Establishment of genetic diagnostics, preventive and development of treatment for atherogenic hypertriglyceridemia

总结了2004 ~ 2006年3年的研究成果。(1)利用电化学阵列（ECA）芯片对脂蛋白脂肪酶（LPL）基因突变检测系统的改进及其在临床样品中的应用（Ikeda）：我们利用ECA芯片和二茂铁萘二亚胺作为指示剂，开发了一种新的同时多重突变检测（SMMD）系统（参考文献# 3和4）。该系统可同时识别单核苷酸多态性、插入、缺失、易位和短串联重复等多种基因突变。通过SMMD系统，我们从高甘油三酯血症患者中新发现了两个LPL基因突变。(2)通过改善LPL基因杂合突变受试者的健康生活方式来治疗高甘油三酯血症（Tamasawa和Ikeda）：杂合子LPL缺乏的受试者（携带者）容易发展为轻度高甘油三酯血症（IV型高脂蛋白血症），当…更复杂的环境因素，如高胰岛素状态和/或高酒精摄入量，刺激肝脏中甘油三酯的合成，而携带者是正常血脂，只要他们没有环境因素（参考文献# 2）。我们的研究表明，携带者的高甘油三酯血症通过让携带者改变更健康的生活方式而变为正常血脂状态。(3)基于LPL基因信息的蛋白酶抗性LPL分子的开发及其在治疗中的应用（Takagi和Ikeda）：由于天然LPL分子在血液中被蛋白酶灭活，通过将靶氨基酸交换为甘氨酸来构建蛋白酶抗性LPL分子。与天然LPL分子相比，抗蛋白酶LPL分子表现出30%的甘油三酯水解功能。由于澄清了该蛋白酶抗性LPL分子与人体血液混合后在37℃下稳定至少6小时，因此证实了该蛋白酶抗性LPL分子能够用作高甘油三酯血症治疗的可能性。为了判断蛋白酶抗性LPL分子对高甘油三酯血症的治疗效果，有必要检测肝甘油三酯脂肪酶（HTGL）的分子特征。因此，建立了测定HTGL质量的ELISA系统（参考文献# 1）。少

We summarize results obtained from the research program of 3 years through 2004 to 2006.(1) Improvement of detection system using electrochemical array (ECA) chip for lipoprotein lipase (LPL) gene mutation and its application to clinical samples (Ikeda) : We developed a new simultaneous multiple mutation detection (SMMD) system using an ECA chip and ferrocenylnapthalene diimide, as the indicator (Ref.# 3 and 4). This SMMD system can simultaneously identify several genetic mutations such as single nucleotide polymorphism, insertion, deletion, translocation and short tandem repeat. By using the SMMD system, we newly identified two LPL gene mutations from subjects with hypertriglyceridemia.(2) Treatment for hypertriglyceridemia developed in subjects with heterozygous LPL gene mutation by improving their lifestyle to a healthful one (Tamasawa and Ikeda) : Subjects (carriers) with heterozygous LPL deficiency are prone to develop mild hypertriglyceridemia (type IV hyperlipoproteinemia) when … More complicated with environmental factors such as a hyperinsulinemic state and/or a high alcohol intake, which stimulate triglyceride synthesis in the liver, while carriers are normolipidemic provided that they do not have the environmental factors (Ref.# 2). Our research revealed that hypertriglyceridemia in the carriers became normolipidemic state by getting the carriers to change to a more healthful lifestyle.(3) Development of a protease-resistant LPL molecule based on LPL gene information and its application to treatment (Takagi and Ikeda) : Because a native LPL molecule was inactivated by proteases in bloodstream, the protease-resistant LPL molecule was constructed by exchanging a target amino acid to a glycine. The protease-resistant LPL molecule exhibited 30% of the triglyceride hydrolyzing function in comparison to that of the native LPL molecule. Since it was clarified that this protease-resistant LPL molecule was steady for at least six hours at 37℃ after the protease-resistant LPL molecule was mixed with human blood, the possibility that the protease-resistant LPL molecule was able to be used as a hypertriglyceridemic treatment was confirmed. To judge the treatment effect of the protease-resistant LPL molecule to hypertriglyceridemia, it was necessary to detect a molecular characteristic of hepatic triglyceride lipase (HTGL). Therefore, ELISA system for the determination of HTGL mass was established (Ref.# 1). Less