Fortilin, p53, and atherosclerosis

描述(由申请人提供)：这项题为“Fortilin，P53与动脉粥样硬化”的赠款提案的总体目标是定义Fortilin的新作用以及Fortilin-P53在动脉粥样硬化中的相互作用，目的是将抗Fortilin疗法建立为一种对抗动脉粥样硬化的Viabl策略。到2025年，全球因动脉粥样硬化及相关并发症死亡的人数预计将超过所有重大疾病，包括癌症、感染和创伤。据估计，仅在美国，每年动脉粥样硬化相关疾病的总成本就高达2,860亿美元。在他汀类药物之后，没有正在酝酿中的突破性战略来抗击这种致命的全球疾病。我们的实验室在过去的15年里一直在研究Fortilin，一种由172个氨基酸组成的多功能蛋白质。2001年，我们首次报道了Fortilin保护细胞免受凋亡的作用。最近，我们发现福尔替林介导抗病毒作用。 通过与肿瘤抑制蛋白P53的结合和抑制而发挥凋亡活性。由于(A)p53不仅可以预防癌症，还可以预防动脉粥样硬化，并且(B)Fortilin阻断了P53，因此我们使用Ldlr-/-Apobec1-/-高胆固醇血症小鼠(一种强大的动脉粥样硬化模型)测试了缺乏Fortilin是否可以改善动脉粥样硬化。值得注意的是，Fortilin+/+Ldlr-/-Apobec1-/-小鼠的动脉粥样硬化程度比Fortilin+/-Ldlr-/-Apobec1-/-小鼠高27%，并伴有广泛的巨噬细胞(M？)在动脉粥样硬化中的渗透。此外，免疫染色显示Fortilin在人类和小鼠动脉粥样硬化病变的M？和泡沫细胞中过表达。此外，冠状动脉粥样硬化患者的单核细胞具有更高的Fortilin水平。最后，我们发现高胆固醇血清能诱导M？的Fortilin。基于这些事实，我们假设：(A)促动脉粥样硬化的环境诱导M？的Fortilin，(B)M？fortilin在诱导时保护M？抵抗P53诱导的细胞凋亡，以及(C)未经抑制的M？增殖并产生促炎细胞因子，从而形成炎症和动脉粥样硬化的恶性循环。M？-fortilin通过P53的促进作用从未被假设或测试过。为了检验这一总体假设，我们提出了四个具体目标。在目标1中，我们使用原代和细胞系M？来研究Fortilin是如何在M？中诱导的，以及诱导的Fortilin在M？中所起的作用。在目标2中，我们首先定义了M-fortilin促进动脉粥样硬化形成的分子机制，使用了Ldlr-/-Apobec1-/-遗传背景上的M？特异性fortilin基因敲除(KO)小鼠(Apobec1-/-)(Apobec1-/-)。接下来，我们研究Fortilin是否通过抑制P53而促进动脉粥样硬化，通过在P53-/-Ldlr-/-Apobec1-/-遗传背景上创建M？特异性Fortilin KO小鼠(Aim 2.2)。在目标3中，我们希望通过口服1，3-D-葡聚糖包裹(GERP)的Fortilin siRNA颗粒给Ldlr-/-Apobec1/-小鼠来证明M？特异性Fortilin沉默重新激活P53并保护动脉粥样硬化。在项目结束时，我们预计M？特异性抗Fortilin疗法将被发现是对抗致命的全球性动脉粥样硬化疾病的一种开创性策略。

DESCRIPTION (provided by applicant): The overall goal of this grant proposal entitled "Fortilin, p53, and atherosclerosis" is to define a novel role of fortilin and the fortilin-p53 interaction in atherosclerosis with the intention of establishing anti- fortilin therapy as a viabl strategy against atherosclerosis. By 2025, worldwide death due to atherosclerosis and associated complications is projected to surpass that of every major disease, including cancer, infection, and trauma. The total cost of atherosclerosis-related diseases in the U.S. alone is estimated to be $286 billion annually. After statins, there is no break-through strategy in the pipeline to combat this deadly global disease. Our laboratory has studied fortilin, a 172-amino acid multi-functional protein for the last 15 years. In 2001, we reported for the first time that fortilin protects cells against apoptosis. More recently, we showed that fortilin mediates its anti apoptotic activity through its binding to and inhibition of p53, a tumor suppressor protein. Since (a) p53 protects against not only cancer but also atherosclerosis and (b) fortilin blocks p53, we tested whether the lack of fortilin ameliorated atherosclerosis using Ldlr-/-Apobec1-/- hypercholesterolemic mice, a robust model of atherosclerosis. Strikingly, fortilin+/+Ldlr-/-Apobec1-/- mice had 27 % more atherosclerosis than fortilin+/-Ldlr-/-Apobec1-/- mice with extensive macrophage (M¿) infiltration in the atheroma. In addition, immunostaining showed that fortilin is overexpressed in M¿ and foam cells within human and mouse atherosclerotic lesions. Further, monocytes from patients with coronary atherosclerosis have higher fortilin levels. Finally, we found that hypercholesterolemic sera induce fortilin in M¿. Based on these facts, we hypothesize that (a) pro-atherosclerotic milieu induces fortilin in M¿, (b) M¿-fortilin, when induced, protects M¿ against p53-induced apoptosis, and (c) unchecked M¿ proliferate and produce pro- inflammatory cytokines creating a vicious cycle of inflammation and atherosclerogenesis. The facilitative role of M¿-fortilin through p53 has never been postulated or tested. To test this overall hypothesis, we propose four Specific Aims. In Aim 1, we investigate how fortilin is induced in M¿ and what role induced fortilin plays in M¿, using primary and cell line M¿. In Aim 2, we first define the molecular mechanism by which M-fortilin promotes atherosclerogenesis, using M¿-specific fortilin knockout (KO) mice on the Ldlr-/- Apobec1-/- genetic background (Aim 2.1.). Next, we examine whether fortilin facilitates atherosclerosis through its inhibition of p53, by creating M¿-specific fortilin KO mice on the p53-/-Ldlr-/-Apobec1-/- genetic background (Aim 2.2.). In Aim 3, we expect to show that M¿-specific fortilin silencing re-activates p53 and protects against atherosclerosis by orally administering ¿1,3-D-glucan-encapsulated (GeRP) fortilin siRNA particles to Ldlr-/- Apobec1-/- mice. At the end of the project, we expect M¿-specific anti-fortilin therapy to be found to be a ground-breaking strategy against the deadly and global disease of atherosclerosis.