Investigation of the role of AMP-activated protein kinase alpha2 (AMPKa2) in bone

骨生成受多种信号通路调节，激酶在其中几种信号通路中起关键作用。 信号通路为了发现调节骨量的新因素，我们筛选了一个人， 激酶siRNA文库寻找参与间充质干细胞成骨分化的激酶 干细胞小干扰RNA介导的腺苷5 '-α 2催化亚基的敲低 间充质干细胞中的单磷酸活化蛋白激酶（AMPK）活性显著增加 碱性磷酸酶，成骨分化的早期标志物，表明AMPKa 2可能具有一个重要的作用。 在成骨细胞中的作用，可能在骨形成中。AMPK是哺乳动物细胞的能量传感器，因为它 直接响应于燃料分子ATP的消耗。虽然AMPK在各种领域中的作用 外周器官，包括肌肉、心脏和肝脏，已被广泛研究，AMPK在 骨稳态尚未被描述。在这里，我们建议阐明AMPKa 2在骨中的作用， 体内平衡首先，我们将通过进行X射线照相来表征AMPKa 2基因敲除小鼠的骨骼表型。 X线摄影、DXA、骨组织形态计量学和显微计算机断层扫描。采用各种 生物化学和分子生物学方法，包括BrdU标记，von Kossa染色，定量 RT-PCR、TUNEL分析和免疫印迹分析，AMPK在成骨细胞增殖中的作用， 成熟和细胞凋亡也将在体内和离体进行研究。由于骨骼是一个动态器官， 易受全身和细胞能量平衡的影响，这项研究将为进一步研究奠定基础。 阐明能量稳态与骨代谢的关系。

Osteogenesis is regulated by multiple signaling pathways and kinases play a critical role in several of these signaling pathways. As an effort to uncover novel factors for bone mass regulation, we screened a human kinase siRNA library searching for kinases that are involved in osteogenic differentiation of mesenchymal stem cells. Small interfering RNA-mediated knockdown of alpha2 catalytic subunit of adenosine 5'- monophosphate-activated protein kinase (AMPK) in mesenchymal stem cells significantly increased activity of alkaline phosphatase, an early marker of osteogenic differentiation, suggesting that AMPKa2 may have a role in osteoblasts, probably in bone formation. AMPK is an energy sensor of mammalian cells because it directly responds to the depletion of the fuel molecule ATP. Although the roles of AMPK in various peripheral organs, including muscle, heart, and liver, have been extensively studied, implication of AMPK in bone homeostasis has not been described. Here, we propose to elucidate the role of AMPKa2 in bone homeostasis. First, we will characterize skeletal phenotype of AMPKa2 knockout mice by carrying out X-ray radiography, DXA, bone histomorphometry, and microcomputed tomography. Employing various biochemical and molecular biological approaches, including BrdU labeling, von Kossa staining, quantitative RT-PCR, TUNEL assay, and immunoblotting analyses, the role of AMPK in osteoblast proliferation, maturation, and apoptosis will also be investigated both in vivo and ex vivo. As bone is a dynamic organ that is susceptible to systemic and cellular energy balance, this study will be groundwork for the further elucidation of the association of energy homeostasis with bone metabolism.