丹酚酸A通过调节氧化应激抑制LPS诱导的人脐静脉血管内皮细胞NLRP3炎性小体的表达  

Salvianolic acid a inhibits LPS-induced human umbilical vein endothelial cells by regulating oxidative stress expression of NLRP3 inflammasome

作　　者：张淼 邓悦[2] 于克英 常丽萍[2] 田腾辉 石锐[2] 邵笑 ZHANG Miao;DENG Yue;YU Keying;CHANG Liping;TIAN Tenghui;SHI Rui;SHAO Xiao(Changchun University of Chinese Medicine,Changchun 130117,China)

机构地区：[1]长春中医药大学 [2]长春中医药大学附属医院心病中心

出　　处：《环球中医药》2022年第3期390-395,共6页Global Traditional Chinese Medicine

基　　金：吉林省教育厅科学技术研究项目(JJKH20210953KJ);国家重点研发计划(2019YFC1709901)。

摘　　要：目的探讨丹酚酸A(salvianolic acid A,Sal A)对脂多糖(lipopolysaccharide,LPS)诱导人脐静脉血管内皮细胞(human umbilical vein endothelial cells,HUVECs)氧化应激及炎症的抑制作用。方法脂多糖建立细胞损伤模型后,将细胞分为对照组、模型组(LPS 1μg/mL)、SalA低剂量组(LPS 1μg/mL+Sal A 6.25μmol/L)、Sal A中剂量组(LPS 1μg/mL+Sal A 12.5μmol/L)及Sal A高剂量组(LPS 1μg/mL+Sal A 25μmol/L)。采用CCK-8法检测HUVECs的活性;酶联免疫吸附法检测Sal A对炎性因子白细胞介素1β(interleukin-1β,IL-1β)、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)表达的影响;活性氧检测剂盒检测HUVECs中活性氧(reactive oxygen species,ROS)的含量;实时荧光定量PCR检测HUVECs中受体蛋白含NLP家族Pyrin域蛋白3重组蛋白(recombinant NLR Family Pyrin domain containing protein 3,NLRP3)、凋亡相关斑点样蛋白(adaptor protein apotosis speck-like protein containing CARD,ASC)半胱氨酸天冬氨酸蛋白酶-1(Caspase-1)、白细胞介素(interleukin,IL)-18、IL-1β的基因表达。蛋白免疫印迹法检测NLRP3、ASC蛋白表达。结果(1)给予LPS刺激后,与对照组比较,模型组细胞活性显著下降(P<0.05),与模型组相比,Sal A各组细胞存活率在一定范围内随药物浓度的递增而升高(P<0.05);(2)与对照组比较,模型组细胞ROS含量显著升高(P<0.05),与模型组比较,Sal A低、中、高剂量组ROS含量显著下降(P<0.05);(3)与对照组比较,HUVECs中NLRP3炎性小体相关指标NLRP3、ASC、Caspase-1表达显著升高(P<0.05),Sal A低、中、高剂量组与模型组比较均显著下降(P<0.05);(4)HUVECs中的炎性因子IL-1β、IL-18、TNF-α在给予LPS刺激后与对照组比较显著升高(P<0.05),给予Sal A后与模型组比较均有所下降(P<0.05)。结论Sal A对LPS诱导的HUVECs损伤具有明显的保护作用,其作用机制可能是通过降低细胞中ROS并抑制NLRP3炎性小体的表达得以实现。Objective To investigate the inhibitory effect of salvianolic acid A(Sal A)on lipopolysaccharide(LPS)-induced oxidative stress and inflammation in human umbilical vein endothelial cells(HUVECs).Methods After the cell injury model established by LPS,the cells were divided into the control group,the model group(LPS 1μg/mL),the low-dose group(LPS 1μg/mL+Sal A 6.25μmol/L),the medium-dose group(LPS 1μg/mL+Sal A 12.5μmol/L)and the high-dose group(LPS 1μg/mL+Sal A 25μmol/L).The CCK-8 method was used to detect the activity of HUVECs,and the enzyme-linked immunosorbent method was used to detect the effects of Sal A on the expressions of inflammatory factors interleukin-1β(IL-1β)and tumor necrosis factor-α(TNF-α),the ROS kit detects the content of reactive oxygen species(reactive oxygen species,ROS)in HUVECs,and Real-time Quantitative polymerase chain reaction(RT-PCR)was used to detect the gene expression of receptor protein NLRP3 and apoptosis-related speck-like protein containing CARD(ASC),Caspase-1,interleukin-1β(IL-1β),interleukin-18(IL-18)in HUVECs.Western blot was performed to detect NLRP3,ASC protein expression.Results(1)After LPS stimulated,cell activity in the model group was significantly decreased compared with the control group(P<0.05),and cell survival in the Sal A group increased with increasing drug concentration within a certain range compared with the model group(P<0.05);(2)compared with the control group,cell ROS content was significantly increased in the model group(P<0.05),and the low-dose group given Sal A ROS content decreased(P<0.05),while the content of ROS in Sal A medium and high dose groups was significantly decreased(P<0.05);(3)after LPS stimulation,NLRP3 inflammatory vesicle-related indexes NLRP3,ASC gene and protein expression were significantly increased in HUVECs compared with the control group(P<0.05),and both Sal A group were significantly decreased compared with the model group(P<0.05);(4)inflammatory factors IL-1β,IL-18,and TNF-αin HUVECs were significantly elevated after

关 键 词：丹酚酸A 内皮细胞 氧化应激 NLRP3炎性小体 动脉粥样硬化 

分 类 号：R285.5[医药卫生—中药学]