槲皮素对氧化低密度脂蛋白诱导的血管平滑肌细胞炎症反应的影响

Effects of Quercetin on Inflammatory Reactions Induced by Oxidized Low Density Lipoprotein in Vascular Smooth Muscle Cells

目的 :研究槲皮素对氧化低密度脂蛋白(ox-LDL)刺激血管平滑肌细胞(VSMCs)炎症因子分泌的影响。方法:MTT法测定不同浓度槲皮素对VSMCs细胞活性的影响。将细胞分为五组:A组(对照组)、B组(ox-LDL干预组)、C组(ox-LDL+槲皮素50μmol/L)、D组(ox-LDL+槲皮素100μmol/L)和E组(ox-LDL+槲皮素200μmol/L),以上各组ox-LDL浓度均为100μg/m L。酶联免疫吸附法(ELISA)测定各组细胞上清液单核细胞趋化蛋白(MCP)-1和肿瘤坏死因子(TNF)-α分泌水平。荧光定量聚合酶链式反应(real-time RT-PCR)法测定各组细胞Toll样受体4(TLR4)、髓样分化因子88(Myd88)m RNA的表达。Western-blot法检测TLR4、Myd88及核内核转录因子(NF)-κB(p65)的蛋白表达。结果:槲皮素在0~250μmol/L浓度范围内未显示明显的细胞毒性。同ox-LDL干预组相比,槲皮素可以有效抑制VSMCs细胞MCP-1和TNF-α的分泌、减少TLR4、My D88 m RNA和蛋白的表达及抑制ox-LDL诱导的NF-κB(p65)核转位,并呈剂量依赖性(P<0.01)。使用TLR4和NF-κB(p65)抑制剂预处理也起到了与槲皮素类似的抗炎作用。结论 :槲皮素能够减少ox-LDL诱导VSMCs细胞炎症因子分泌,这种作用可能依赖于其对TLR4介导的炎症通路的调控而得以实现的。

Objective:To investigate the effects of quercetin(Que) on oxidized low density lipoprotein(ox-LDL)-induced cytokines secretion in vascular smooth muscle cells(VSMCs). Methods:The effect of quercetin with different concentrations on the activity of VSMCs was determined by MTT method. VSMCs were divided into 5 groups: A group(control group), B group(ox-LDL group), C group(ox-LDL+Que 50 μmol/L), D group(ox-LDL+Que 100 μmol/L) and E group(ox-LDL+Que 200 μmol/L), and the concentration of ox-LDL in all groups was 100 μg/m L.The levels of monocyte chemoattractant protein(MCP-1) and Tumor necrosis factor(TNF)-α in cell supernatant in each group was determined by ELISA. m RNA expression of Toll-like receptor 4(TLR4) and myeloid differentiation factor 88(Myd88) was determined by fluorescent quantitative PCR(real-time RT-PCR). The protein expression of TLR-4, Myd88 and nuclear transcription factor(NF)-κB(p65) was determined by Western-blot method. Results:Quercetin did not show significant cytotoxicity in the range of 0~250 mol/L. Quercetin can significantly inhibit MCP-1 and TNF-α secretion in VSMCs, reduce TLR4, My D88 m RNA and protein expression and inhibit ox-LDLinduced NF-κB(p65) nuclear transposition compared with ox-LDL group, showing dose-dependent manner(P<0.01). Pretreatment of TLR4 and TNF-α(p65) inhibitor has similar effect to quercetin. Conclusion:Quercetin can reduce oxLDL-induced cytokines secretion in VSMCs, which may depend on regulating TLR4-mediated signaling pathway.