Lysophosphatidic acid & tissue factor in atherosclerosis

描述（由申请人提供）：本提案的长期目标是探索LPA（氧化LDL的一种成分）在动脉粥样硬化中的病理作用。该提案的驱动假设是，LPA（oxLDL的一种成分）导致动脉粥样硬化的机制之一是通过上调TF表达。这一假说是基于我们的新发现：1）脂质溶血磷脂酸（LPA），最近被证明在动脉粥样硬化病变中以高浓度蓄积，并被鉴定为oxLDL的组分，显著增加SMC中TF mRNA、TF蛋白的产生和TF活性; 2）LPA诱导的TF基因表达需要特定的G蛋白、MAPK、PKC、新型蛋白激酶D（PKD）和核糖体S6激酶（p90 RSK）的激活; 3）LPA可诱导动物心脏、肾脏和主动脉组织中TF mRNA的表达。这一假说也得到了其他人最近报道的观察结果的支持：1）组织因子（TF）在动脉粥样硬化的发展中起重要作用; 2）LPA在动脉粥样硬化病变中以高浓度蓄积; 3）LPA诱导大鼠主动脉新生内膜形成。这一假设是创新的，因为在动脉粥样硬化的发展中，这种磷脂LPA的作用是完全未知的，它在LDL氧化过程中形成，并从活化的血小板中分泌出来。本提案的目标将通过以下具体目标来实现：目标1：鉴定和确定LPA受体，特定的G-蛋白，和特定亚型的蛋白激酶C在LPA刺激的TF表达中的作用，使用LPA受体的特异性拮抗剂，亲和标记，反义寡核苷酸，以及显性阴性的方法。 目的2：利用选择性抑制剂和显性负调控策略，探索PKD和p90 RSK在TF基因调控中的新生物学作用。 目标3：使用成熟的动脉粥样硬化模型-ApoE基因敲除小鼠，以确定i）LPA在动脉粥样硬化发展中的作用; ii）TF在体内介导LPA在动脉粥样硬化病变发展中的病理作用中的作用。 从拟议的研究中获得的信息可能会导致识别新的治疗靶点，用于预防和治疗动脉粥样硬化。拟议的研究也将有助于了解介导LPA生物学效应的细胞信号通路，特别是对基因调控的影响。拟议的研究还将提供新的见解PKD和p90 RSK在血管壁中的生物学和细胞功能。

DESCRIPTION (provided by applicant): The long-term objective of this proposal is to explore the pathological role of LPA, a component of oxidized LDL, in atherosclerosis. The driving hypothesis of the proposal is that one of the mechanisms by which LPA, a component of oxLDL, contributes to atherosclerosis is via up-regulation of TF expression. This hypothesis is based on our novel findings that 1) lipid lysophosphatidic acid (LPA), which has recently been shown to accumulate in high concentrations in atherosclerotic lesions and identified as a component of oxLDL, markedly increases TF mRNA, TF protein production and TF activity in SMC; 2) LPA induced-TF gene expression requires the activation of a particular G-protein, MAPK, PKC, the novel protein kinase D (PKD), and ribosome S6 kinase (p90RSK); and 3) LPA induces TF mRNA expression in heart, kidney and aorta of the living animal. This hypothesis is also supported by the observations recently reported by others: 1) Tissue factor (TF) plays an important role in the development of atherosclerosis; 2) LPA accumulates in high concentrations in atherosclerotic lesions; and 3) LPA induces neointimal formation in rat aorta. This hypothesis is innovative because in the development of atherosclerosis, the role of this phospholipid LPA, which forms during oxidation of LDL and is secreted from activated platelets, is totally unknown. The goals of this proposal will be achieved through the following specific aims: Aim 1:To identify and determine the role of LPA receptor, specific G-protein, and the particular isoform of protein kinase C in LPA-stimulated TF expression, using specific antagonists of LPA receptors, affinity labeling, antisense oligonucleotides, as well as the dominant-negative approach. Aim 2: To explore the novel biological role of PKD and p90RSK in TF gene regulation, using selective inhibitors and the dominant-negative strategy. Aim 3: Using a well-established atherosclerotic model-ApoE knockout mouse, to determine i) the role of LPA in the development of atherosclerosis; ii) the role of TF in mediating the pathological effect of LPA in atherosclerotic lesion development in vivo. The information obtained from the proposed studies may lead to the identification of novel therapeutic targets for the prevention and treatment of atherosclerosis. The proposed studies would also contribute to the understanding of the cellular signaling pathway that mediates the biological effects of LPA, specifically the effects on gene regulation in general. The proposed studies would also provide novel insight into the biological and cellular function of PKD and p90RSK in the vascular wall.