A path to thrombosis in primary myelofibrosis

摘要 控制血栓形成是各种病理处理的核心。这里提出的研究将确定一个 新的、意想不到的血栓形成途径，涉及赖氨酰氧合酶(LOX)。拟议的工作着眼于 LOX对原发性骨髓纤维化(PMF)患者血小板功能和血栓形成的影响 骨髓中巨核细胞的增殖和聚集，骨髓纤维化。来自瑞典的数据 1980年至2009年的癌症登记显示，在11,155名骨髓增生症患者的队列中 与44,620名匹配的健康对照组相比，肿瘤患者动脉血栓形成的风险显著增加了4.9倍 与匹配的对照组相比，患者的升高(4.8p&5.0p&lt；0.001)，突出了 研究与此病理相关的控制血栓形成的调节因子。众所周知，鲑鱼会被氧化 肽基赖氨酸，导致骨髓壁龛中的交联型基质蛋白。我们发表的研究报告 发现巨核细胞中LOX上调与骨髓纤维化进展之间的联系 小鼠骨髓纤维化模型。重要的是，最近我们发现LOX的表达在 PMF患者的血小板(大多数检测出携带JAK2V617F或钙网织蛋白基因) 突变)，与匹配的对照组相比，从PMF患者分离的血小板与 I型胶原蛋白，与对照组相比。此外，转基因小鼠血小板被设计成过表达LOX，at 类似于骨髓纤维化小鼠模型细胞的水平，显示与单体胶原的粘附性增加， 而且在活体内动脉血栓形成的倾向显著增加。考虑到LOX的已知活动，我们 假设它通过至少一种胶原受体的赖氨酸氧化来影响血小板， 进一步得到了涉及Oxy-Western印迹分析的初步研究的支持。因此，我们提出了两个具体的 研究目的：目的：1.探讨LOX影响血小板对胶原反应的机制 蛋白质组学方法，以及小鼠和人类MPN样本；目的2.确定LOX- 调节骨髓纤维化小鼠体内血栓形成，以及胶原受体在这一作用中的作用。追求 这些目标将通过以下方式实现：1)我们实验室培育的一种新的转基因小鼠系，在其中，血小板LOX 水平在野生型背景下上调(不含MPN)，以及2)新LOX抑制剂的可用性， LOX基因功能丧失的研究。这项工作的创新之处在于，我们第一次揭示了 LOX和胶原蛋白受体激活。此外，这些受体以前并未被怀疑受 氧化。我们的研究具有重要意义，因为它将为研究血小板活化的基本机制提供新的线索。 一般，并涉及与PMF相关的血栓事件中的LOX，从而认识到LOX途径是新的 是未来抗血栓药物开发的潜在靶点。

Abstract Controlling thrombosis is central to management of various pathologies. Studies proposed here will identify a new, unexpected path to thrombosis, involving the enzyme Lysyl Oxidase (LOX). The proposed work looks at LOX effects on platelet function and thrombosis in primary myelofibrosis (PMF), a pathology hallmarked by proliferation and clustering of megakaryocytes, and myelofibrosis in bone marrow. Data from the Swedish Cancer Register from 1980 to 2009 showed that in a cohort of 11,155 patients with myeloproliferative neoplasms and 44, 620 matched healthy controls, the risk of arterial thrombosis was significantly 4.9-fold increased (4.8-5.0 p<0.001) in the patients compared to matched controls, highlighting the importance of studying regulators implicated in controlling thrombosis associated with this pathology. LOX is known to oxidize peptidyl lysines, leading to cross-linked matrix proteins in the bone marrow niche. Our published studied identified a link between LOX upregulation in megakaryocytes and bone marrow fibrotic progression in a mouse model of myelofibrosis. Importantly, more recently we found LOX expression to be vastly upregulated in platelets of patients with PMF (of which the majority tested carry the JAK2V617F or calreticulin gene mutations), compared to matching controls, and platelets isolated from PMF patients have greater adhesion to type I collagen, compared to controls. Further, transgenic mouse platelets engineered to overexpress LOX, at a level similar to cells from a myelofibrotic mouse model, show increased adhesion to monomeric collagen, and significantly greater propensity for arterial thrombosis in vivo. Considering LOX known activity, we hypothesized that it influences platelets through lysine oxidation of at least one of the collagen receptors, further supported by preliminary studies involving Oxy-Western blot analysis. Thus, we propose two specific aims of research: Aim 1. To explore the mechanism by which LOX affects platelet response to collagen using proteomics approaches, and mouse and human MPN samples; Aim 2. To determine the relevance of LOX- regulated thrombosis in vivo in myelofibrotic mice, and the role of collagen receptors in this effect. Pursuing these aims will be facilitated by: 1) a new transgenic mouse line produced in our lab, in which platelet LOX level is upregulated under wild type background (free of MPN), and 2) the availability of a new LOX inhibitor, and Lox gene loss of function studies. This work is innovative in that we are the first to reveal a link between LOX and collagen receptors activation. Further, these receptors were not suspected before to be regulated by oxidation. Our research is significant in that it will shed new light on basic mechanisms of platelet activation, in general, and implicate LOX in thrombotic events related to PMF, thus, recognizing the LOX pathway as new potential target for future anti-thrombosis drug development.