Objective·To explore the potential role of SUMO-specific protease 1(SENP1)in ferroptosis.Methods·The Cancer Genome Atlas(TCGA)database was used to analyze the correlation between the expression levels of SENP1 and the ferroptosis-related genes,acyl-CoA synthetase long chain family member 4(ACSL4)and glutathione peroxidase 4(GPX4).Ferroptosis in human fibrosarcoma HT1080 cells,murine fibrosarcoma MCA-205 cells,and human embryonic kidney 293T cells was induced by RAS-selective lethal 3(RSL3).Quantitative real-time PCR(RT-qPCR)and Western blotting were used to detect the expression of SENP1.In 293T cells,immunoprecipitation-mass spectrometry was used to investigate the interacting proteins of SENP1 in the process of ferroptosis.The Flag-SENP1 plasmid was transiently transfected into 293T cells,and the overexpression efficiency of SENP1,along with the expression levels of ferroptosis-related genes ACSL4 and GPX4,was assessed by RT-qPCR and Western blotting.Results·TCGA database analysis showed that the expression of SENP1 was positively correlated with ACSL4 and negatively correlated with GPX4 in most tumor tissues.RT-qPCR and Western blotting showed that the expression level of SENP1 was significantly down-regulated in RSL3-treated HT1080,MCA-205,and 293T cells.Immunoprecipitation-mass spectrometry showed that SENP1 enriched SUMO molecules in the process of ferroptosis.Western blotting showed that the level of ACSL4 protein increased after SENP1 overexpression,and there was no significant change in the level of GPX4 protein.RT-qPCR showed that after SENP1 overexpression,there was no significant change in the mRNA levels of ACSL4 and GPX4.Conclusion·SENP1 gene expression is downregulated during ferroptosis,and may regulate the stability of ferroptosis-related protein ACSL4.

Objective·To explore the potential role of SUMO-specific protease 1(SENP1)in ferroptosis.Methods·The Cancer Genome Atlas(TCGA)database was used to analyze the correlation between the expression levels of SENP1 and the ferroptosis-related genes,acyl-CoA synthetase long chain family member 4(ACSL4)and glutathione peroxidase 4(GPX4).Ferroptosis in human fibrosarcoma HT1080 cells,murine fibrosarcoma MCA-205 cells,and human embryonic kidney 293T cells was induced by RAS-selective lethal 3(RSL3).Quantitative real-time PCR(RT-qPCR)and Western blotting were used to detect the expression of SENP1.In 293T cells,immunoprecipitation-mass spectrometry was used to investigate the interacting proteins of SENP1 in the process of ferroptosis.The Flag-SENP1 plasmid was transiently transfected into 293T cells,and the overexpression efficiency of SENP1,along with the expression levels of ferroptosis-related genes ACSL4 and GPX4,was assessed by RT-qPCR and Western blotting.Results·TCGA database analysis showed that the expression of SENP1 was positively correlated with ACSL4 and negatively correlated with GPX4 in most tumor tissues.RT-qPCR and Western blotting showed that the expression level of SENP1 was significantly down-regulated in RSL3-treated HT1080,MCA-205,and 293T cells.Immunoprecipitation-mass spectrometry showed that SENP1 enriched SUMO molecules in the process of ferroptosis.Western blotting showed that the level of ACSL4 protein increased after SENP1 overexpression,and there was no significant change in the level of GPX4 protein.RT-qPCR showed that after SENP1 overexpression,there was no significant change in the mRNA levels of ACSL4 and GPX4.Conclusion·SENP1 gene expression is downregulated during ferroptosis,and may regulate the stability of ferroptosis-related protein ACSL4.

Objective·To establish a reliable alcoholic liver disease mouse model (ALDNM) that mimics the drinking pattern of alcoholic liver disease (ALD) patients.Methods·Using the self-designed feeding tubes and liquid diet,ALDNM model was developed through chronic feeding combined with acute gavage of ethanol based on Lieber-DeCarli model and Gao-Binge model.C57BL/6 mice were administered with control liquid diet for adaptation for first 5 d,and then divided into pair-fed group and ethanol-fed group (10 mice each group).Ethanol-fed mice were fed with the liquid diet in which ethanol accounts for 30％ of total energy,while the pair-fed mice were fed with the control diet for 10 d.At the 16th day,ethanol-fed mice and pair-fed mice were respectively gavaged a single dose of 31.5％ ethanol or isocaloric maltose dextrin,and euthanized 9 h later.Sera and livers were collected.The general physiological condition,hepatic tissue pathological changes and serum indexes between Lieber-DeCarli models and ALDNM models were compared.The liver lipids of ALDNM mice were determined by Oil red O (ORO) staining and hepatic triacylglyceride (TAG) test.Meanwhile,the mRNA levels of interleukin-6 (IL-6),tumor necrosis factor α (TNF-α),fatty acid synthase (Fas),long chain fatty acid elongase 6 (Elovl6) and stearyl-CoA desaturase (Scdl) were detected by real-time PCR in ALDNM models.Western blotting was used to detect the changes of phosphorylated signal transduction and transcriptional activator (p-STAT3) in the livers.Results·Lieber-DeCarli model mice were generally in poor condition,and there was no significant change in serum glutamic-pyruvic transaminase (GPT) and glutamic-oxaloacetic transaminase (GOT) compared to pair-fed group.However,in ALDNM models,H-E staining showed that the hepatocytes of ethanol-fed mice were extremely swollen with round volume,increased cytoplasm and filled with large amounts of fat vacuoles.ORO staining analyses showed obvious microsteatosis in the liver cells from all ethanol-fed mice.The hepatosomatic index,liver TAG content,serum GPT and GOT of ALDNM models were significantly higher than those in the pair-fed group,while the serum HDL significantly decreased compared to the pair-fed group.Moreover,the expression levels of both lipid synthesis pathways and inflammatory signaling pathways related genes in livers significantly increased in the ethanol-fed mice of ALDNM model.Conclusion·ALDNM model was successfully constructed.This model is cost-and time-efficient.Moreover,ALDNM model mimics the drinking pattern and pathogenesis of ALD patients with the advantages of stable food intake,good repeatability,and obvious liver damage.