Croons Valerie, Martinet Wim, Herman Arnold G, De Meyer Guido R Y
Division of Pharmacology, University of Antwerp, Universiteitsplein 1, B-2610 Wilrijk, Belgium.
J Pharmacol Exp Ther. 2008 Jun;325(3):824-32. doi: 10.1124/jpet.107.132944. Epub 2008 Mar 5.
Recent evidence indicates that the protein synthesis inhibitor cycloheximide triggers selective macrophage death in rabbit atheroma-like lesions without affecting smooth muscle cells (SMCs) or the endothelium, thereby favoring a stable plaque phenotype. In this study, we report that puromycin, a protein synthesis inhibitor with a different mode of action but with similar ability to inhibit de novo protein synthesis, did not reveal plaque-stabilizing effects. The macrophage and the SMC content readily decreased in puromycin-treated atheroma-like lesions in rabbit carotid arteries. Moreover, puromycin induced apoptosis in macrophages and SMCs in vitro. Puromycin-treated SMCs showed signs of endoplasmic reticulum (ER) stress, as demonstrated by CCAAT/enhancer-binding protein homologous protein (CHOP) protein expression, splicing of X-box-binding protein 1 mRNA, and phosphorylation of eukaryotic translation initiation factor 2alpha. The ER stress inducer thapsigargin up-regulated CHOP protein expression in SMCs without affecting their viability, indicating that ER stress not necessarily results in cell death. Puromycin, but not thapsigargin, activated the ER stress-related caspase-12. Treatment of SMCs with a combination of cycloheximide and puromycin inhibited ER stress and partially improved SMC viability. In addition, puromycin, but not cycloheximide or thapsigargin, induced intracellular accumulation of polyubiquitinated proteins in SMCs, whereas the proteasome function was not affected. Taken together, puromycin, in contrast to cycloheximide, induces SMC apoptosis, thereby favoring an unstable plaque phenotype. SMC death upon puromycin treatment could only be partially prevented by cycloheximide, which completely blocked ER stress. However, other or additional mechanisms, such as increased polyubiquitination of proteins, might be involved in puromycin-induced SMC death.
最近有证据表明，蛋白质合成抑制剂放线菌酮可在兔动脉粥样硬化样病变中引发选择性巨噬细胞死亡，而不影响平滑肌细胞（SMC）或内皮细胞，从而有利于形成稳定的斑块表型。在本研究中，我们报告称，嘌呤霉素是一种蛋白质合成抑制剂，其作用方式不同，但抑制从头蛋白质合成的能力相似，并未显示出斑块稳定作用。在嘌呤霉素处理的兔颈动脉粥样硬化样病变中，巨噬细胞和SMC含量迅速减少。此外，嘌呤霉素在体外可诱导巨噬细胞和SMC凋亡。嘌呤霉素处理的SMC表现出内质网（ER）应激迹象，CCAAT/增强子结合蛋白同源蛋白（CHOP）蛋白表达、X盒结合蛋白1 mRNA剪接以及真核翻译起始因子2α磷酸化均证明了这一点。ER应激诱导剂毒胡萝卜素上调了SMC中CHOP蛋白的表达，但不影响其活力，这表明ER应激不一定导致细胞死亡。嘌呤霉素而非毒胡萝卜素激活了与ER应激相关的半胱天冬酶-12。用放线菌酮和嘌呤霉素联合处理SMC可抑制ER应激并部分改善SMC活力。此外，嘌呤霉素而非放线菌酮或毒胡萝卜素可诱导SMC中多聚泛素化蛋白的细胞内积累，而蛋白酶体功能未受影响。综上所述，与放线菌酮相反，嘌呤霉素可诱导SMC凋亡，从而有利于形成不稳定的斑块表型。放线菌酮可部分预防嘌呤霉素处理导致的SMC死亡，它能完全阻断ER应激。然而，其他或额外的机制，如蛋白质多聚泛素化增加，可能参与了嘌呤霉素诱导的SMC死亡。
