Lysyl Oxidase Like 2: A Novel Target in Aging Associated Vascular Stiffening

项目概要： 老龄化中的中心血管硬度增加是主要不良心血管事件的独立预测因子 事件和老年人发病率和死亡率的主要原因。虽然血管硬化的机制 正在出现，靶向治疗尚未可用。血管硬化及其后遗症在临床上仍然存在 目前，治疗依赖于生活方式的改变和药物靶向 原发性高血压，尽管有不同的病因和细胞分子机制，导致晕厥， 老年人的福尔斯。随着人口统计学向老年人、临床、社会和 血管硬化的经济负担增加了。因此，迫切需要查明和 表征血管硬化临床管理的潜在目标并减轻其后遗症。 使用无偏蛋白质组学筛选，我们已经确定赖氨酰氧化酶样2（LOXL 2）作为一种新的蛋白质 介导血管硬化我们的假设得到了初步数据的有力支持，即LOXL 2 调节基质组成、沉积和结构，调节血管平滑肌细胞（VSMC） 微力学和收缩性，并有助于小鼠血管硬度的老化相关增加。在 在这项研究中，我们建议在老年人中研究LOXL 2介导的血管硬化的机制。 从细胞到衰老小鼠模型的分层方式。我们创造了新的试剂，小鼠模型， 并组建了一个强大的多学科研究团队，成功地执行了拟议的工作。 在目标1中，我们将确定一氧化氮和蛋白水解加工对LOXL 2调节的机制。 通过因子Xa在VSMCs中。细胞和脉管系统中LOXL 2调节的这些方面尚不清楚。在Aim中 2、我们推测清道夫受体富含半胱氨酸（SRCR）的结构域和胺氧化酶（LOX）结构域 存在于L0 XL 2中的蛋白具有独特的功能，其独特地促成血管硬化。我们将1）定义 LOXL 2的ECM重塑机制，特别强调其推定的支架功能，以及2） 确定LOXL 2如何调节VSMC微观力学，细胞骨架动力学和行为/功能。在 目的3，我们将确定靶向L0 XL 2是否减轻小鼠模型中年龄相关的血管硬化。 使用遗传（他莫昔芬诱导的全局、VSMC特异性和EC特异性LOXL 2-/-小鼠）的组合， 和药理学（PAT-1251）方法来靶向L0 XL 2，我们将鉴定1）L0 XL 2的细胞来源， 老化血管，2）研究LOXL 2靶向是否可以停止或逆转老化中的血管硬化，以及3） 确定VSMC硬化或ECM沉积是否是LOXL 2在VSMC中起作用的主要机制。 老船总之，这些研究将促进我们对LOXL 2蛋白生物化学和生物学的了解， 建立LOXL 2在脉管系统中调节的机制，通过以下方式确定机制： 其中LOXL 2驱动年龄相关的血管硬化，并为这一发现奠定科学基础， 开发用于血管硬化临床管理的新型有效疗法。

Project summary: Increased central vascular stiffness in aging is an independent predictor of major adverse cardiovascular events and the leading cause of morbidity and mortality in the elderly. While mechanisms of vascular stiffening are emerging, targeted therapies are not yet available. Vascular stiffening and its sequelae remain clinically challenging to manage and currently, care relies on lifestyle modifications and pharmacologic agents targeting essential hypertension despite the distinct etiology and cellular molecular mechanisms, leading to syncope and falls in the elderly. and with the shift of population demographics towards the elderly, the clinical, societal, and financial burden of vascular stiffening has grown. Therefore, there is an urgent need to identify and characterize potential targets for clinical management of vascular stiffening and mitigate its sequelae. Using an unbiased proteomic screen, we have identified lysyl oxidase like 2 (LOXL2) as a novel protein that mediates vascular stiffening. Our hypothesis, strongly supported by preliminary data, is that LOXL2 regulates matrix composition, deposition, and architecture, modulates vascular smooth muscle cell (VSMC) micromechanics and contractility, and contributes to aging associated increase in vascular stiffness in mice. In this grant, we propose to examine the mechanisms of LOXL2-mediated vascular stiffening in aging in a hierarchical fashion from cells to mouse models of aging. We have generated novel reagents, mouse models, and assembled a strong, multidisciplinary research team to successfully execute the proposed work. In Aim 1, we will determine the mechanism of LOXL2 regulation by nitric oxide and proteolytic processing by Factor Xa in VSMCs. These aspects of LOXL2 regulation in cells and vasculature are not yet known. In Aim 2, we postulate that the scavenger receptor cysteine rich (SRCR) domains and amine oxidase (LOX) domains present in LOXL2 have distinct functions that contribute uniquely to vascular stiffening. We will 1) define the mechanisms of ECM remodeling by LOXL2 with specific emphasis on its putative scaffolding function and 2) determine how LOXL2 modulates VSMC micromechanics, cytoskeletal dynamics, and behavior/function. In Aim 3, we will determine if targeting LOXL2 mitigates age-associated vascular stiffening in mouse models. Using a combination of genetic (tamoxifen inducible global, VSMC-specific, and EC-specific LOXL2-/- mice), and pharmacologic (PAT-1251) approaches to target LOXL2, we will identify 1) the cellular source of LOXL2 in the aging vessel, 2) investigate if LOXL2 targeting can halt or reverse vascular stiffening in aging, and 3) identify whether VSMC stiffening or ECM deposition is the primary mechanism by which LOXL2 acts in the aging vessel. In sum, these studies will advance our knowledge of LOXL2 protein biochemistry and biology, establish the mechanisms by which LOXL2 is regulated in the vasculature, determine the mechanisms by which LOXL2 drives age-associated vascular stiffening, and lay the scientific foundation for the discovery and development of novel and effective therapies for the clinical management of vascular stiffening.