Role of Glucose metabolism in Chondrocyte Mechanotransduction

所有的细胞都受到机械力的影响，并对机械力做出反应，比如压缩。然而，分子机制 将机制与生物反应联系起来还不完全清楚。我们的模型系统的细胞， 软骨的软骨细胞在体内经历压缩，并且这些细胞可以将压缩成 生物信号有证据表明，软骨细胞中的葡萄糖利用受压缩和 生理压力刺激糖酵解，这是软骨细胞制造ATP的主要途径。这 这种现象与软骨细胞维持软骨的能力有关。因此，葡萄糖的研究 新陈代谢与NIH有关，因为数百万人患有软骨细胞驱动的软骨退化， 骨关节炎目前的骨关节炎治疗涉及关节运动，这是违反直觉的。我们展示了 第一次，生理相关的培养条件能够在体外压缩软骨细胞。这 项目测试的假设，生理压缩正常和骨关节炎软骨细胞 导致葡萄糖代谢中特定的代谢物模式，其支持蛋白质产生以维持 细胞微环境。前提是通过量化软骨细胞中的葡萄糖代谢， 该项目将开发使用机械加载来产生软骨修复的构建块的策略。 目的1 -体外实验将检查碳源（葡萄糖或谷氨酰胺）和 调控因变量包括性别、供体年龄和施加的压缩水平（低或高）。 靶向代谢组学数据将从正常和骨关节炎软骨细胞产生， 在不同的实验条件下。目的2 -使用自愿接受免疫治疗的小鼠的实验 跑步将评估体内机械传导。因变量包括性别和跑步的持续时间。 读数将包括靶向代谢物和免疫组织学标记物，检查细胞内的 葡萄糖代谢检测将采用高度特异性的酶抑制剂，允许逐步分析 关键的代谢途径。该项目具有实质性的创新，包括一个新的系统生物学模型 以及计算葡萄糖代谢中每一步反应的相对速率的分析方法。这些 建模结果将用于完善现有假设并产生新的假设。这个目标 该项目是为了确定小代谢物的模式的变化，导致压缩正常和 骨关节炎软骨细胞预期的结果是确定候选目标反应， 葡萄糖代谢，以增加机械驱动的氨基酸前体的产生来修复软骨。 了解这些机制可能有助于开发翻译策略，通过以下方式治愈软骨： 激活现有的机械敏感通路。深入了解软骨细胞对压力的反应将有助于 通过为软骨修复提供新的治疗靶点， 临床上取得了实质性进展。

All cells are subject to and respond to mechanical forces like compression. However the molecular mechanisms linking the mechanics to biological responses are not fully understood. The cells of our model system, the chondrocytes of cartilage, undergo compression in vivo, and these cells can transduce compression into biological signals. There is evidence that glucose utilization in chondrocytes is regulated by compression and that physiologic compression stimulates glycolysis, the main pathway chondrocytes use to make ATP. This phenomenon has been linked to the ability of chondrocytes to maintain cartilage. Thus, the study of glucose metabolism is relevant to NIH because millions suffer from chondrocyte-driven cartilage deterioration in osteoarthritis. Current osteoarthritis treatments involve joint motion, which is counterintuitive. We show for the first time that physiologically relevant culture conditions enable in vitro compression of chondrocytes. This project tests the hypothesis that physiological compression of both normal and osteoarthritic chondrocytes results in a specific pattern of metabolites within glucose metabolism that support protein production to maintain the cellular microenvironment. The premise is that by quantifying glucose metabolism in chondrocytes this project will develop strategies that use mechanical loading to produce the building blocks for cartilage repair. Aim 1 - In vitro experiments will examine the source of carbon (glucose or glutamine) and the mechanism of regulation. Dependent variables include sex, donor age and the level (low or high) of applied compression. Targeted metabolomics data will be generated from normal and osteoarthritic chondrocytes subjected to compression under different experimental conditions. Aim 2 - Experiments using mice subjected to voluntary running will assess in vivo mechanotransduction. Dependent variables include sex and the duration of running. Readouts will include both targeted metabolites and immunohistological markers examining regulation of glucose metabolism. Assays will employ highly specific enzyme inhibitors that will allow a step-by-step analysis of critical metabolic pathways. This project has substantial innovation including a novel systems biology model and analytical approach that calculate the relative rates of reaction for each step in glucose metabolism. These modeling results will be used both to refine existing hypotheses and to generate new ones. The goal of this project is to identify changes in patterns of small metabolites that result from compression for normal and osteoarthritic chondrocytes. The expected outcome is to identify candidate target reactions that leverage glucose metabolism to increase mechanically driven production of amino acid precursors to repair cartilage. Understanding these mechanisms may prove useful in developing translational strategies to heal cartilage by activating existing mechanosensitive pathways. Insight into how chondrocytes respond to compression will advance osteoarthritis translation by providing new therapeutic targets for cartilage repair and enabling substantial clinical progress.