Fortilin, p53, and atherosclerosis

描述（由申请人提供）：本授权提案题为“Fortilin、p53和动脉粥样硬化”的总体目标是确定Fortilin和Fortilin-p53相互作用在动脉粥样硬化中的新作用，目的是建立抗Fortilin治疗作为抗动脉粥样硬化的可行策略。到2025年，全球因动脉粥样硬化和相关并发症导致的死亡预计将超过每一种主要疾病，包括癌症，感染和创伤。仅在美国，动脉粥样硬化相关疾病的总成本估计为每年2860亿美元。在他汀类药物之后，没有突破性的战略可以对抗这种致命的全球性疾病。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，我们首次报道了福替林保护细胞免于凋亡。最近，我们发现，fortilin介导其抗 通过其与肿瘤抑制蛋白p53的结合和抑制其凋亡活性。由于（a）p53不仅可以预防癌症，还可以预防动脉粥样硬化，（B）fortilin可以阻断p53，因此我们使用Ldlr-/-Apobec 1-/-高胆固醇血症小鼠（一种动脉粥样硬化的稳健模型）测试缺乏fortilin是否改善动脉粥样硬化。值得注意的是，fortilin+/+Ldlr-/-Apobec 1-/-小鼠的动脉粥样硬化比fortilin+/-Ldlr-/-Apobec 1-/-小鼠多27%，动脉粥样硬化中有广泛的巨噬细胞（M？）浸润。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 ²和泡沫细胞中过表达。此外，来自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，高胆固醇血症血清诱导fortilin在M？。基于这些事实，我们假设：（a）促动脉粥样硬化环境诱导M <$中的fortilin，（B）M <$-fortilin，当被诱导时，保护M <$免受p53诱导的细胞凋亡，和（c）未受抑制的M <$增殖并产生促炎细胞因子，产生炎症和动脉粥样硬化形成的恶性循环。Mü-fortilin通过p53的促进作用从未被假设或测试过。为了验证这一假设，我们提出了四个具体目标。在目的1中，我们研究了福替林是如何在M <$中诱导的，以及诱导的福替林在M <$中起什么作用，使用原代和细胞系M <$。在目标2中，我们首先使用Ldlr-/-Apobec 1-/-遗传背景下的M-特异性fortilin敲除（KO）小鼠，定义了M-fortilin促进动脉粥样硬化形成的分子机制（目标2.1.）。接下来，我们通过在p53-/-Ldlr-/-Apobec 1-/-遗传背景下建立M？特异性fortilin KO小鼠，研究fortilin是否通过抑制p53促进动脉粥样硬化（目的2.2.）。在目标3中，我们希望通过向Ldlr-/-Apobec 1-/-小鼠口服1，3-D-葡聚糖封装（GeRP）fortilin siRNA颗粒来证明M ²特异性fortilin沉默重新激活p53并预防动脉粥样硬化。在该项目结束时，我们预计M？特异性抗fortilin疗法将被发现是一种针对致命的全球性动脉粥样硬化疾病的突破性策略。

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.