The Role Of Osteoclast PODXL, A Mediator Of Cell Adhesion, In Bone Metabolism

描述（由申请人提供）：我们的长期目标是描述破骨细胞促进骨代谢的分子机制，并期望这些知识将指导更有效的治疗方法来促进骨骼健康。理想情况下，减少破骨细胞介导的骨质流失的新治疗目标将阻断破骨细胞，同时保持骨形成。这可能具有挑战性，因为越来越多的证据表明，破骨细胞无论是否能够吸收骨，都是成骨细胞形成正常骨所必需的。我们检查了造血/破骨细胞谱系细胞中CD34家族成员PODXL缺失的小鼠的椎骨（hem lineage- podxldel小鼠）。骨密度和成骨细胞数量均升高。出乎意料的是，破骨细胞的数量和大小也明显增加。因此，早期破骨细胞谱系细胞中PODXL的缺失减少了骨吸收，同时保留了成骨细胞的募集和骨形成。我们的体外研究支持PODXL在破骨细胞谱系细胞中的表达可减少破骨细胞的分化。然而，破骨细胞肌动蛋白环的形成和骨吸收需要相对低水平的PODXL表达。我们的研究和已知的PODXL功能导致了我们的中心假设，即包括CXCR4和NHERF1在内的PODXL膜复合物的早期祖膜表达水平必须降低，以允许前体融合，激活破骨细胞需要较低水平的该复合物，使它们能够吸收骨。以下具体目标旨在验证这一假设1。确定破骨细胞谱系PODXL表达变化在骨骼获得和骨骼维持中的作用。我们将对hem谱系- podxldel小鼠进行综合表型分析，确定卵巢切除术对hem谱系- podxldel小鼠骨代谢的影响，确定合成代谢性甲状旁腺激素治疗过程中早期破骨细胞谱系细胞中PODXL缺失的影响，并将晚期破骨细胞谱系细胞中PODXL缺失的小鼠与hem谱系- podxldel小鼠进行比较。2. 探究PODXL、CXCR4和NHERF-1复合物影响破骨细胞谱系细胞的分子机制。我们将确定在分化过程中参与复合物形成、膜表达、分化和内化的PODXL和NHERF1的结构域，解析M-CSF下调PODXL、CXCR4和NHERF1基因表达的机制，确定PODXL及其结合伙伴对整合素相互作用、肌动蛋白环形成和破骨细胞功能的影响。

DESCRIPTION (provided by applicant): Our long term goal is to delineate the molecular mechanisms by which osteoclasts contribute to bone metabolism with the expectation that this knowledge will guide more effective therapies to promote skeletal health. Ideally, new therapy targets to reduce osteoclast-mediated bone loss will block osteoclasts while also preserving bone formation. This may be challenging since evidence is mounting that osteoclasts, whether or not they are capable of resorbing bone, are needed for normal bone formation by osteoblasts. We have examined the vertebrae of mice in which the CD34 family member PODXL is deleted in hematopoietic/osteoclast lineage cells (hem lineage-PODXLdel mice). Bone density and osteoblast numbers were both elevated. Unexpectedly, the number and size of osteoclasts were also significantly higher. Thus, loss of PODXL in early osteoclast lineage cells reduces bone resorption while preserving osteoblast recruitment and bone formation. Our in vitro studies support that PODXL expression in osteoclast lineage cells reduces osteoclast differentiation. However, a relatively low level of PODXL expression is required for osteoclast actin ring formation and bone resorption. Our studies and the known functions of PODXL lead to our central hypothesis that early progenitor membrane expression levels of a PODXL membrane complex including CXCR4 and NHERF1 must be reduced to allow for precursor fusion and lower levels of this complex are required for activation of osteoclasts, enabling them to resorb bone. The following Specific Aims are designed to test this hypothesis 1. Determine the role of osteoclast lineage PODXL expression changes in skeletal acquisition and bone maintenance. We will comprehensively phenotype the hem lineage-PODXLdel mice, establish the effects of ovariectomy on bone metabolism in hem lineage-PODXLdel mice, define the impact of loss of PODXL in early osteoclast lineage cells during anabolic PTH treatment, and compare mice with PODXL deleted in late osteoclast lineage cells to the hem lineage-PODXLdel mice. 2. Interrogate the molecular mechanisms by which the PODXL, CXCR4, and NHERF-1 complex influences osteoclast lineage cells. We will identify structural domains of PODXL and NHERF1 involved in complex formation, membrane expression, differentiation, and internalization during differentiation, resolve the mechanism of down regulation of PODXL, CXCR4, and NHERF1 gene expression by M-CSF, and ascertain the influences of PODXL and binding partners on integrin interactions, actin ring formation, and osteoclast function. PUBLIC HEALTH RELEVANCE: Bone loss associated with aging and postmenopausal osteoporosis is due to excess osteoclast-mediated bone resorption. Excess bone loss leads to risks of debilitating fractures. An estimated 44 million Americans are at risk of osteoporosis. Within the U.S., 10 million people are estimated to already have this disease and an additional 34 million are suspected of having low bone mass and at risk of developing the disease. There are 1.5 million osteoporotic fractures in the U.S. each year and the cost of caring for these fractures is estimated at $12 billion to $18 billion. Indirect costs in lost productivity of patients and care givers add billions to this cost. Thus, therapies that target bone resorbing osteoclasts while preserving bone formation have the potential to have a significant impact on patients.