RSL3是谷胱甘肽过氧化物酶4(GPx4)的常见抑制剂,可诱导铁凋亡。铁凋亡是铁离子依赖的,氧化型程序性细胞死亡。在这项研究中,用RSL3刺激幼体/成年斑马鱼构建铁凋亡模型,以CYP2R1-/-斑马鱼为1,25(OH)2D3敲低模型,探讨1,25(OH)2D3/VD3对RSL3诱导的铁凋亡的调控作用及机制。结果表明,1,25(OH)2D3/VD3减轻了RSL3诱导的斑马鱼幼虫/成虫肝脏线粒体损伤,逆转了GPx4活性的下降,减少了ROS的积累,LPO和MDA。VD3还抑制成年鱼肝中的铁调素(HEPC),促进了运铁素(FPN)的生产,减少了Fe2+的聚集。外源性1,25(OH)2D3增加RSL3治疗后CYP2R1-/-存活和肝脏GPx4活性。在基因层面,1,25(OH)2D3/VD3激活Keap1-Nrf2-GPx4并抑制NFκB-hepcidin轴。在铁性死亡的背景下,cyp2r1基因的缺失导致gpx4表达更严重的下降,但外源性1,25(OH)2D3增加了RSL3处理后CYP2R1-/-斑马鱼肝脏中GPx4基因和蛋白的表达。结果表明,1,25(OH)2D3/VD3可以通过提高机体的抗氧化能力和调节铁离子的转运,从而抑制RSL3诱导的斑马鱼幼虫/成年斑马鱼肝脏的铁凋亡。外源性1,25(OH)2D3逆转CYP2R1-/-斑马鱼肝脏中GPx4的下调。与铁凋亡抑制剂Fer-1相比,1,25(OH)2D3/VD3的作用机制是多样化且非特异性的。这项研究表明,在斑马鱼的不同发育阶段,VD3对RSL3诱导的铁细胞凋亡具有抗性。
RSL3 is a common inhibitor of glutathione peroxidase 4 (GPx4) that can induce ferroptosis. Ferroptosis is an iron ion-dependent, oxidative-type of programmed cell death. In this study, larval/adult zebrafish were stimulated with RSL3 to construct a ferroptosis model, and
CYP2R1-/- zebrafish was used as a 1,25(OH)2D3 knock-down model to explore the regulatory effect and mechanism of 1,25(OH)2D3/VD3 on RSL3-induced ferroptosis. The results showed that 1,25(OH)2D3/VD3 alleviated RSL3 induced mitochondrial damage in liver of larval/adult zebrafish, reversed the decline of GPx4 activity, and reduced the accumulation of ROS, LPO and MDA. VD3 also inhibited hepcidin (HEPC) in adult fish liver, promoted the production of ferroportin (FPN), and reduced the aggregation of Fe2+. Exogenous 1,25(OH)2D3 increased the
CYP2R1-/- survival and liver GPx4 activity after RSL3 treatment. At the gene level, 1,25(OH)2D3/VD3 activated Keap1-Nrf2-GPx4 and inhibited the NFκB-hepcidin axis. In the ferroptosis context, deletion of the
cyp2r1 gene resulted in a more severe decline in gpx4 expression, but the exogenous 1,25(OH)2D3 increased the expression of the GPx4 gene and protein in
CYP2R1-/- zebrafish liver after RSL3 treatment. The collective results indicated that 1,25(OH)2D3/VD3 can inhibit ferroptosis induced by RSL3 in liver of larval/adult zebrafish by improving the antioxidant capacity and regulating iron ion transport. Exogenous 1,25(OH)2D3 reverses the downregulation of GPx4 in the
CYP2R1-/- zebrafish liver in the ferroptosis state. Compared with the ferroptosis inhibitor Fer-1, the mechanism of action of 1,25(OH)2D3/VD3 is diversified and nonspecific. This study demonstrated the resistance of VD3 to RSL3-induced ferroptosis at different developmental stages in zebrafish.