乙酸代谢影响间充质干细胞成骨成脂分化命运及骨质疏松的机制研究

细胞的能量物质代谢紊乱是导致骨稳态失衡的重要因素。我们前期研究揭示谷氨酰胺和葡萄糖等碳源物质作为线粒体三羧酸循环的代谢供体，影响间充质干细胞（MSC）的成骨分化及矿化。进一步探索发现短链脂肪酸乙酸可能是MSC分化的新能量源，本项目将研究代谢酶AceCS1/2介导乙酸在胞质/线粒体中的代谢转变和调控机制，揭示乙酸代谢影响MSC成脂/成骨分化命运的作用。拟采用13C同位素示踪、能量代谢分析仪等技术研究乙酸代谢途径和能量产生；通过慢病毒沉默/过表达、双荧光素酶报告系统、ChIP等方法重点研究线粒体代谢酶AceCS2的基因转录和活性调控机制，揭示衰老、代谢应激等条件下乙酸代谢对线粒体活性和稳态的影响；构建AceCS2条件性敲除小鼠和生酮饮食诱导代谢模型，在体内研究乙酸代谢异常影响MSC分化命运及其与骨质疏松发生的关联机制，探索基于代谢通路调控MSC分化、干预骨质疏松的新策略方向。

Dysregulation of energy metabolism is believed to be involved in imbalance of bone homeostasis leading to degenerative bone diseases such as osteoporosis. Our previous work revealed a regulatory pathway in which carbon sources such as glutamine and glucose are funneled into the mitochondrial TCA cycle generating energy and intermediates that contribute to osteogenic differentiation and mineralization of mesenchymal stem cells (MSC). On the other hand, we recently discovered that acetate, which is the main short chain fatty acid found in the circulation, might be an alternative fuel for MSC differentiation. In the current study, we aim to investigate the metabolic shift of acetate between cytoplasmic and mitochondrial metabolism under the regulation of AceCS1/2, exploring the underlying mechanism by which acetate metabolism contributes to the fate determination of MSC. We will employ techniques such as 13C-stable isotopically labeled tracer and Seahorse XF cell energy phenotype test to analyze the metabolites and energy production of acetate metabolism. Through manipulating gene expression by lentivirus infection, Duo-luciferase assay and ChIP assay, we will focus on defining the regulatory mechanism by which mitochondrial AceCS2 expression and enzymatic activity are fined tuned, discovering how metabolites and its regulators affect mitochondrial oxidative phosphorylation activity and thus mitochondrial function. In addition, we will generate Prx1-cre; AceCS2-flox conditional knockout mice and utilize ketone diet-induced metabolic models to investigate the effect of metabolic changes of acetate and its relationship with pathological progress of osteoporosis in vivo. Our study will provide a further understanding of metabolic changes involved in bone diseases, paving ways to preventing or reversing degenerative bone diseases via novel metabolic pathway manipulations.