中国兽医学报2024,Vol.44Issue(9) :1992-1998.DOI:10.16303/j.cnki.1005-4545.2024.09.19

镉通过Nrf2/HO-1通路诱导猪肾PK-15细胞铁死亡

Cadmium induces ferroptosis by modulating Nrf2/HO-1 pathway in PK-15 cells

宋圣哲 罗通旺 沈灵俊 吴亚 王书杰 郁孝强 宋厚辉 邵春艳
中国兽医学报2024,Vol.44Issue(9) :1992-1998.DOI:10.16303/j.cnki.1005-4545.2024.09.19

镉通过Nrf2/HO-1通路诱导猪肾PK-15细胞铁死亡

Cadmium induces ferroptosis by modulating Nrf2/HO-1 pathway in PK-15 cells

宋圣哲 1罗通旺 1沈灵俊 1吴亚 1王书杰 1郁孝强 1宋厚辉 1邵春艳1
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作者信息

  • 1. 浙江农林大学动物科技学院/动物医学院浙江省畜禽绿色生态健康养殖应用技术研究重点实验室/动物健康互联网检测技术浙江省工程实验室,动物医学与健康管理浙江省国际科技合作基地/中澳动物健康大数据分析联合实验室,浙江 杭州 311300
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摘要

镉(Cd)是非必需且难降解的重金属元素之一,镉蓄积可导致肾损伤.本研究旨在揭示镉暴露诱导猪肾PK-15细胞铁死亡的作用机制.首先,CCK-8法检测梯度浓度氯化镉(CdCl2)对PK-15细胞活力的影响,以筛选出合适的工作浓度.其次,用10 μmol/L CdCl2处理PK-15细胞3、6和12 h,通过比色法检测细胞上清中乳酸脱氢酶(LDH)和细胞中丙二醛(MDA)的含量,Western blot检测铁死亡相关蛋白的表达水平,荧光探针法检测细胞中亚铁离子(Fe2+)含量.最后,用10 μmol/L的HO-1抑制剂锌原卟啉(ZnPP)预处理PK-15细胞6 h,再用10 μmol/L CdCl2处理PK-15细胞12 h,通过相差显微镜观察细胞形态,Western blot检测铁死亡相关蛋白表达水平.结果显示,CdCl2处理后细胞活力下降,LDH、MDA和Fe2+水平极显著升高(P<0.01),Nrf2、HO-1和ALOX5蛋白表达水平极显著升高(P<0.01),FTH1蛋白表达水平极显著下降(P<0.01).与CdCl2处理组相比,ZnPP可以显著改善CdCl2处理诱导的猪肾PK-15细胞铁死亡.ZnPP预处理组细胞形态明显好转,Nrf2、HO-1和ALOX5蛋白表达水平极显著下降(P<0.01),FTH1蛋白表达水平极显著升高(P<0.01).结果表明,镉通过Nrf2/HO-1通路引发细胞铁过载和脂质过氧化,诱导猪肾PK-15细胞铁死亡.

Abstract

Cadmium is a non-essential,non-degradable heavy metal element,the accumulation of cadmium can cause kidney damage.The purpose of this study was to reveal the ferroptosis mecha-nism induced by cadmium exposure in porcine kidney PK-15 cells.First of all,cell viability of gradi-ent concentration of cadmium chloride(CdCl2)on PK-15 cells was detected by CCK-8 method to screen suitable work concentration of CdCl2.Secondly,PK-15 cells were treated with 10 μmol/L CdCl2 for 3,6 and 12 h.The contents of lactate dehydrogenase(LDH)malondialdehyde(MDA)were detected by colorimetry,the expression of ferroptosis related protein were detected by West-ern blot,and the content of ferrous ion(Fe2+)was detected by fluorescence probe.Finally,PK-15 cells were pretreated with 10 μmol/L HO-1 inhibitor zinc protoporphyrin(ZnPP)for 6 h,and then treated with 10 μmol/L CdCl2 for 12 h.The morphology of PK-15 cells was observed by phase contrast microscope and the expression of ferroptosis related proteins was detected by West-ern blot.The results showed that CdCl2 treatment caused a significant decrease in cell viability.The levels of LDH,MDA and Fe2+,the protein levels of Nrf2,HO-1 and ALOX5,and the expression level of FTH1 protein were significantly decreased in CdCl2 treatment cells(P<0.01).ZnPP could significantly improve the ferroptosis of PK-15 cells induced by CdCl2 treatment.Compared with CdCl2 treatment cells,the cell morphology was significantly improved,the protein levels of Nrf2,HO-1 and ALOX5 protein were significantly decreased,and the protein levels of FTH1 protein were significantly increased in ZnPP pretreatment group(P<0.01).The results showed that cad-mium induced iron overload and lipid peroxidation,which result in ferroptosis in PK-15 cells through Nrf2/HO-1 pathway.

关键词

/PK-15/HO-1/铁死亡

Key words

cadmium/PK-15/HO-1/ferroptosis

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基金项目

浙江农林大学科研发展基金人才启动基金资助项目(2034020072)

浙江省大学生科技创新活动计划暨新苗人才计划资助项目(2023R412021)

出版年

2024
中国兽医学报
吉林大学

中国兽医学报

CSTPCDCSCD北大核心
影响因子:0.702
ISSN:1005-4545
参考文献量17
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