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

脂蛋白(A)[Lp(A)]是由Kare Berg在1963年发现的，但对这种独特的脂蛋白仍有许多需要了解的地方，它是心血管疾病(CVD)的一个独立危险因素。Hoff等人在30年前证明了Lp(A)在动脉粥样硬化病变中的作用，在Jupiter试验(一项高剂量他汀类药物试验)中，Lp(A)是接受罗舒伐他汀治疗的受试者残余心血管风险的重要决定因素。Lp(A)水平很难改变，但新批准和开发的药物显著降低了Lp(A)的血浆浓度。Lp(A)是由载脂蛋白(A)[apo(A)]与载脂蛋白B[apoB]共价结合而成，Lp(A)-apo(A)和Lp(A)-apoB均参与CVD的发生发展。虽然Lp(A)在生化上与低密度脂蛋白密切相关，但已在超速离心法分离的不同密度组分的apoB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(apo(A)和apoB100颗粒在哪里以及如何结合)以及如何将其从循环中清除仍然是关键问题。我们的实验室使用稳定的同位素来检查人类体内脂蛋白的清除和产生。质谱学的最新进展使我们能够利用针对apo(A)的免疫沉淀法从血浆中分离出Lp(A)，以检测整个Lp(A)粒子、其apo(A)和apoB组分以及apo(A)的异构体的清除和产生。Lp(A)水平在很大程度上由KVI-2型重复序列(亚型)的变化决定。目前的应用解决了Lp(A)代谢中存在的差距，并检查了Lp(A)新陈代谢和异构体大小之间的关系。目的1：用免疫沉淀法从血浆中分离Lp(A)，并用稳定同位素持续输注和新建立的质谱学方法测定Lp(A)-apo(A)和Lp(A)-apoB的清除率和分泌率。我们还将确定所有受试者中KIV-2重复的数量。我们将招收低和高Lp(A)浓度(N=20，Lp(A)15-30nmol/L和N=20，Lp(A)75-350nmol/L)的个体。我们已经开发了全面的模型来描述Lp(A)代谢的途径。目标2：我们将用两种可检测到的载脂蛋白(A)亚型来检查目标1中的15名受试者。在对Lp(A)进行ip后，我们将通过非变性电泳法分离两种apo(A)亚型(每个亚型都与apoB100相连)，并通过MS测定分离的复合体中Lp(A)-apo(A)和Lp(A)-apoB的FCR。随着Lp(A)治疗方法的改进，我们必须详细和准确地了解调节Lp(A)水平的因素，以及KIV-2型重复序列是否对这一心血管危险因子的代谢产生影响。

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.