Gravitational Regulation of Osteoblast Genomics and Metabolism

描述（由申请人提供）：地球上重力在骨骼发育和维持中的作用是有争议的。我们目前的理解是，骨的重力卸载导致骨质流失（骨质减少），骨质疏松症通常发展为骨质疏松症并最终骨折。这是一个严重的问题，卧床不起的病人和老年人的经验。在轨道上的宇航员每月骨质流失约1%至3%，这相当于地球上骨质疏松症患者一年的骨质流失。了解骨骼对不同程度重力的反应对于开发合理的药物设计或物理治疗方法以减轻骨质减少至关重要。目前，将骨头暴露在长期缺乏重力的环境下的唯一方法是在国际空间站上进行轨道自由落体。这些研究既昂贵又难以在国际空间站上进行。研究骨动力学的重力调节的理想方法是有一种基于地面的方法来消除或减少骨上的重力。创造一个能加速骨质流失的环境将给我们提供一个工具来加速我们对潜在的基因组、蛋白质组学和代谢组学机制的理解。提出的研究旨在确定地面磁悬浮是否适合用于细胞培养研究的轨道自由落体模拟。研究细胞培养物而不是整个动物，可以进行高度控制和可重复的实验。我们计划在与地面悬浮磁铁和国际空间站空间微重力环境兼容的生物反应器中培养MC3T3成骨细胞和原代破骨细胞。我们独特的悬浮磁铁集成了一个37℃的孵化器集成到磁铁孔允许悬浮数周。在国际空间站上进行实验的执行伙伴是位于科罗拉多州博尔德的BioServe空间技术公司。BioServe公司与美国宇航局在近地轨道上进行生物实验的合作已有20年的历史。在先前的轨道飞行任务中，成骨细胞和破骨细胞显示出微重力影响细胞分化和基因表达的证据。拟议研究的指标包括细胞形态学研究，微阵列分析测量差异基因表达，ELISA和RT-PCR鉴定成骨细胞和破骨细胞分化标记，代谢组学，通过H-1核磁共振光谱和MAPK， Twsg1和NF-kB信号通路的定量。通过这一套分析，我们将确定模拟微重力和国际空间站微重力环境中许多代谢物和生物标志物的异同。该项目将有助于确定可以解决的机制和途径，以减轻地球上的骨质流失。

DESCRIPTION (provided by applicant): The role of gravity in the development and maintenance of bone on earth is controversial. Our current understanding is that gravitational unloading of bone results in bone loss (osteopenia) which typically progresses to osteoporosis and ultimately bone fracture. This is a significant problem that bed- ridden patients and the elderly experience. Astronauts in orbit experience significant osteopenia of approximately 1% to 3% of bone loss per month, which is equivalent to a year of bone loss for osteoporotic individuals on earth. Understanding bone response to varying levels of gravitational force is critical toward developing rational approaches in drug design or physical therapy toward mitigating osteopenia. Presently, the only means of exposing bone to a long-term lack of gravitational force is through orbital free fall aboard the ISS. These studies are expensive and logistically difficult to do aboard the ISS. An ideal method of study gravitational regulation of bone dynamics is to have a ground-based method to remove or reduce the gravitational force on bone. Creating an environment that amplifies the rate of bone loss will give us a tool to accelerate our understanding of the underlying genomic, proteomic and metabolomic mechanisms. The proposed study seeks to determine if ground-based magnetic levitation is a suitable simulation of orbital free fall for cell culture studies. Studying a cell culture rather than an entire animal allows for highly controlled and reproducible experiments. We plan to grow MC3T3 osteoblastic and primary osteoclastic cells in a bioreactor compatible with our ground-based levitation magnet and the ISS space-based microgravity environment. Our unique levitation magnet integrates a 37 C incubator integrated into the magnet bore allowing levitation for several weeks. The implementation partner for experiments aboard the ISS is BioServe Space Technologies in Boulder, CO. BioServe has a two-decade record of accomplishment of working with NASA on delivering biological experiments to and from low-earth orbit. Osteoblasts and osteoclasts flown on previous orbital missions show evidence that microgravity affects cell differentiation and gene expression. The metrics for the proposed study include cell morphology studies, microarray analysis to measure differential gene expression, ELISA and RT-PCR to identify markers of osteoblast and osteoclast differentiation, metabolomics, via H-1 NMR spectroscopy and quantification of MAPK, Twsg1 and NF-kB signaling pathways. With this suite of assays, we will determine similarities and differences in a number of metabolites and biomarkers for simulated microgravity and ISS microgravity environments. This project will help identify mechanism(s) and pathways that can be addressed to mitigate bone loss on earth.