Wu C C, Hsieh C W, Lai P H, Lin J B, Liu Y C, Wung B S
Institute of Biotechnology, National Chiayi University, Chiayi, Taiwan.
Toxicol Appl Pharmacol. 2006 Aug 1;214(3):244-52. doi: 10.1016/j.taap.2005.12.013. Epub 2006 Feb 15.
Acrolein is a highly electrophilic alpha,beta-unsaturated aldehyde that is present in cigarette smoke. Heme oxygenase-1 (HO-1) is a cytoprotective enzyme activated by various such electrophilic compounds. In this study, the regulatory effects of acrolein upon the expression of HO-1 were investigated in endothelial cells (ECs). We demonstrate that acrolein induces the elevation of HO-1 protein levels, and subsequent enzyme activity, at non-cytotoxic concentrations. An additional alpha,beta-unsaturated aldehyde, cinnamaldehyde, was also found to increase HO-1 expression and have less cytotoxicity than acrolein. Moreover, acrolein-mediated HO-1 induction is abrogated in the presence of actinomycin D and cycloheximide. Nrf2 is a transcription factor involved in the induction of HO-1 through an antioxidant response element (ARE) in the promoter region of the HO-1 gene. We show that acrolein induces Nrf2 translocation and ARE-luciferase reporter activity. Acrolein was also found to induce the production of both superoxide and H2O2 at levels greater than 100 microM. However, with the exception of NAC, no antioxidant generated any effect upon acrolein-dependent HO-1 expression in ECs. Our present findings suggest that reactive oxygen species (ROS) may not be a major modulator for HO-1 induction. Using buthionine sulfoximine to deplete the intracellular GSH levels further enhanced the effects of acrolein. We also found that cellular GSH level was rapidly reduced after both 10 and 100 microM acrolein treatment. However, after 6 h of exposure to ECs, only 10 microM acrolein treatment increases GSH level. In addition, only the JNK inhibitor SP600125 and tyrosine kinase inhibitor genistein had any significant inhibitory impact upon the upregulation of HO-1 by acrolein. Pretreatment with a range of other PI3 kinase inhibitors, including wortmannin and LY294002, showed no effects. Hence, we show in our current experiments that a sublethal concentration of acrolein is in fact a novel HO-1 inducer, and we further identify the principal underlying mechanisms involved in this process.
丙烯醛是一种高亲电性的α,β-不饱和醛，存在于香烟烟雾中。血红素加氧酶-1（HO-1）是一种由各种此类亲电化合物激活的细胞保护酶。在本研究中，我们在内皮细胞（ECs）中研究了丙烯醛对HO-1表达的调节作用。我们证明，在无细胞毒性浓度下，丙烯醛可诱导HO-1蛋白水平升高及随后的酶活性增强。另一种α,β-不饱和醛肉桂醛也被发现可增加HO-1表达，且细胞毒性比丙烯醛小。此外，在放线菌素D和环己酰亚胺存在的情况下，丙烯醛介导的HO-1诱导作用被消除。Nrf2是一种转录因子，通过HO-1基因启动子区域的抗氧化反应元件（ARE）参与HO-1的诱导。我们发现丙烯醛可诱导Nrf2易位和ARE-荧光素酶报告基因活性。还发现丙烯醛在浓度大于100 microM时可诱导超氧化物和H2O2的产生。然而，除了NAC外，没有抗氧化剂对ECs中丙烯醛依赖的HO-1表达产生任何影响。我们目前的研究结果表明，活性氧（ROS）可能不是HO-1诱导的主要调节因子。使用丁硫氨酸亚砜胺耗尽细胞内谷胱甘肽水平进一步增强了丙烯醛的作用。我们还发现，在10 microM和100 microM丙烯醛处理后，细胞内谷胱甘肽水平迅速降低。然而，在ECs暴露6小时后，只有10 microM丙烯醛处理可增加谷胱甘肽水平。此外，只有JNK抑制剂SP600125和酪氨酸激酶抑制剂染料木黄酮对丙烯醛上调HO-1有显著抑制作用。用一系列其他PI3激酶抑制剂（包括渥曼青霉素和LY294002）预处理没有效果。因此，我们在当前实验中表明，亚致死浓度的丙烯醛实际上是一种新型的HO-1诱导剂，并且我们进一步确定了该过程中涉及的主要潜在机制。
