Regulation Of Iron Overload-induced Ferroptosis And Expression Of TRF By PPARα

Ferroptosis is a new regulated cells death manner defined as results of iron-dependent accumulation of lipid peroxidation.Ferroptosis is morphologically,biochemically and genetically distinct from other forms of cell death,including apoptosis,necrosis and autophagy.Ferrousiron(Fe²⁺)accumulation and lipid peroxidation are critical events in the induction of ferroptosis,which is inhibited by iron chelators and lipophilic antioxidants.Peroxisome proliferators-activated receptorα（PPARα）is a nuclear receptor involved in the regulation of lipid metabolism.Upon ligand-induced activation,PPARαstimulates the expression of target genes directly through binding to PPAR response elements（PPRE）in the promoter regions of target genes,and then regulates the expression of genes involved in lipid metabolism and peroxisome proliferation.The biological roles of iron and iron metabolism in ferroptosis remain poorly understood.Transferrin（TRF）,a serum glycoprotein,plays an essential role in the transport of iron from sites of absorption and storage to iron-requiring cells.Moreover,iron overload-induced ferroptosis damage was rescued by the ferroptosis inhibitor Fer-1in mouse livers.As an essential regulator of ferroptosis,Glutathione peroxidase 4（GPX4）acts through the suppression of lipid peroxidation generation to play a central role in ferroptosis.In this paper,we investigated the molecular mechanisms of regulation of iron overload-induced ferroptosis and expression of TRF by PPARα,and results were as the following:（1）PPARαactivation suppresses TRF expression in vitro and in vivoHep1-6 cells were treated with PPARαligand GW7647,and the expression of TRF was significantly decreased in m RNA levels.Mice were gavaged with either vehicle or GW7647 and the expression of TRF was examined,and it is showed that TRF was significantly decreased in m RNA and protein level after PPARαligand treatment.All of the results showed that TRF expression was suppressed by PPARαin vitro and in vivo.（2）PPARαactivation suppresses trans-activity of the TRFHep1-6 cells were transfected with the TRF reporter plasmid.Cells were subsequently treated with GW7647 for 24 h.Dual luciferase reporter gene assay showed that the transcriptional activity of TRF was significantly decreased following PPARαactivation.（3）Either PPARαagonist GW7647 or ferroptosis inhibitor Fer-1 suppresses iron overload-induced cell deathPPARα^-/-mice were fed a high-iron diet and treated with Fer-1 for rescue purpose.High-iron diet strongly elevated the content of iron in liver,while Fer-1 administration significantly decreased the Malondialdehyde（MDA）content,the iron content,the iron content and Prostaglandinendoperoxidesynthase2（PTGS2）m RNA levels in mice.These data indicated that iron overload induced ferroptosis in vivo.Wild type（WT）mice were fed a high-iron diet and treated either with Fer-1 or GW7647.The MDA content,the iron content,the iron content and PTGS2 m RNA levels were both significantly decreased in mice after GW7647 gauage or Fer-1 administration.All the above data indicated that iron overload–induced liver ferroptosis was rescued by PPARαactivation,and it had the comparable effect as Fer-1 supplementation on iron overload-induced ferroptosis inhibition.（4）GPX4 overexpression alleviates iron overload ferroptosis in PPARα^-/-mouse liversHep1-6 cells were transfected with GPX4 and control eukaryotic expression plasmids.The expression of GPX4 was significantly increased in m RNA and protein levels.PPARα^-/-mice were fed a high-iron diet following GPX4 overexpression.The content of iron in liver was significantly elevated after high-iron diet,and as expected,GPX4 overexpression significantly decreased the MDA content,the iron content,the iron content and PTGS2 m RNA levels in mice.These data indicated that GPX4overexpression alleviated iron overload ferroptosis in PPARα^-/-mouse livers.