Department of Molecular Medicine and Surgery, Karolinska Institutet, Karolinska University Hospital (Solna), SE-171 76 Stockholm, Sweden.
Biochem Biophys Res Commun. 2012 May 4;421(2):335-42. doi: 10.1016/j.bbrc.2012.03.154. Epub 2012 Apr 7.
Aspirin may exert part of its antithrombotic effects through platelet-independent mechanisms. Diabetes is a condition in which the beneficial effects of aspirin are less prominent or absent - a phenomenon called "aspirin resistance". We investigated whether acetylation and glycation occur at specific sites in fibrinogen and if competition between glucose and aspirin in binding to fibrinogen occurs. Our hypothesis was that such competition might be one explanation to "aspirin resistance" in diabetes. After incubation of fibrinogen in vitro with aspirin (0.8 mM, 24 h) or glucose (100 mM, 5-10 days), we found 12 modified sites with mass spectrometric techniques. Acetylations in the α-chain: αK191, αK208, αK224, αK429, αK457, αK539, αK562, in the β-chain: βK233, and in the γ-chain: γK170 and γK273. Glycations were found at βK133 and γK75, alternatively γK85. Notably, the lysine 539 is a site involved in FXIII-mediated cross-linking of fibrin. With isotope labeling in vitro, using [(14)C-acetyl]salicylic acid and [(14)C]glucose, a labeling of 0.013-0.084 and 0.12-0.5 mol of acetylated and glycated adduct/mol fibrinogen, respectively, was found for clinically (12.9-100 μM aspirin) and physiologically (2-8 mM glucose) relevant plasma concentrations. No competition between acetylation and glycation could be demonstrated. Thus, fibrinogen is acetylated at several lysine residues, some of which are involved in the cross-linking of fibrinogen. This may mechanistically explain why aspirin facilitates fibrin degradation. We find no support for the idea that glycation of fibrin(ogen) interferes with acetylation of fibrinogen.
阿司匹林可能通过血小板非依赖性机制发挥部分抗血栓作用。糖尿病是一种阿司匹林的有益效果不明显或不存在的情况——这种现象称为“阿司匹林抵抗”。我们研究了纤维蛋白原中是否存在特定部位的乙酰化和糖化，以及葡萄糖和阿司匹林在与纤维蛋白原结合时是否存在竞争。我们的假设是，这种竞争可能是糖尿病中“阿司匹林抵抗”的一个解释。在体外将纤维蛋白原与阿司匹林（0.8 mM，24 小时）或葡萄糖（100 mM，5-10 天）孵育后，我们使用质谱技术发现了 12 个修饰位点。α 链上的乙酰化：αK191、αK208、αK224、αK429、αK457、αK539、αK562，β 链上的乙酰化：βK233，以及γ 链上的乙酰化：γK170 和 γK273。βK133 和γK75 或γK85 上发现了糖化。值得注意的是，赖氨酸 539 是 FXIII 介导的纤维蛋白交联的一个位点。通过体外同位素标记，使用[（14）C-乙酰]水杨酸和[（14）C]葡萄糖，发现临床（12.9-100 μM 阿司匹林）和生理（2-8 mM 葡萄糖）相关血浆浓度下纤维蛋白原的乙酰化和糖化加合物分别为 0.013-0.084 和 0.12-0.5 mol/mol。未发现乙酰化和糖化之间的竞争。因此，纤维蛋白原在几个赖氨酸残基上被乙酰化，其中一些残基参与纤维蛋白原的交联。这可能从机制上解释了为什么阿司匹林促进纤维蛋白降解。我们没有发现纤维蛋白原（原）糖化干扰纤维蛋白原乙酰化的证据。
