's has recently been shown to be an important component of the mechanosensing apparatus in bone. Our results suggest that G-protein coupled receptor kinase 2 (GRK2) is involved in the desensitization of P2Y activation in response to fluid flow. Interestingly, our preliminary results suggest that P2Y2 deficient mice exhibit moderate bone phenotype. More importantly, our data suggest that the osteogenic response to mechanical loading in P2Y2 deficient mice is suppressed. Thus, our central hypothesis is that biophysical signals, such as fluid flow, regulate bone cell behavior via a mechanism involving P2Y purinergic receptors which are desensitized by G-protein coupled receptor kinases. To test this hypothesis we will conduct a series of in vitro and in vivo experiments to examine the role of P2Y receptors in bone cell mechanotransduction (aim 1), the regulation of P2Y receptors by GRK (aim 2) and parathyroid hormone (PTH) (aim 3), and mechanical loading effects on intact bone from mice deficient in P2Y and GRK (aim 4). PUBLIC HEALTH RELVANCE. Osteoporosis is a significant health problem that affects over 44 million Americans. The proposed project is to determine the molecular mechanism responsible for mechanotransduction in bone via P2Y receptors. Completion of this project will ultimately lead to novel targets for pharmacological intervention in bone diseases that have a mechanical component, such as osteoporosis.