Genetics of Osteoblast Differentiation

描述(由申请人提供)：从间充质干细胞(MSCs)中适当地发育成熟的、有功能的成骨细胞对数百万患有骨质疏松症的美国人的骨骼健康至关重要。成骨细胞负责新骨组织的形成和矿化，在骨稳态中起着关键作用。一种潜在的治疗骨质疏松症的方法是促进成骨细胞的活动，以对抗许多患者中发生的骨材料丢失。然而，成骨细胞的发育过程和分化在分子水平上还不足以确定潜在的新药靶点。在这项研究中，我们建议使用RNA-seq技术在从5个不同品系的实验室小鼠的纯化细胞培养中检测从MSC到成熟成骨细胞分化过程中的全基因组转录动力学。同时，我们将测量我们培养物的体外矿化动力学，以及同一品系小鼠骨形成的体内组织形态计量学测量。我们将分析这些RNA-SEQ数据以确定成骨细胞形成的关键转录事件，包括所有被检查菌株共同的显著表达变化和不同遗传背景的独特表达变化。通过将这些表达数据与现有的体内表型数据以及将在该项目中测量的体外和体内表型相结合，我们将把转录变化与临床上重要的表型联系起来。通过这一过程确定的基因和蛋白质将包括潜在的药物靶点，可以在未来的项目中进一步评估和研究。 与公共健康相关：据预测，目前有4370万50岁以上的美国人已经或正处于患骨质疏松症的严重风险中。根据卫生局局长最近的一份报告，美国每年有超过150万例骨质疏松性骨折，其中包括28万例髋部骨折和50万例脊椎骨折。每10名女性中有2名和每10名男性中有3名髋部骨折患者将在骨折后1年内死亡，一半人将永远不会回到以前的生活状态，70%-80%的人将永远不能在没有人帮助的情况下行走。在美国，每年用于治疗骨质疏松症的直接医疗费用超过183亿美元。越来越多的证据表明，骨髓间充质干细胞(MSCs)向功能性成骨细胞的正确分化对骨的健康和动态平衡起着关键作用。在小鼠和人身上的研究表明，随着年龄的增长，骨髓中的成骨细胞数量减少，骨髓肥胖度增加。骨质疏松症最有希望的治疗途径之一是开发促进成骨细胞功能的合成代谢药物。目前，FDA只批准了一种促进成骨细胞的药物(特瑞帕替德)，但这种化合物具有重大风险，因此建议使用不超过两年。为了更好地了解骨质疏松症的遗传基础并开发新的治疗方法，我们必须从分子和遗传学的角度了解成骨细胞分化的过程，这也是本提案的目标。

DESCRIPTION (provided by applicant): The proper development of mature, functional osteoblasts from mesenchymal stem cells (MSCs) is critical to the bone health of millions of Americans suffering from osteoporosis. Osteoblasts play a key role in bone homeostasis as they are responsible for the formation and mineralization of new bone tissue. A potential treatment for osteoporosis is to promote osteoblast activity in order to combat the loss of bone material that occurs in many patients. However, the developmental progression and differentiation of osteoblasts is inadequately characterized at a molecular level to identify potential new drug targets. In this study, we propose to measure whole genome transcriptional dynamics using RNA-seq technology throughout the differentiation process from MSC to mature osteoblast in purified cell cultures derived from 5 diverse strains of laboratory mouse. In parallel, we will measure in vitro mineralization dynamics of our cultures, as well as in vivo histomorphometric measures of bone formation in mice of the same strains. We will analyze these RNA-seq data to indentify the key transcriptional events of osteoblastogenesis, including significant expression changes common to all examined strains and unique to different genetic backgrounds. By integrating these expression data with existing in vivo phenotypic data as well as the in vitro and in vivo phenotypes that will be measured in this project, we will correlate transcriptional changes with clinically important phenotypes. The genes and proteins identified through this process will include potential drug targets that can be further evaluated and studied in future projects. PUBLIC HEALTH RELEVANCE: It is predicted that currently 43.7 million Americans over the age of 50 already have or are at serious risk of developing osteoporosis. Based on a recent surgeon general's report, in the United States there are over 1.5 million osteoporotic fractures each year, including 280,000 hip fractures and 500,000 vertebral fractures. Two out of every 10 women and 3 out of every 10 men who suffer a fracture of the hip will die within 1 year of that fracture, half will never return to their previous living situation, and 70-80% will never walk again unaided. The annual direct health care costs for treatment of osteoporosis in the United States is over $18.3 billion. Increasing evidence shows that proper differentiation of mesenchymal stem cells (MSCs) into functional osteoblasts plays a key role in bone health and homeostasis. Studies in both mouse and human have shown that with age, the number of osteoblasts in marrow decreases and the degree of marrow adiposity increases. One of the most promising therapeutic avenues for the treatment of osteoporosis is the development of anabolic agents which promote osteoblast function. Currently, only a single osteoblast promoting drug is approved by the FDA (teriparatide), but this compound carries significant risks and is thus indicated for use for no more than two years. In order to better understand the genetic bases of osteoporosis and to develop new treatments, we must understand the process of osteoblast differentiation from both a molecular and genetic perspective, which is the goal of this proposal.