摘要
目的探讨虾青素(AST)调节沉默信息调节因子1(SIRT1)/核因子E2相关因子2(Nrf2)信号通路对脂多糖(LPS)诱导的肾小管上皮细胞(HK-2细胞)铁死亡的影响。方法取生长较好的HK-2细胞进行分组,依次分为对照组、溶剂组(0.1%二甲基亚砜)、LPS组(1μg·mL^(-1)LPS)、AST组(1μg·mL^(-1)LPS+20μmol·L^(-1)AST)、AST+烟酰胺组(1μg·mL^(-1)LPS+20μmol L^(-1)AST+20μmol·L^(-1)烟酰胺)。用细胞计数试剂盒8(CCK-8)分析细胞活力;用流式细胞仪分析活性氧(ROS)水平;用试剂盒法检测胱甘肽过氧化物酶(GSH-Px)、铁离子、丙二醛(MDA)含量;用实时荧光定量聚合酶链反应(qRT-PCR)检测铁死亡标志谷胱甘肽过氧化物酶4(GPX4)、非糖基化的xCT(SLC7A11)mRNA表达水平;用蛋白质印迹(Western blot)法检测SIRT1、Nrf2、GPX4、SLC7A11蛋白表达。结果对照组、溶剂组、LPS组、AST组、AST+烟酰胺组细胞活力分别为(100.00±0.00)%、(94.27±4.44)%、(39.62±3.97)%、(69.09±6.91)%和(40.51±4.06)%,SLC7A11蛋白表达水平分别为0.95±0.10、1.02±0.11、0.26±0.03、0.88±0.09和0.45±0.05,GPX4蛋白表达水平分别为1.02±0.11、0.93±0.10、0.43±0.05、0.95±0.10和0.62±0.07,SIRT1蛋白表达水平分别为1.06±0.11、0.93±0.10、0.48±0.05、1.02±0.11和0.72±0.08,Nrf2蛋白表达水平分别为0.77±0.08、0.79±0.08、0.33±0.04、0.74±0.09和0.44±0.05,ROS表达水平分别为(216.33±21.64)%、(235.14±23.52)%、(1899.34±189.94)%、(549.32±54.94)%和(1216.23±121.63)%,MDA表达量分别为(6.63±0.67)、(6.75±0.68)、(15.82±1.59)、(9.09±0.91)和(13.96±1.40)ng·mL^(-1),铁离子含量分别为(2.74±0.28)、(3.04±0.31)、(16.72±1.68)、(8.41±0.85)和(14.26±1.43)ng·mL^(-1),LPS组上述指标与对照组比较,AST组与LPS组比较,AST+烟酰胺组与AST组比较,差异均有统计学意义(均P<0.05)。结论AST可通过SIRT1/Nrf2信号通路的活化抑制铁死亡,缓解LPS诱导的HK-2细胞损伤。
Objective To investigate the influence of astaxanthin(AST)on lipopolysaccharide(LPS)induced ferroptosis in renal tubular epithelial cells(HK-2 cells)by regulating silent information regulator 1(SIRT1)/nuclear factor E2 related factor 2(Nrf2)signaling pathway.Methods The well growing HK-2 cells were divided into control group,solvent group(dimethyl sulfoxide,0.1%),LPS group(1μg·mL^(-1) LPS),AST group(1μg·mL^(-1) LPS+20μmol·L^(-1) AST),and AST+nicotinamide group(1μg·mL^(-1) LPS+20μmol L^(-1) AST+20 mmol L^(-1) nicotinamide).Cell counting kit-8(CCK-8)was used to analyze cell viability;the level of reactive oxygen species(ROS)was analyzed by flow cytometry;the contents of glutathione peroxidase(GSH-Px),iron ion and malondialdehyde(MDA)were analyzed in turn with the kits;the mRNA expression levels of glutathione peroxidase 4(GPX4)and non-glycosylated xCT(SLC7A11)were detected by quantitative reverse transcription-polymerase chain reaction(qRT-PCR);Western blot was used to detect the expression of SIRT1,Nrf2,GPX4 and SLC7A11 proteins.Results The cell viability of the control group,solvent group,LPS group,AST group,and AST+nicotinamide group were(100.00±0.00)%,(94.27±4.44)%,(39.62±3.97)%,(69.09±6.91)%and(40.51±4.06)%,respectively;the expression levels of SLC7A11 protein were 0.95±0.10,1.02±0.11,0.26±0.03,0.88±0.09 and 0.45±0.05,respectively;the expression levels of GPX4 protein were 1.02±0.11,0.93±0.10,0.43±0.05,0.95±0.10 and 0.62±0.07,respectively;the expression levels of SIRT1 protein were 1.06±0.11,0.93±0.10,0.48±0.05,1.02±0.11 and 0.72±0.08,respectively;the expression levels of Nrf2 protein were 0.77±0.08,0.79±0.08,0.33±0.04,0.74±0.09 and 0.44±0.05,respectively;the expression levels of ROS were(216.33±21.64)%,(235.14±23.52)%,(1899.34±189.94)%,(549.32±54.94)%and(1216.23±121.63)%,respectively;the expression levels of MDA were(6.63±0.67),(6.75±0.68),(15.82±1.59),(9.09±0.91)and(13.96±1.40)ng·mL^(-1),respectively;the levels of iron ions were(2.74±0.28),(3.04±0.31),(16.72±1.
作者
孔春艳
李纪华
王艳丽
KONG Chun-yan;LI Ji-hua;WANG Yan-li(Department of Physiology,Henan Medical College,Zhengzhou 451191,Henan Province,China;Department of Urology,First People’s Hospital of Zhengzhou City,Zhengzhou 450000,Henan Province,China;Department of Pediatrics,South China Hospital Affiliated to Shenzhen University,Shenzhen 518100,Guangdong Province,China)
出处
《中国临床药理学杂志》
CAS
CSCD
北大核心
2023年第21期3092-3096,共5页
The Chinese Journal of Clinical Pharmacology
关键词
虾青素
沉默信息调节因子1/核因子E2相关因子2信号通路
脂多糖
肾小管上皮细胞
铁死亡
astaxanthin
silent information regulator 1/nuclear factor E2 related factor 2 signal pathway
lipopolysaccharide
renal tubular epithelial cells
ferroptosis
