Zha Ruo-peng, Xu Wei, Wang Wen-yi, Dong Li, Wang Yi-ping
State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.
Acta Pharmacol Sin. 2007 Aug;28(8):1143-8. doi: 10.1111/j.1745-7254.2007.00595.x.
To investigate the effect of 3,5-dicaffeoylquinic acid (3,5-diCQA) on lipopolysaccharide (LPS)-induced injury in human dermal microvascular endothelial cells (HMEC-1).
The anti-oxidant effect was detected using the malondialdehyde (MDA) assay in a rat liver microsome model of lipid peroxidation. Cell viability was analyzed using the 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide assay. Cell lipid peroxide injury was measured by lactate dehydrogenase (LDH) release. Apoptotic cells were detected by flow cytometry, and confirmed by DNA fragmentation analysis. Caspase-3 activity was measured using a specific assay kit. The level of intracellular reactive oxygen species (ROS) was determined by flow cytometry with a 2,7-dichlorodihydro-fluorescein diacetate fluorescence probe.
The exposure of microsomes to ascorbate-Fe2+ resulted in lipoperoxidation according to an increase in the level of MDA. MDA formation decreased in a dose-dependent manner on treatment with 5, 10, or 50 micromol/L 3,5-diCQA. Treatment with LPS for 16 h resulted in a 60% decrease in cell viability and an increase in LDH release from 47.6% to 61.5%. DNA laddering was observed by agarose gel electrophoresis. The level of apoptotic cells peaked at 27% after treatment with LPS for 12 h. Following treatment with LPS for 12 h, intracellular ROS and caspase-3 activity increased. Pretreatment with 3,5-diCQA at 5, 10, or 50 micromol/L for 1 h attenuated LPS-mediated endothelial cell injury. The anti-apoptotic action of 3,5-diCQA was partially dependent on its capacity for anti-oxidation and the suppression of caspase-3 activity.
3,5-diCQA displays anti-oxidative and anti-apoptotic activity in HMEC-1 due to scavenging of intracellular ROS induced by LPS, and the suppression of caspase-3 activity.
研究3,5-二咖啡酰奎宁酸（3,5-diCQA）对脂多糖（LPS）诱导的人皮肤微血管内皮细胞（HMEC-1）损伤的影响。
在大鼠肝微粒体脂质过氧化模型中，使用丙二醛（MDA）检测法检测抗氧化作用。采用3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐检测法分析细胞活力。通过乳酸脱氢酶（LDH）释放来测定细胞脂质过氧化损伤。通过流式细胞术检测凋亡细胞，并通过DNA片段化分析进行确认。使用特定检测试剂盒测量半胱天冬酶-3活性。用2,7-二氯二氢荧光素二乙酸荧光探针通过流式细胞术测定细胞内活性氧（ROS）水平。
微粒体暴露于抗坏血酸-Fe2+导致脂质过氧化，MDA水平升高。用5、10或50 μmol/L 3,5-diCQA处理后，MDA形成呈剂量依赖性降低。用LPS处理16小时导致细胞活力降低60%，LDH释放从47.6%增加到61.5%。通过琼脂糖凝胶电泳观察到DNA梯状条带。用LPS处理12小时后，凋亡细胞水平在27%达到峰值。用LPS处理12小时后，细胞内ROS和半胱天冬酶-3活性增加。用5、10或50 μmol/L 3,5-diCQA预处理1小时可减轻LPS介导的内皮细胞损伤。3,5-diCQA的抗凋亡作用部分取决于其抗氧化能力和对半胱天冬酶-3活性的抑制。
3,5-diCQA在HMEC-1中表现出抗氧化和抗凋亡活性，这归因于其清除LPS诱导的细胞内ROS以及抑制半胱天冬酶-3活性。
