Unraveling the Complexity of Lipoprotein(a) Metabolism: Human Kinetic Studies

脂蛋白(a) [Lp(a)]于1963年由Kare Berg发现，但这种独特的脂蛋白作为心血管疾病（CVD）的独立危险因素，仍有许多有待了解的地方。Hoff等人在30年前证实Lp(a)存在于动脉粥样硬化病变中，在JUPITER试验（一项高剂量他汀试验）中，Lp(a)是接受瑞舒伐他汀治疗的受试者剩余心血管疾病风险的重要决定因素。Lp(a)水平很难改变，但新批准和正在开发的药物可显著降低Lp(a)血浆浓度。Lp(a)由载脂蛋白(a) [apo(a)]与载脂蛋白B [apoB]共价结合组成，Lp(a)-apo(a)和Lp(a)-apoB都与CVD的发生有关。尽管Lp(a)在生化上与LDL密切相关，但在通过超离心分离的载脂蛋白ob的不同密度组分中发现了Lp(a)。许多研究强调了Lp(a)在疾病中的重要性，然而，关于Lp(a)在肝脏和/或血浆中的组装（载脂蛋白(a)和载脂蛋白100颗粒结合的位置和方式）以及它如何从循环中移除的关键问题仍然存在。我们的实验室使用稳定同位素来检查人体脂蛋白的清除和产生。质谱技术的最新进展使我们能够利用免疫沉淀法从血浆中分离出针对载脂蛋白(a)的Lp(a)来检测整个Lp(a)颗粒、其载脂蛋白(a)和载脂蛋白ob组分以及载脂蛋白(a)的同型异构体的清除和产生。Lp(a)水平在很大程度上取决于kvi - 2型重复序列（同种异构体）的变异。目前的应用解决了Lp(a)代谢中存在的空白，并检查了Lp(a)代谢与同工异构体大小之间的关系。我们有两个目标：目标1：我们将通过免疫沉淀（IP）从血浆中分离Lp(a)，并使用稳定同位素的引物恒定输注和新建立的质谱（MS）方法确定Lp(a)-apo(a)和Lp(a)-apoB的分数清除率和分泌率。我们还将确定所有受试者中KIV-2重复的数量。我们将招募低浓度和高浓度Lp(a)的个体(N=20, Lp（a) 15 -30nmol/L和N=20, Lp(a) 75-350nmol/L）。我们已经开发了全面的模型来表征Lp(a)代谢的途径。目的2：我们将检查来自目的1的15名具有两种可检测的载脂蛋白(a)亚型的受试者。在Lp(a)的IP后，我们将通过非变性电泳分离出两个载脂蛋白(a)亚型（每个都将与apoB100相连），并通过ms测定分离复合物中Lp(a)-载脂蛋白(a)和Lp(a)-载脂蛋白ob的fcr。随着Lp(a)治疗方法的改进，我们有必要详细和精确地了解调节Lp(a)水平的因素，以及KIV- 2型重复是否对这一心血管危险因素的代谢有影响。

Lipoprotein (a) [Lp(a)] was identified by Kare Berg in 1963, but much remains to be learned about this unique lipoprotein, which is a an independent risk factor for cardiovascular disease (CVD). Hoff et al demonstrated Lp(a) within atherosclerotic lesions 30 years ago, and in the JUPITER trial (a high dose statin trial), Lp(a) was a significant determinant of residual CVD risk in subjects receiving rosuvastatin. Lp(a) levels have been difficult to alter, but newly approved and developing drugs lower Lp(a) plasma concentrations significantly. Lp(a) is comprised of apolipoprotein (a) [apo(a)] covalently bound to apolipoprotein B [apoB], and both components Lp(a)-apo(a) and Lp(a)-apoB, are implicated in the development of CVD. Although it is closely related to LDL biochemically, Lp(a) has been found in varying density fractions of apoB isolated via ultracentrifugation. Many studies have highlighted the importance of Lp(a) in disease, however, key questions remain about the assembly of Lp(a) (where and how the apo(a) and apoB100 particle bind) within the liver and/or plasma and how it is removed from the circulation. Our laboratory uses stable isotopes to examine the clearance and production of lipoproteins in humans. Recent advances in mass spectrometry has allowed us to use Lp(a) isolated from plasma by immunoprecipitation directed against apo(a) to examine clearance and production of the whole Lp(a) particle, its apo(a) and apoB components, and isoforms of apo(a). Lp(a) levels are largely determined by variation in KVI-type 2 repeats (Isoforms). The current application addresses existing gaps in Lp(a) metabolism and examines relationships between Lp(a) metabolism and isoform size. We have two aims: Aim 1: We will isolate Lp(a) from plasma by immunoprecipitation (IP) and determine the fractional clearance rates and secretion rates of Lp(a)-apo(a) and Lp(a)-apoB using primed constant infusion of stable isotopes and newly established mass spectrometry (MS) methods. We will also determine the number of KIV-2 repeats in all subjects. We will enroll individuals with low and high Lp(a) concentrations (N=20, Lp(a) 15 -30nmol/L and N=20, Lp(a) 75-350nmol/L). We have developed comprehensive models to allow characterization of pathways of Lp(a) metabolism. Aim 2: We will examine 15 subjects from Aim 1 with two detectable apo(a) isoforms. After IP of Lp(a), we will isolate two apo(a) isoforms (each will be linked to apoB100) via non-denaturing electrophoresis and determine the FCRs of Lp(a)-apo(a) and Lp(a)-apoB within the isolated complexes via MS. As improved therapeutic approaches to Lp(a) move forward, it is imperative that we have a detailed and precise understanding of the factors that regulate Lp(a) levels and if the KIV- type 2 repeats have an effect on metabolism of this cardiovascular risk factor.