Du Youjia, Taylor Carla G, Aukema Harold M, Zahradka Peter
Department of Physiology & Pathophysiology, University of Manitoba, Winnipeg, Manitoba, Canada; Canadian Centre for Agri-Food Research in Health and Medicine, St. Boniface Hospital Albrechtsen Research Centre, Winnipeg, Manitoba, Canada.
Department of Physiology & Pathophysiology, University of Manitoba, Winnipeg, Manitoba, Canada; Department of Food and Human Nutritional Sciences, University of Manitoba, Winnipeg, Manitoba, Canada; Canadian Centre for Agri-Food Research in Health and Medicine, St. Boniface Hospital Albrechtsen Research Centre, Winnipeg, Manitoba, Canada.
Biochim Biophys Acta Mol Cell Biol Lipids. 2022 Jul;1867(7):159156. doi: 10.1016/j.bbalip.2022.159156. Epub 2022 Mar 30.
Oxylipins are oxygenated derivatives of polyunsaturated fatty acids, generated by COX, LOX and CYP enzymes, that regulate various aspects of endothelial cell physiology. Although 15-LOX and its products are positively associated with atherosclerosis, the relevant mechanisms have not been explored. The current study examined the effects of PD146176 (PD), a putative 15-LOX inhibitor, on EA.hy926 endothelial cell functions in the growing and confluent states. The effects of PD on endothelial cell oxylipin production (profiled by LC/MS/MS), cell viability, proliferation, eNOS activity, ICAM-1 and VE-cadherin levels were assessed. The contribution of signaling pathways relevant to endothelial function (p38 MAPK, Akt, PPARα) were also investigated. PD treatment for 30 min did not block formation of individual 15-LOX oxylipins, but 20 μM PD stimulated the accumulation of total LOX and COX products, while reducing several individual CYP products generated by epoxygenase. At 20 μM, the accumulated total oxylipins were primarily LOX-derived (86%) followed by COX (12%) and CYP (2%). PD altered cell functions by upregulating p38 MAPK and PPARα and downregulating Akt in a dose-dependent fashion. These observations suggest a link between PD-induced changes in oxylipins and altered endothelial cell functions via specific signaling pathways. In conclusion, the results of this study imply that PD does not function as a 15-LOX inhibitor in EA.hy926 endothelial cells, and instead inhibits CYP epoxygenase. These findings suggest that the cellular function changes induced by PD may be contingent upon its ability to modulate total oxylipin production, particularly by the LOX and CYP families.
氧化脂质是多不饱和脂肪酸的氧化衍生物，由环氧化酶（COX）、脂氧合酶（LOX）和细胞色素P450（CYP）酶生成，调节内皮细胞生理的各个方面。尽管15-脂氧合酶及其产物与动脉粥样硬化呈正相关，但相关机制尚未得到探索。本研究检测了一种假定的15-脂氧合酶抑制剂PD146176（PD）对生长状态和汇合状态的EA.hy926内皮细胞功能的影响。评估了PD对内皮细胞氧化脂质生成（通过液相色谱/串联质谱分析）、细胞活力、增殖、内皮型一氧化氮合酶（eNOS）活性、细胞间黏附分子-1（ICAM-1）和血管内皮钙黏蛋白（VE-钙黏蛋白）水平的影响。还研究了与内皮功能相关的信号通路（p38丝裂原活化蛋白激酶（p38 MAPK）、蛋白激酶B（Akt）、过氧化物酶体增殖物激活受体α（PPARα））的作用。PD处理30分钟并未阻断单个15-脂氧合酶氧化脂质的形成，但20μM的PD刺激了总脂氧合酶和环氧化酶产物的积累，同时减少了由环氧化酶产生的几种单个细胞色素P450产物。在20μM时，积累的总氧化脂质主要来源于脂氧合酶（86%），其次是环氧化酶（12%）和细胞色素P450（2%）。PD通过剂量依赖性地上调p38 MAPK和PPARα以及下调Akt来改变细胞功能。这些观察结果表明，PD诱导的氧化脂质变化与通过特定信号通路改变的内皮细胞功能之间存在联系。总之，本研究结果表明，PD在EA.hy926内皮细胞中并非作为15-脂氧合酶抑制剂发挥作用，而是抑制细胞色素P450环氧化酶。这些发现表明，PD诱导的细胞功能变化可能取决于其调节总氧化脂质生成的能力，特别是脂氧合酶和细胞色素P450家族的能力。
