Role of GSK3beta in Osteoblast Cell Cycle, Differentiat*

描述（由申请人提供）：骨质疏松症是糖皮质激素（GC）用于改善自身免疫性和炎症性疾病症状的主要不良反应。导致gc治疗患者骨质流失的最重要的单一机制是抑制成骨细胞骨形成。我们最近发现了在培养成骨细胞分化过程中的一个承诺阶段（“鹅卵石”阶段），在这个阶段，GC发挥了抑制作用。在这个阶段，增殖持续存在，但与融合前的细胞周期控制相比，增殖受到独特的调节。在这个承诺阶段，而不是更早，GC抑制细胞周期进程，这种抑制是通过糖原合成酶激酶3 β (gskβ)的Ser9抑制磷酸化的衰减介导的。GSK3beta3在细胞增殖和分化中起关键作用。它与Wnt β - catenin通路和生长因子激活的PI3激酶/Akt通路均有关联。大量研究表明，生长因子如igf - 1和FGF-2在成骨细胞分化中的作用，但gsk3 β的参与尚未得到充分研究。最近的一些报道表明Wnt信号在成骨细胞分化中的作用，但gsk3 β的参与也没有得到证实。因此，我们建议研究gsk3 β在成骨细胞分化相关的细胞周期、成骨细胞表型的发展以及gc介导的这些过程的抑制中的作用。在Specific Aim 1中，我们将研究gsk3 β在PI3激酶和Akt下游的作用。在Specific Aim 2中，我们将研究GSK3beta在Wnt下游的作用。在这两个特定目标中，我们将测量gc处理与未处理的成骨细胞培养中每个途径的各种成分的水平和活性。此外，为了绘制直接受GC影响的步骤，我们将增强或抑制每个途径中的信号分子，并检查GC处理和未处理培养物对gsk3 β、细胞周期以及分化标记物的影响。这些研究将揭示成骨细胞生长和分化的新方面，以及这些过程如何受到糖皮质激素的不利影响。

DESCRIPTION (provided by applicant): Osteoporosis is a major adverse effect of glucocorticoid (GC) treatment for the amelioration of symptoms of autoimmune and inflammatory diseases. The most important single mechanism contributing to bone loss in GC-treated patients is inhibition of osteoblastic bone formation. We recently identified a commitment stage during osteoblast differentiation in culture (the "cobblestone" stage) in which GC exert their inhibitory effect. During this stage, proliferation persists, but is regulated uniquely compared to control of the cell cycle prior to confluence. At this commitment stage, but not earlier, GC inhibit cell cycle progression and this inhibition is mediated by attenuation of the Ser9 inhibitory phosphorylation of Glycogen Synthase Kinase 3beta(GSKbeta). GSK3beta3 plays pivotal roles in cell proliferation and differentiation. It has been implicated in both the Wnt beta - catenin pathway and the growth factor-activated PI3 kinase/Akt pathway. Numerous studies demonstrated roles for growth factors such as IGF-I and FGF-2 in osteoblast differentiation, but the involvement of GSK3beta has not been sufficiently studied. Several recent reports suggest a role for Wnt signaling in osteoblast differentiation, but again the involvement of GSK3beta has not been tested. Thus, we propose to examine the role of GSK3beta in the osteoblast differentiation-related cell cycle, in the development of the osteoblast phenotype and in GC-mediated inhibition of these processes. In Specific Aim 1, we will examine the role that GSK3beta plays down stream of PI3 Kinase and Akt. In Specific Aim 2, we will examine the role that GSK3beta plays downstream of Wnt. In both Specific Aims, we will measure levels and activities of various components of each pathway in GC-treated versus non-treated osteoblast cultures. In addition, to map the step that is directly affected by GC, we will either augmnent or suppress signaling molecules in each pathway, and examine the effects on GSK3beta, the cell cycle as well as differentiation markers in GC-treated and non-treated cultures. These studies will shed light on novel aspects of osteoblast growth and differentiation, and how these processes are adversely affected by glucocorticoids.