CELL CYCLE IN OLIGODENDROCYTE DEVELOPMENT

描述：申请人的目的是了解如何 少突胶质细胞的发育和髓鞘形成在CNS中受到调节， 旨在设计脱髓鞘疾病的治疗方法。 Miskimins博士 初步数据显示， 少突胶质细胞祖细胞的表达伴随着 细胞周期蛋白依赖性激酶抑制剂p27kip1（和细胞周期蛋白E的减少）。 她 建议增加p27kip1是发展所必需的， 少突胶质细胞的分化。 具体来说，p27kip1的增加 可以解释在分化之前增殖的停止 也可能在分化程序的激活中发挥作用。 最初的目标是确定少突胶质细胞的特定阶段 p27kip1水平上升的发展。 在这项研究中，Miskimins博士将 利用少突胶质细胞祖细胞（O-2A细胞）以及CG-4细胞 她报告说，这条线保留了O-2A细胞的许多特性。 另一组研究是确定生长因子的作用， 信号分子（bFGF，TGFB和cAMP）已知中度少突胶质细胞 p27kip1表达的研究进展。 申请人预测， 成纤维细胞生长因子（bFGF）将干扰 p27 kip 1表达增加，转化生长因子B（TGF B）和cAMP表达增加 p27kip1水平。 在这些研究中，她将使用O-2A细胞并监测 p27 kip1蛋白免疫荧光检测，定量mRNA 通过北方印迹法。 第三个目的是确定是否表达 p27kip1能促进O-2A细胞的分化。 构建体 为了在诱导型启动子的控制下表达p27kip1， 转染到CG-4或增殖的O-2A细胞中。 申请人 预测p27kip1的诱导应该使增殖停止， 促进分化。 反义技术将用于确定 如果p27kip1是少突胶质细胞发育所必需的。 增编 提供的证据表明，细胞周期蛋白E水平也应监测 这是提议。

DESCRIPTION: The applicant s aim is to gain understanding of how oligodendrocyte development and myelination are regulated in the CNS, with a view to design therapies for demyelinating disorders. Dr. Miskimins presents preliminary data indicating that the differentiation of oligodendrocyte progenitor cells is accompanied by a marked increase in the cyclin dependent kinase inhibitor p27kip1 (and a decrease in cyclin E). She proposes that the increase in p27kip1 is necessary for development and differentiation of oligodendrocytes. Specifically, the increase in p27kip1 would account for the cessation of proliferation prior to differentiation and could also play a role in the activation of the differentiation program. An initial goal is to define the specific stage of oligodendrocytes development at which p27kip1 levels rise. For this study Dr. Miskimins will utilize the oligodendrocyte progenitor (O-2A cell), as well as the CG-4 cell line which she reports retains many of the properties of the O-2A cell. Another set of studies is to define the effects of growth factors and signaling molecules (bFGF, TGFB and cAMP) known to moderate oligodendrocyte development on the expression of p27kip1. The applicant predicts that basic fibroblasts growth factor (bFGF) will interfere with the elevation of p27kip1 while transforming growth factor b (TGF b) and cAMP should increase p27kip1 levels. For these studies she will utilize O-2A cells and monitor p27kip1 by immunofluorescence in Western blots and she will quantitate mRNA by Northern blotting. The third aim is to determine if expression of p27kip1 is sufficient to promote differentiation of O-2A cells. Constructs for expression of p27kip1 under the control of an inducible promoter will be transfected into either CG-4 or proliferating O-2A cells. The applicant predicts that induction of p27kip1 should bring proliferation to a halt and promote differentiation. Antisense technology will be utilized to determine if p27kip1 is required for oligodendrocyte development. An addendum provides evidence suggesting that cyclin E levels should also be monitored and this is proposed.