骨肉瘤中HIF-1α清除ROS的机制研究

Ser/Thr protein kinase activation, which is a common molecular phenomenon in osteosarcoma, can induce over-expression of hypoxia inducible factor-1α（HIF-1α）and subsequent activation of downstream targets. Recent study demonstrated that AKT activation can increase the accumulation of intracellular reactive oxygen species (ROS), and evidence showed that ROS can further induce AKT activation, thus a positive feedback of AKT-ROS formed. Accumulation of ROS will result in cell apoptosis, but resistance to apoptosis could be seen in osteosarcoma with AKT activation. The above findings indicated that AKT activation related over-expression of HIF-1αmight initiate a mechanism of ROS degeneration. Our previous studies indicated that high expression of HIF-1α in osteosarcoma tissues significantly associated with poor survival, inhibition of HIF-1α in osteosarcoma cell line MG-63 resulted in accumulation of intracellular ROS. The machanism that HIF-1α degeneates ROS remains unclear, and there were few reports about the implication in osteosarcoma. Our study aim to investigate the molecular mechanism of the degeneration of ROS by HIF-1αin AKT activation osteosarcoma cell lines with previously constructed interference vector and over-expression vector, focusing on the generation and degeration of intracellular ROS. Furthermore, we will perform an investigational experiment for novel therapies based on the mechanism.

丝氨酸/苏氨酸蛋白酶（AKT）活化是骨肉瘤细胞中常见的分子现象，它可以引起肿瘤细胞表达乏氧诱导因子-1α（HIF-1α）并活化下游靶基因。最新的研究发现AKT活化容易导致活性氧（ROS）在细胞内的累积，而ROS又可以进一步活化AKT，形成正反馈。过量的ROS累积可导致肿瘤细胞死亡，而AKT活化的骨肉瘤往往存在凋亡抵抗。这提示AKT活化引起HIF-1α高表达后，可能启动了对ROS的降解。我们的前期研究表明，在骨肉瘤组织中HIF-1α的表达与预后相关；在骨肉瘤细胞系MG-63中抑制HIF-1α的表达后，可导致ROS累积。但目前HIF-1α降解ROS的机制未阐明，是否具有临床应用价值少有报道。本课题拟在AKT活化的骨肉瘤细胞系中，利用已构建的HIF-1α干扰载体和过表达载体，从ROS的生成和降解两方面进行研究，深入探讨HIF-1α降解ROS的分子机制，并进一步探索骨肉瘤治疗新的方案。

丝氨酸/苏氨酸蛋白酶（AKT）活化是骨肉瘤细胞中常见的分子现象，它可以引起肿瘤细胞表达乏氧诱导因子-1α（HIF-1α）并活化下游靶基因。最新的研究发现AKT活化容易导致活性氧（ROS）在细胞内的累积，而ROS又可以进一步活化AKT，形成正反馈。过量的ROS累积可导致肿瘤细胞死亡。这提示AKT活化引起HIF-1α高表达后，可能启动了对ROS的降解。我们的前期研究表明，在骨肉瘤组织中HIF-1α的表达与预后相关；在骨肉瘤细胞系MG-63中抑制HIF-1α的表达后，可导致ROS累积。本研究验证了在AKT活化的骨肉瘤细胞系中HIF-1α可降解ROS，改变细胞内HIF-1α的表达水平可导致骨肉瘤细胞内ROS的变化。利用已构建的HIF-1α干扰载体和过表达载体，发现HIF-1α通过抑制线粒体呼吸减少ROS的生长，并上调FOXO1及其下游相关基因的表达来降解ROS，HIF1α对FOXO1的作用为直接结合在其启动子区的乏氧反应元件(HRE) 来完成。HIF-1α抑制剂和ROS诱导剂在体外和体内都具有抗肿瘤作用，且表现出了一定的协同效应。如期完成实验结果方面的目标，实验结果正在整理总结和分析，部分相关文章已投稿。