摘要
目的分析高尿酸血症是否通过诱导亮氨酸富集的核苷酸结合寡聚结构域3蛋白(NLRP3)炎症小体活化蛋白质-gasdermin D(GSDMD)导致内皮细胞焦亡,观察尿酸对血管内皮细胞的作用机制。方法选择ATCC细胞库的120份人脐静脉内皮细胞(HUVEC),制备HUVEC培养基、内皮细胞(ECM)培养基,培养HUVEC细胞,按随机数表法将培养成功的120份HUVEC细胞分为3组,分别为HUVEC+ECM+500μmol/L浓度尿酸,HUVEC+ECM+1000μmol/L浓度尿酸,HUVEC+ECM+1500μmol/L浓度尿酸。分别将各浓度尿酸置于5%CO295%空气细胞培养箱内分别培养24 h、48 h、72 h。计算不同浓度内皮细胞活力下降率;采用Western blot与免疫荧光测定细胞caspase-3、caspase-4、NLRP3及GSDMD表达。结果在处理12 h的实验模型内,仅有1500μmol/L浓度的尿酸能够明显降低HUVECS的细胞活力,细胞活力下降率为50.00%;在处理24 h的实验模型内,各组细胞活力下降率分别为25.00%、65.00%、80.00%,随着尿酸浓度增加,HUVECS细胞的细胞活力随之下降,各浓度比较(P<0.05);随着尿酸浓度的升高,内皮细胞caspase-3、caspase-4、NLRP3及GSDMD表达逐渐上升(P<0.05)。结论高尿酸血症可引起内皮细胞NLRP3炎症小体激活,诱发细胞炎症反应,通过介导细胞中GSDMD活性,从而引起内皮细胞焦亡的发生。
Objective To analyze whether hyperuricemia induces nucleotide-binding oligomerization domainlike receptor protein 3(NLRP3)inflammasome activating protein-gasdermin D(GSDMD)leading to the pyroptosis of endothelial cell,and to observe the mechanism of action of uric acid on vascular endothelial cell.Methods 120 human umbilical vein endothelial cells(HUVEC)in ATCC cell bank were selected,the HUVEC medium and endothelial cell medium(ECM)were prepared.The HUVEC were cultured and divided into 3 groups randomly with 500μmol/L uric acid,1000μmol/L uric acid and 1500μmol/L uric acid added for culture.Cells in each group were placed in 5%CO2 and 95%air cell incubators for 24 h,48 h,and 72 h,respectively.The decrease rate of endothelial cell viability under different uric acid concentrations was calculated;the expression of caspase-3,caspase-4,NLRP3 and GSDMD was determined by Western blot and immunofluorescence.Results After adding uric acid for 12 hours,only 1500μmol/L concentration of uric acid can significantly reduce the cell viability of HUVECS,and the rate of cell viability decline is 50.00%.Twenty-four hours after the addition of uric acid,the cell viability reduction rates of each group were 25.00%,65.00%,and 80.00%,respectively.As the concentration of uric acid increased,the cell viability of HUVECS cells decreased accordingly,and the difference between the concentrations was statistically significant(P<0.05).With the increase of uric acid concentration,the expression of caspase-3,caspase-4,NLRP3 and GSDMD gradually increased(P<0.05).Conclusion Hyperuricemia can cause the activation of NLRP3 inflammatory bodies of endothelial cells,inducing inflammatory reaction of cells,by mediating the activity of GSDMD in cells,thereby causing the occurrence of pyrolysis of endothelial cells.
作者
蒋云
沈成兴
潘静薇
王占成
宋艳秋
Jiang Yun;Shen Chengxing;Pan Jingwei;Wang Zhancheng;Song Yanqiu(Department of Cardiovascular Medicine,Shanghai Sixth People's Hospital,Shanghai 200235,China;不详)
出处
《中国循证心血管医学杂志》
2020年第5期612-615,共4页
Chinese Journal of Evidence-Based Cardiovascular Medicine
