IDENTIFYING COLLAGEN-I-INDUCED MRNAS IN OSTEOBLASTS

骨质慢性病是一个日益严重的健康问题 这需要改进诊断和治疗。的重要性 细胞外基质（ECM）组分如I型胶原蛋白与细胞外基质（ECM）组分如I型胶原蛋白结合。 骨的生理学通过在骨中发现的胶原蛋白突变来说明。 骨生成障碍和绝经后骨质疏松症患者。 然而，ECM介导的骨生理调节还没有得到很好的定义。 ECM-成骨细胞的相互作用可能是中央的病理生理， 骨质疾病。本提案的目标是确定 成骨细胞-1型胶原的末端反应（诱导蛋白表达） 交互. 这一目标将通过克隆成骨细胞（OB）培养来实现 模型和cDNA文库的消减cDNA探针筛选以鉴定 cDNA（代表胶原蛋白I诱导的蛋白质）独特地存在于 I型胶原刺激的OB。因此，第一个具体目标是 建议将是建立cDNA文库，代表谱 由a）静息OB和B）胶原I刺激的OB表达的蛋白质。的 第二个具体目标是鉴定编码蛋白质的cDNA， 表达是特异性的胶原蛋白I刺激的OB，将需要两个cDNA a）通过苯酚再结合合成消减探针 杂交技术（pERT）和B）筛选胶原I刺激的 OB cDNA文库。最后，第三个目标是确保 鉴定的蛋白质是OB与胶原I相互作用的结果 矩阵 这将通过北方印迹比较 未刺激和胶原I刺激的OB转录物表达 在特定目标2中鉴定为cDNA。 最近对骨重建的综述表明， 骨量的临床管理将需要识别 骨重建的分子组成部分。拟议查明 ECM刺激的OB表达的蛋白质代表了一个细胞周期的启动。 一项旨在确定成骨细胞的分子决定因素的研究计划， 生理学和骨量。

Chronic diseases of bone mass represent a growing health care problem which demands improved diagnosis and treatment. The importance of extracellular matrix (ECM) components such as type I collagen to the physiology of bone is illustrated by the collagen mutations found in individuals with osteogenesis imperfecta and post menopausal osteoporosis. Yet, ECM-mediated regulation of osseous physiology is not well defined. ECM-ostoblast interactions may be central to the pathophysiology of diseases of bone mass. The goal of the present proposal is to identify end-responses (induced protein expression) of osteoblast -type 1 collagen interactions. This goal will be accomplished using a clonal osteoblast (OB) cell culture model and subtracted cDNA probe screening of a cDNA library to identify cDNAs (representing collagen I-induced proteins) uniquely present in collagen I-stimulated OB. Therefore, the first specific aim of this proposal will be to create cDNA libraries which represent the spectrum of proteins expressed by a) resting OB and b) collagen I-stimulated OB. The second specific aim, to identify the cDNAs that encode proteins whose expression is specific to collagen I-stimulated OB, will require both cDNA libraries to a) synthesize a subtracted probe by the Phenol Reassociation Hybridization Technique (pERT) and b)to screen the collagen I-stimulated OB cDNA library. Finally, the third aim will be to assure that the identified proteins are the result of OB interaction with a collagen I matrix. This will be accomplished by Northern blot comparison of unstimulated and collagen I-stimulated OB expression of transcripts identified as cDNAs in Specific Aim 2. A recent review of bone remodeling suggested that improvements in the clinical management of bone mass will require the identification of the molecular components of bone remodelling. The proposed identification of proteins expressed by ECM-stimulated OB represents the initiation of a research program that seeks to define molecular determinants of osteoblast physiology and bone mass.