Immunological evidence that non-carboxymethyllysine advanced glycation end-products are produced from short chain sugars and dicarbonyl compounds in vivo
Immunological evidence that non-carboxymethyllysine advanced glycation end-products are produced from short chain sugars and dicarbonyl compounds in vivo
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DOI：
10.1007/bf03401779
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发表时间：
2000-02-01
影响因子：
5.7
通讯作者：
Kameda, Y
中科院分区：
文献类型：
--
作者：
Takeuchi, M;Makita, Z;Kameda, Y
背景：导致晚期糖基化终产物（AGE）形成的美拉德反应在糖尿病患者血管病变的发病机制和衰老过程中起着重要作用。近年来，人们提出AGE不仅由葡萄糖产生，还由美拉德反应、糖的自氧化等葡萄糖代谢途径衍生的二羰基化合物产生。在这项研究中，我们开发了四种类型的非羧甲基赖氨酸（CML）抗age抗体，它们识别经短链糖和二羰基化合物孵育修饰的蛋白质。材料与方法：将兔血清白蛋白与甘油醛、乙醇醛、甲基乙二醛或乙二醛孵育制备age修饰的血清白蛋白。兔免疫后获得4种AGE特异性抗血清，对AGE修饰具有特异性。为了分离非cml AGE抗体（Ab） （non-CML AGE-Ab-2、-3、-4和-5），这些抗AGE抗血清在与四种AGE牛血清白蛋白（BSA）或CML-BSA偶联的基质上进行亲和层析。这些非cml AGE抗体用于研究糖尿病血液透析患者血清中AGE的含量。结果：免疫亲和层析获得的4种非cml AGE抗体均采用竞争性ELISA和免疫印迹法进行鉴定。非cml AGE-Ab-2与甘油醛或乙醇醛修饰的蛋白发生交叉反应。非cml的AGE-Ab-3和-Ab-4分别与乙醇醛和甲基乙二醛修饰的蛋白特异性交叉反应。Non-CML AGE-Ab-5与乙二醛、甲基乙二醛和乙醇醛修饰的蛋白发生交叉反应。糖尿病患者血清中检测到3种非cml AGE （AGE-2、-4、-5）为3个峰，表观分子量分别为200、1.15、0.85 kD；AGE-3为2个峰，表观分子量分别为200和0.85 kD。结论：不同类型的非cml AGE是由美拉德反应、糖自氧化和糖代谢形成的。这些抗体使我们能够在体内识别由美拉德反应产生的化合物。
Background: The Maillard reaction that leads to the formation of advanced glycation end-products (AGE) plays an important role in the pathogenesis of angiopathy in diabetic patients and in the aging process. Recently, it was proposed that AGE were not only created by glucose, but also by dicarbonyl compounds derived from the Maillard reaction, autoxidation of sugars and other metabolic pathways of glucose. In this study, we developed four types of non- carboxymethyllysine (CML) anti-AGE antibodies that recognized proteins modified by incubation with short chain sugars and dicarbonyl compounds.Materials and Methods: AGE-modified serum albumins were prepared by incubation of rabbit serum albumin with glyceraldehyde, glycolaldehyde, methylglyoxal or glyoxal. After immunization of rabbits, four types of AGE-specific antisera were obtained that were specific for the AGE modification. To separate non-CML AGE antibodies (Ab) (non-CML AGE-Ab-2, -3, -4, and -5), these anti-AGE antisera were subjected to affinity chromatography on a matrix coupled with four kinds of AGE bovine serum albumin (BSA) or CML-BSA. These non-CML AGE antibodies were used to investigate the AGE content of serum obtained from diabetic patients on hemodialysis.Results: Characterization of the four types of non-CML AGE antibodies obtained by immunoaffinity chromatography was performed by competitive ELISA and immunoblot analysis. Non-CML AGE-Ab-2 crossreacted with the protein modified by glyceraldehyde or glycolaldehyde. Non-CML AGE-Ab-3 and -Ab-4 specifically cross-reacted with protein modified by glycolaldehyde and methylglyoxal, respectively. Non-CML AGE-Ab-5 cross-reacted with protein modified with glyoxal as well as methylglyoxal and glycolaldehyde. Three kinds of non-CML AGE (AGE-2, -4, and -5) were detected in diabetic serum as three peaks with apparent molecular weights of 200, 1.15, and 0.85 kD; whereas, AGE-3 was detected as two peaks with apparent molecular weights of 200 and 0.85 kD.Conclusion: We propose that various types of non-CML AGE are formed by the Maillard reaction, sugar autoxidation and sugar metabolism. These antibodies enable us to identify such compounds created by the Maillard reaction in vivo.