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目的研究高游离脂肪酸(HFFA)血症致血管内皮细胞功能紊乱是否与全身氧化应激增强以及血管局部烟酰胺腺嘌呤二核苷酸磷酸(NADPH)氧化酶p47-phox亚基表达增强有关。方法正常6周龄雄性SD大鼠分为对照组(NC组,静脉输注生理盐水),HFFA组(输注脂肪乳及肝素),以及还原型谷胱甘肽(GSH)干预组(HFFA+GSH组,同时输注脂肪乳、肝素及GSH)。输液前后测血浆FFA水平及GSH/GSSG比值。输液后行超声多普勒检测股动脉内皮依赖性血管舒张功能(EDV)与非内皮依赖性血管舒张(NEDV)功能,实时荧光定量RT-PCR检测胸主动脉NADPH氧化酶p-47phox的mRNA水平,Western blot检测内皮细胞及平滑肌细胞p-47phox蛋白的表达。结果与NC组相比,HFFA组血浆FFA水平明显增高(P<0.05),GSH/GSSG比值下降(P<0.05),EDV明显降低(P<0.01);HF-FA+GSH组血浆GSH/GSSG比值介于HFFA组与NC组之间,EDV较HFFA组明显改善(P<0.05);NADPH氧化酶p-47phox mRNA及蛋白水平HFFA组较NC组明显增加,该增加趋势在HFFA+GSH组中受到明显抑制;p-47phox mRNA和蛋白水平与血浆GSH/GSSG比值呈正相关关系。结论外源性升高血浆FFA水平能诱导机体氧化应激,促进血管局部NADPH氧化酶P-47phox亚基基因的转录与翻译,加重血管局部氧化应激损伤。阻断HFFA血症导致的氧化应激能抑制主动脉NADPH氧化酶表达,部分恢复EDV,提示HFFA血症致血管内皮细胞功能受损与全身及血管局部氧化应激增强有关。
Objective To investigate whether dysfunction of vascular endothelial cells induced by high free fatty acid (HFFA) is associated with increased systemic oxidative stress and enhanced expression of the p47-phox subunit of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. Methods Normal 6-week-old male Sprague-Dawley rats were divided into control group (NC group, intravenous infusion of saline), HFFA group (trans fat emulsion and heparin), and GSH intervention group (HFFA + GSH group, while trans fat emulsion, heparin and GSH). Before and after infusion of plasma FFA levels and GSH / GSSG ratio. After transfusion, the endothelium-dependent vasodilation (FEV) and non-endothelium-dependent vasodilation (NEDV) in femoral artery were detected by Doppler ultrasound. The mRNA level of NADPH oxidase p-47phox in the thoracic aorta was detected by real-time fluorescence quantitative RT- Western blot was used to detect the expression of p-47phox protein in endothelial cells and smooth muscle cells. Results Compared with NC group, plasma FFA level was significantly increased in HFFA group (P <0.05), GSH / GSSG ratio was decreased (P <0.05), EDV was significantly decreased (P <0.01) (P <0.05). The levels of p-47phox mRNA and protein of NADPH oxidase in HFFA group were significantly higher than those in NC group (P <0.05). The increasing trend was in HFFA + GSH group Was significantly inhibited; p-47phox mRNA and protein levels and plasma GSH / GSSG ratio was positively correlated. Conclusions Exogenous elevated plasma FFA levels can induce oxidative stress in the body and promote the transcription and translation of the P-47phox subunit gene of NADPH oxidase, which aggravates the local oxidative stress injury in blood vessels. Blockade of HFFA-induced oxidative stress can inhibit aortic NADPH oxidase expression, partial recovery of EDV, suggesting that impaired HFFA vascular endothelial cell dysfunction and systemic and vascular local oxidative stress-related.