Hydralazine inhibits rapid acrolein-induced protein oligomerization: Role of aldehyde scavenging and adduct trapping in cross-link blocking and cytoprotection
Hydralazine inhibits rapid acrolein-induced protein oligomerization: Role of aldehyde scavenging and adduct trapping in cross-link blocking and cytoprotection
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DOI：
10.1124/mol.105.018168
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发表时间：
2006-03-01
影响因子：
3.6
通讯作者：
Pyke, SM
中科院分区：
文献类型：
--
作者：
Burcham, PC;Pyke, SM
肼苯哒嗪强烈抑制丙烯醛的毒性，丙烯醛是一种导致许多健康障碍的反应性醛。至少有两种机制可能是细胞保护的基础，这两种机制都涉及肼苯哒嗪所具有的亲核肼。在最简单的情况下，肼苯哒嗪直接清除游离丙烯醛，降低细胞内丙烯醛的可用性，从而抑制大分子的加合。在第二种“加合物捕获”机制中，药物与细胞蛋白中的丙烯醛衍生的迈克尔加合物形成腙，防止可能引发细胞死亡的加合蛋白的次级反应。为了确定最重要的机制，我们探讨了这两个途径在小鼠肝细胞中毒的丙烯醛前体烯丙醇。肼苯哒嗪对细胞蛋白质中的强烈浓度依赖性加合物捕获作用伴随着毒性抑制。然而，肼苯哒嗪的保护浓度并没有改变细胞外游离丙烯醛水平，细胞谷胱甘肽损失，或蛋白质羰基化，这表明细胞保护不是由于细胞内丙烯醛的可用性最小化。为了探索加合物捕获可能赋予细胞保护的方式，检查肼苯哒嗪对丙烯醛诱导的蛋白质交联的影响。使用牛胰腺核糖核酸酶A作为模型蛋白质，丙烯醛引起快速的时间和浓度依赖性交联，与二聚化的蛋白质在45分钟内开始蛋白质修饰检测。赖氨酸加合在单体蛋白之前的低聚物的外观，而蛋白质胺基团的还原甲基化取消加合和寡聚化。肼苯哒嗪抑制交联，如果加入后30分钟开始丙烯醛暴露，但无效，如果加入后90分钟的延迟。肼苯哒嗪对交联的抑制作用与加合物捕获密切相关。这些发现表明，交联阻断可能有助于肼苯哒嗪的细胞保护。
Hydralazine strongly suppresses the toxicity of acrolein, a reactive aldehyde that contributes to numerous health disorders. At least two mechanisms may underlie the cytoprotection, both of which involve the nucleophilic hydrazine possessed by hydralazine. Under the simplest scenario, hydralazine directly scavenges free acrolein, decreasing intracellular acrolein availability and thereby suppressing macromolecular adduction. In a second "adduct-trapping" mechanism, the drug forms hydrazones with acrolein-derived Michael adducts in cell proteins, preventing secondary reactions of adducted proteins that may trigger cell death. To identify the most important mechanism, we explored these two pathways in mouse hepatocytes poisoned with the acrolein precursor allyl alcohol. Intense concentration-dependent adduct- trapping in cell proteins accompanied the suppression of toxicity by hydralazine. However, protective concentrations of hydralazine did not alter extracellular free acrolein levels, cellular glutathione loss, or protein carbonylation, suggesting that the cytoprotection is not due to minimization of intracellular acrolein availability. To explore ways whereby adduct-trapping might confer cytoprotection, the effect of hydralazine on acrolein-induced protein crosslinking was examined. Using bovine pancreas ribonuclease A as a model protein, acrolein caused rapid time-and concentration-dependent cross-linking, with dimerized protein detectable within 45 min of commencing protein modification. Lysine adduction in monomeric protein preceded the appearance of oligomers, whereas reductive methylation of protein amine groups abolished both adduction and oligomerization. Hydralazine inhibited cross-linking if added 30 min after commencing acrolein exposure but was ineffective if added after a 90-min delay. Adduct-trapping closely accompanied the inhibition of cross-linking by hydralazine. These findings suggest that cross-link blocking may contribute to hydralazine cytoprotection.