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Detemination of The Main Advanced Glycation End Product of The Maillard Reaction and Its Significance as A Biochemical Marker for Diabetic Complications

美拉德反应主要高级糖基化终产物的测定及其作为糖尿病并发症生化标志物的意义

基本信息

批准号：
07557076
负责人：
HORIUCHI Seikoh
金额：
$ 9.28万
依托单位：
Kumamoto University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
1995
资助国家：
日本
起止时间：
1995 至 1996
项目状态：
已结题

项目摘要

Incubation of glucose with proteins in vitro results in formation of early-products such as Schiff base and Amadori product. Upon further incubations, these early-products leads to formation of advanced glycation end products (AGE) which are characterized by fluorescence, brown color and intra-and intermolecular crosslinking. Recent immunological studies using anti-AGE antibody suggest a potential role of AGE-modification in aging and age-enhanced disease processes such as atherosclerosis and diabetic complications. To solve this suggestion, a main AGE-structure (s) expressed in vivo is to be determined. In the present study, we isolated the main AGE-structure from AGE-lysine derivative and determined its chemical structure.The following results were obtained by two-years projects.[1] The AGE-sample was obtained after long-term incubation of alpha-tosyl-lysine-methyl ester with glucose and the main fluorescent compound named as X1 was purified from this sample. Chemical analyzes by mass spectroscopy and 1H-NMR and 13C-NMR successfully identified its structure as 1,7-disubstituted-5- (1,2,3,4-tetrahydroxybutyl) -1,4-dihydro-4-oxo-1,7-naphthyridinium cation, indicating two lysine residues are crosslinked by pyridinium structure derived from two glucose molecules.[2] Immunological studies using the rabbit polyclonal anti-X1 antibody demonstrated the presence of X1 in (i) human lens proteins, (ii) foam cells as well as in extracellular spaces in human atherosclerotic lesions, (iii) glomerulus and renal tubules of diabetic nephropathy, (iv) in Carpal tunnel deposits (beta2-microglobulin) of dialysis-related amyloidosis, (v) elastin fibers of skin dermis in actinic elastosis. These results indicate that X1, one of the main AGE-structures in vivo, plays an important role in aging and/or age-enhanced disease processes such as diabetic complications and atherosclerosis of vascular walls.

葡萄糖与蛋白质在体外孵育导致早期产物如Schiff碱和Amadori产物的形成。进一步孵育后，这些早期产物导致形成晚期糖基化终产物（AGE），其特征为荧光、棕色以及分子内和分子间交联。最近的免疫学研究表明，抗AGE抗体的AGE修饰在衰老和年龄增强的疾病过程中的潜在作用，如动脉粥样硬化和糖尿病并发症。为了解决这个问题，需要确定体内表达的主要AGE结构。本研究通过两年的项目研究，从AGE-赖氨酸衍生物中分离出主要的AGE-结构，并确定了其化学结构。[1]在α-甲苯磺酰-赖氨酸-甲酯与葡萄糖长期孵育后获得AGE样品，并从该样品中纯化命名为X1的主要荧光化合物。通过质谱和1H-NMR和13 C-NMR的化学分析成功地鉴定了其结构为1，7-二取代-5-（1，2，3，4-四羟基丁基）-1，4-二氢-4-氧代-1，7-萘啶鎓阳离子，表明两个赖氨酸残基通过衍生自两个葡萄糖分子的吡啶鎓结构交联。[2]使用兔多克隆抗X1抗体进行的免疫学研究证明，X1存在于（i）人透镜蛋白，（ii）泡沫细胞以及人动脉粥样硬化病变的细胞外间隙，（iii）糖尿病肾病的肾小球和肾小管，（iv）透析相关淀粉样变性的腕管沉积物（β 2-微球蛋白），（v）光化性弹性组织变性皮肤真皮的弹性蛋白纤维中。这些结果表明，X1是体内主要的AGE结构之一，在衰老和/或年龄增强的疾病过程中起着重要作用，如糖尿病并发症和血管壁的动脉粥样硬化。