The role of the atypical PKCs in osteoclast function

非典型 PKC 在破骨细胞功能中的作用

• 批准号: 8410564
• 负责人: JULIA Therese WARREN
• 金额: $ 4.72万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-03-01 至 2015-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8410564
• 关键词: Actins; Bone Density; Bone Marrow; Bone Resorption; Cell Differentiation process; Cell Lineage; Cell physiology; Cells; Complex; Cytokine Receptors; Cytoskeletal Modeling; Data; Elderly; Family; Foundations; Fracture; Genes; Homeostasis; Imagery; In Vitro; Knock-out; Knockout Mice; Ligands; Mediating; Mediator of activation protein; Morbidity - disease rate; Mouse Strains; Muramidase; Mus; NF-kappa B; Osteoclasts; Osteogenesis; Osteoporosis; Pathogenesis; Phenotype; Phosphorylation; Physiological; Process; Relative (related person); Risk; Role; Signal Transduction; Structure; Testing; Therapeutic Human Experimentation; atypical protein kinase C; bone; bone loss; cathepsin K; in vivo; macrophage; monocyte; mortality; mutant; novel therapeutics; osteoclastogenesis; public health relevance; receptor; skeletal; skeletal disorder; therapeutic development

DESCRIPTION (provided by applicant): Osteoporosis is a skeletal disease characterized by bone resorption in excess of bone formation. The osteoclast is the sole bone resorbing cell and therefore provides an important avenue for novel therapeutic development. The mature osteoclast is derived from the monocyte/macrophage lineage and requires proper differentiation and cell polarization for its function, both of which are downstream of signaling through the cytokine receptor activator of NF-kB ligand (RANKL). One potential mediator of RANKL-induced osteoclast function is the atypical protein kinase C (aPKC) sub-family, composed of PKC; and PKC6. The role of these molecules in the osteoclast has not been elucidated, but preliminary data suggests that they may be important for osteoclast function. Specifically, our lab has found that RANKL can induce phosphorylation of the aPKCs and their association with an active polarization complex composed of Par-3, Par-6, and the aPKCs. It is also possible that these molecules may be important for the differentiation of the osteoclast. Therefore, we propose to study the role of the aPKCs in osteoclast differentiation and function (Specific Aim #1) by utilizing several mouse strains lacking the aPKCs alone or in combination. We will determine the effect of aPKC deletion on osteoclastogenesis in vitro (Sub-aim A) and on osteoclast function (Sub-aim B) using several different parameters (actin ring visualization, cathepsin K localization, media CTx release, and bone pit formation). Then, we will generate mutants of the aPKCs which we will retrovirally transduce into osteoclasts lacking the appropriate gene, allowing us to perform structure/function analysis of these molecules(Sub-aim C). Finally, we will examine the in vivo phenotype of mice lacking the aPKCs (using histomorphometry and micro-CT) to determine their role in skeletal homeostasis or pathological bone loss (Specific Aim #2). Overall, we will determine the role of the aPKCs in osteoclast differentiation and function in vitro and in vivo.

描述（由申请人提供）：骨质疏松症是一种骨骼疾病，其特征是骨吸收超过骨形成。破骨细胞是唯一的骨吸收细胞，因此为新的治疗开发提供了重要途径。成熟破骨细胞来源于单核细胞/巨噬细胞谱系，需要适当的分化和细胞极化才能发挥功能，这两者都是通过NF-κ B配体的细胞因子受体激活剂（RANKL）进行信号传导的下游。RANKL诱导破骨细胞功能的一种潜在介质是非典型蛋白激酶C（aPKC）亚家族，由PKC和PKC 6组成。这些分子在破骨细胞中的作用尚未阐明，但初步数据表明，它们可能对破骨细胞功能很重要。具体而言，我们的实验室发现RANKL可诱导aPKC磷酸化及其与由Par-3、Par-6和aPKC组成的活性极化复合物的结合。这些分子也可能对破骨细胞的分化很重要。因此，我们建议通过利用单独或组合缺乏aPKC的几种小鼠品系来研究aPKC在破骨细胞分化和功能中的作用（具体目标#1）。我们将使用几种不同的参数（肌动蛋白环可视化、组织蛋白酶K定位、介质CTx释放和骨陷窝形成）确定aPKC缺失对体外破骨细胞生成（子目标A）和破骨细胞功能（子目标B）的影响。然后，我们将产生aPKC的突变体，我们将逆转录病毒转染到缺乏适当基因的破骨细胞中，使我们能够对这些分子进行结构/功能分析（子目标C）。最后，我们将检查缺乏aPKC的小鼠的体内表型（使用组织形态计量学和micro-CT），以确定其在骨骼稳态或病理性骨丢失中的作用（具体目标#2）。总之，我们将在体外和体内确定aPKCs在破骨细胞分化和功能中的作用。