GAPDH, DNA Repair and Atherosclerosis

项目摘要/摘要 DNA损伤的累积被认为是动脉粥样硬化发生和发展的原因之一。 血管平滑肌细胞(SMC)中的基因组不稳定性导致细胞凋亡，并有助于 动脉粥样硬化斑块易损性。甘油醛3-磷酸脱氢酶(GAPDH)是最主要的 细胞感受器最终负责维持细胞内环境的稳定，但其在 动脉粥样硬化的形成是完全未知的。已有研究表明，一种关键的促动脉粥样硬化的脂质OxLDL GAPDH在人主动脉SMC中的表达下调，且GAPDH在 动脉粥样硬化斑块中的SMC和低水平的GAPDH与增加的细胞凋亡有关。GAPDH 通过一种新的涉及核的分子机制减少DNA损伤和抑制SMC凋亡 GAPDH与主要的氧化DNA修复酶--脱嘌呤/脱嘧啶核酸内切酶1(APE1)相互作用。 SMC特异性GAPDH过表达减少DNA损伤，减少斑块SMC凋亡 减少动脉粥样硬化的负担。重要的是，GAPDH过表达小鼠的动脉粥样硬化斑块 SMC水平升高，胶原增加，坏死核减少，富含SMC的纤维帽更厚，提示 增强斑块稳定性。当前建议的主要重点是研究调解机制 人参皂苷脱氢酶诱导的抗动脉粥样硬化和稳定斑块作用及测定人参皂苷脱氢酶是否 多肽(GMP)可减轻动脉粥样硬化小鼠的负担，改善斑块的稳定性。主 假说是GAPDH通过以下途径减轻动脉粥样硬化负担并增强斑块稳定性 促进APE1依赖的DNA修复和抑制SMC的凋亡。这将检验该假设在 以下是具体目标： 具体目的1：证明SMC特异性GAPDH调节DNA修复、细胞凋亡、 动脉粥样硬化负荷与斑块稳定性特征及识别机制。 特异性目的2：确定GAPDH型多肽(GMP)是否能激活APE1，刺激 DNA修复，抑制细胞凋亡，减轻动脉粥样硬化。 建议使用腺相关病毒(AAV)进行SMC靶向GMP cDNA转移 动脉粥样硬化小鼠。基于AAV的载体被批准用于人类，因此长期目标是使用 AAV-GMP载体作为一种创新的DNA修复和抗凋亡治疗方法治疗不稳定 动脉硬化。GAPDH抗动脉粥样硬化作用的研究及GMP诱导的实验研究 稳定斑块的作用将导致开发新的靶向疗法来治疗动脉粥样硬化和 预防急性血管事件。

Project Summary/Abstract Accumulated DNA damage is recognized as a causal factor in the initiation and progression of atherosclerosis. Genomic instability in the vascular smooth muscle cells (SMC) leads to cell apoptosis and contributes to atherosclerotic plaque vulnerability. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) is the major cellular sensor ultimately responsible for maintaining of cellular homeostasis, however its specific role in atherogenesis is completely unknown. It has been shown that a key pro-atherogenic lipid OxLDL downregulated GAPDH in human aortic SMC and that GAPDH expression was markedly decreased in the atherosclerotic plaque SMC and that low GAPDH level was associated with increased apoptosis. GAPDH reduced DNA damage and suppressed SMC apoptosis via a novel molecular mechanism involving nuclear GAPDH interaction with apurinic/apyrimidinic endonuclease 1 (Ape1), the major oxidized DNA repair enzyme. SMC-specific GAPDH overexpression decreased DNA damage, reduced plaque SMC apoptosis and decreased atherosclerotic burden. Importantly, atherosclerotic plaques in GAPDH-overexpressing mice had elevated SMC levels, increased collagen, reduced necrotic cores and thicker SMC-rich fibrous caps suggesting enhanced plaque stability. The major focus of the current proposal is to study the mechanism mediating GAPDH-induced anti-atherosclerotic and plaque stabilizing effect and to determine whether GAPDH mimicking peptide (GMP) reduces atherosclerotic burden and improve plaque stability in atherosclerotic mice. The main hypothesis is that GAPDH reduces atherosclerotic burden and enhances features of plaque stability via stimulation of Ape1-dependent DNA repair and suppression of SMC apoptosis. This will test the hypothesis in following Specific Aims: Specific Aim 1: To demonstrate that SMC-specific GAPDH regulates DNA repair, apoptosis, atherosclerotic burden and features of plaque stability and identify mechanism. Specific Aim 2: To determine whether GAPDH-mimicking peptide (GMP) will activate Ape1, stimulate DNA repair, suppress cell apoptosis and reduce atherosclerosis. Proposal will use adeno-associated viruses (AAVs) to perform SMC-targeted GMP cDNA transfer in atherosclerotic mice. AAV-based vectors are approved to use in humans, therefore the long-term goal is to use the AAV-GMP vector as an innovative pro-DNA repair and anti-apoptotic therapy to treat unstable atherosclerosis. Studying of anti-atherosclerotic effects of GAPDH and demonstration that GMP induces plaque-stabilizing effect will lead to the development of novel targeted therapies to treat atherosclerosis and prevent acute vascular events.