Patricia M K, Natarajan R, Dooley A N, Hernandez F, Gu J L, Berliner J A, Rossi J J, Nadler J L, Meidell R S, Hedrick C C
Division of Cardiology, Department of Experimental Pathology, University of California Los Angeles, USA.
Circ Res. 2001 Apr 13;88(7):659-65. doi: 10.1161/hh0701.088838.
The lipoxygenase (LO) pathway has been implicated as an important mediator of chronic glucose and platelet-derived growth factor (PDGF)-induced effects in the vascular system. Endothelial cells treated with 12LO products or cultured in high glucose showed enhanced monocyte adhesion, an important step in atherogenesis. We have previously reported that PDGF increased HETE levels in porcine aortic smooth muscle cells. Although several pharmacological inhibitors to the LO pathway are available, most lack specificity and may harbor undesirable side effects. Therefore, we developed a recombinant adenovirus expressing a hammerhead ribozyme (AdRZ) targeted against the porcine leukocyte-type 12LO mRNA to investigate the involvement of LO in glucose- and PDGF-mediated effects in vascular cells. Infection of porcine aortic endothelial cells with AdRZ reduced the level of glucose-enhanced 12LO mRNA expression as determined by quantitative, real-time reverse transcriptase-polymerase chain reaction. Reverse-phase HPLC and RIA analysis also revealed a corresponding decrease in glucose-stimulated 12HETE production in both the cellular and supernatant fractions. In the ribozyme-treated porcine aortic endothelial cells, there was marked inhibition of high glucose-stimulated monocyte adhesion. Infection with AdRZ also reduced PDGF-induced porcine aortic smooth muscle cell migration by approximately 50%. These studies demonstrate the efficacy of recombinant adenovirus expressing 12LO ribozyme in studying the effects of 12LO in vascular wall cells. They document an important role for the 12LO pathway in regulating inflammatory changes in endothelial cells and smooth muscle cells.
脂氧合酶（LO）途径被认为是慢性葡萄糖和血小板衍生生长因子（PDGF）诱导的血管系统效应的重要介质。用12-LO产物处理或在高糖环境中培养的内皮细胞显示单核细胞粘附增强，这是动脉粥样硬化形成中的重要一步。我们之前报道过PDGF会增加猪主动脉平滑肌细胞中的HETE水平。尽管有几种针对LO途径的药理抑制剂，但大多数缺乏特异性，可能存在不良副作用。因此，我们构建了一种表达针对猪白细胞型12-LO mRNA的锤头状核酶的重组腺病毒（AdRZ），以研究LO在血管细胞中葡萄糖和PDGF介导的效应中的作用。用AdRZ感染猪主动脉内皮细胞后，通过定量实时逆转录聚合酶链反应测定，葡萄糖增强的12-LO mRNA表达水平降低。反相高效液相色谱和放射免疫分析还显示，细胞和上清液部分中葡萄糖刺激的12-HETE产生相应减少。在核酶处理的猪主动脉内皮细胞中，高糖刺激的单核细胞粘附受到明显抑制。用AdRZ感染还使PDGF诱导的猪主动脉平滑肌细胞迁移减少了约50%。这些研究证明了表达12-LO核酶的重组腺病毒在研究12-LO对血管壁细胞作用方面的有效性。它们证明了12-LO途径在调节内皮细胞和平滑肌细胞炎症变化中的重要作用。
