BIOLOGY OF PROTEOLYTIC DERIVATIVES OF LP(A)

描述：存在载脂蛋白(A)的免疫反应片段， 载脂蛋白(A)，已显示在血浆，尿液和动脉粥样硬化的人。 此外，脂蛋白(A)、脂蛋白(A)和载脂蛋白(A)的衍生物可以在 弹性蛋白酶和金属蛋白酶(MMPs)作用的玻璃体 家人。我们提出的研究的目标是阐明这些产品， 就它们在心血管致病中的潜在参与而言 OL Lp(A)。特别是，我们希望检验一种假设，即基质金属蛋白酶的酶 体外显示的在连接区热点中切割Lp(a；)的家族 在apo(A)的IV-4和IV-5环之间，在组织部位产生2 Mair 衍生品。一种是mini Lp(A)，这是一种以apoB 100为蛋白质部分的颗粒 与P2相连，apo(A)的C-末端片段能够与 血管基质。另一个是F1，代表SPO(A)的N-末端结构域。 含有Kringle IV-2重复序列，缺乏基质结合功能。一个 F1的一部分一旦从组织部位的载脂蛋白(A)释放出来，就会返回到 然后迅速排到尿液中。反过来。F2。无论是免费的还是 小Lp(A)的成员，将优先保留在 形成，优先在它经历的心内膜下内膜 致动脉粥样硬化的变化。这一假设将通过四种相关的方法进行检验： 1)载脂蛋白(A)性质的结构、功能和免疫学研究 为了定义它的各个结构域，以及切割的基础 MMPs的特异性；2)在连接体4是最多的前提下 易受基质金属蛋白酶切割，体外表达具有连接子4的载脂蛋白(A) 突变为抗基质金属蛋白酶裂解，然后比较在体外和在 这些突变体与野生型突变体的活体特性； 通过腺病毒技术在apoE小鼠中的导入及其与apoE-/-杂交 和人类载脂蛋白B100转基因在动脉粥样硬化形成的不同阶段，要么是F1， P2、野生型或连接子4突变的apo(A)ANC评估这些基因的定位 未受影响和病变区域的产品以及MMI的测量 活动；4)研究人颈动脉的外科节段 用免疫化学和化学方法确定稳定斑块或不稳定斑块 方法比较载脂蛋白(A)和片段在这两种类型中的定位。 并确定这些碎片是否与 体外用MMPs消化Lp(A)/apo(A)。体外与体外相结合 活体研究有望提高我们对血管活性物质特性的了解 载脂蛋白(A)的不同结构域，并阐明了 基质金属蛋白酶介导的蛋白分解可能在脂蛋白(A)/载脂蛋白(A)在动脉粥样硬化中的作用 动脉粥样硬化的一般炎症理论的背景。

DESCRIPTION: The presence of immunoreactive fragments of apolipoprotein(a), apo(a), have been shown in the plasma, urine anc atheromas of human subjects. Moreover, derivatives of lipoprotein(a), Lp(a), and apo(a) can be generated in vitrc by the action of enzymes of the elastase and metalloproteinase (MMP) families. The goal of our proposed studies is to shed light on these products, in terms of their potential participation in the cardiovascular pathogenicity ol Lp(a). In particular, we wish to test the hypothesis that enzymes of the MMP family shown in vitro to cleave Lp(a; in the hot spot of the linker region between kringle IV-4 and IV-5 of apo(a), generate at tissue sites, two mair derivatives. One, miniLp(a), a particle that has as a protein moiety apoB 100 linked to P2, the C-terminal fragmen of apo(a) able to bind to members of the vascular matrix. The other, F1 representing the N- terminal domain of spo(a) containing the kringle IV-2 repeats and, lacking matrix binding function. A portion of F1 once releasec from apo(a) at tissue sites, would return to the plasma and then rapidly excreted into the urine. In turn. F2. eithe free or a member of a miniLp(a), would be preferentially retained at the sites of formation, preferentially in the sub endotheial intima where it undergoes atherogenic changes. This hypothesis wifi be tested by four relatec approaches: 1) structural, functional and immunological studies on the properties of apo(a) in order to define its various domains, and the basis for the cleavage specificities by MMPs; 2) on the premise that the linker 4 is thc one most susceptible to MMP cleavage, express in vitro apo(a) species having linker 4 mutated to be resistant t MMP cleavage, and then compare the in vitro and in vivo properties of these mutants with those of their wild-type counterpart; 3) introduce via adenovirus technology in apoE mice and crosses between apoE-/- and human apoB100 transgenics in different stages of atherogenesis, either Fl, P2, wild-type or linker 4-mutated apo(a) anc assess the localization of these products in unaffected and lesion areas along with measurements of MMI activities; 4) study surgical segments of human carotid arteries with either stable or unstable plaques to determine using immunochemical and chemical methods, the comparative localization of apo(a) and fragments in these tw types of plaques and also determine whether the fragments resemble those generated in vitro from the digestion o Lp(a)/apo(a) with MMPs. The combined in vitro and in vivo studies are expected to improve our knowledge oi the properties of the various domains of apo(a) and shed light on the potential role that MMP-mediated proteoIysis may play in the atherogenicity of Lp(a)/apo(a) in the context of the general inflammatory theory of atherosclerosis.