MOLECULAR MECHANISMS OF OSTEOCLASTIC BONE RESORPTION

描述(改编自《调查员摘要》)：的总体目标 拟议的研究是为了确定OC-116 kDa，a 新描述的分子，在破骨细胞的功能和应用这一知识 控制溶骨性疾病。拟议的研究涉及一个 深入分析OC-116 kDa的表达、调控和功能 在破骨细胞性骨吸收中。拟议研究的假设是 OC-116 kDa在破骨细胞中选择性表达；它构成 破骨细胞质子泵的组成部分；它起着关键作用 在亚碎屑陷窝酸化中的功能作用。在AIM 1、用RT-PCR方法检测OC-116 kDa在体内的表达， 核糖核酸酶保护，原位杂交和免疫化学，在细胞 和超微结构水平。在目标2中，OC-116 kDa的结合是 破骨细胞质子泵的浓度将通过共沉淀法确定 OC-116 kDa，与泵的已知亚基结合，并在功能上通过反义 体外抑制作用。在目标3中，OC-116 kDa的功能将是 通过靶向破坏内源基因而建立的。的影响 胚胎表型和破骨细胞功能的零突变， 将测定新生小鼠和成年小鼠。在目标4中，对 骨吸收刺激和抑制因子对破骨细胞OC-116 kDa的影响 将会被评估。此外，对细胞特异性的机制也进行了研究。 OC-116 kDa的调控将通过启动子-记者启动 转基因动物。在目标5中，OC-116 kDa的特定抑制剂将是 寻求并确定其对骨吸收的影响。凭借其 OC-116 kDa的选择性表达可作为特异性靶点 吸收的治疗性抑制剂。

DESCRIPTION (Adapted from the investigator's Abstract): The overall goal of the proposed studies is to determine the role of OC-116 kDa, a newly-described molecule, in osteoclast function and to apply this knowledge to the control of osteolytic diseases. The proposed studies involve an in-depth analysis of the expression, regulation, and function of OC-116 kDa in osteoclastic bone resorption. The hypothesis of the proposed studies is that OC-116 kDa is expressed selectively in osteoclasts; that it constitutes an integral part of the osteoclast proton pump; and that it plays a key functional role in the acidification of the subosteoclastic lacuna. In Aim 1, the expression of OC-116 kDa will be determined in vivo, using RT-PCR, RNase protection, in situ hybridization and immunochemistry, at the cellular and ultrastructural levels. In Aim 2, the association of OC-116 kDa as part of the osteoclast proton pump will be determined by coprecipitation of OC-116 kDa with known subunits of the pump, and functionally by antisense inhibition in vitro. In Aim 3, the function of OC-116 kDa will be established using targeted disruption of the endogenous gene. The effect of the null mutation on phenotype and osteoclast function in embryonic, neonatal and adult mice will be determined. In Aim 4, the regulation of OC-116 kDa in osteoclasts by stimulators and inhibitors of bone resorption will be assessed. In addition, studies of the mechanisms of cell-specific regulation of OC-116 kDa will be initiated using promoter-reporter transgenic animals. In Aim 5, specific inhibitors of OC-116 kDa will be sought and their effect on bone resorption determined. By virtue of its selective expression, OC-116 kDa may be useful as a target of specific therapeutic inhibitors of resorption.