Ferroptosis is a type of programmed cell death dependent on iron.It is different from other forms of cell death such as apoptosis,classic necrosis and autophagy.Ferroptosis is involved in many neurodegenerative diseas...Ferroptosis is a type of programmed cell death dependent on iron.It is different from other forms of cell death such as apoptosis,classic necrosis and autophagy.Ferroptosis is involved in many neurodegenerative diseases.The role of ferroptosis in glutamate-induced neuronal toxicity is not fully understood.To test its toxicity,glutamate(1.25–20 mM)was applied to HT-22 cells for 12 to 48 hours.The optimal experimental conditions occurred at 12 hours after incubation with 5 mM glutamate.Cells were cultured with 3–12μM ferrostatin-1,an inhibitor of ferroptosis,for 12 hours before exposure to glutamate.The cell viability was detected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay.Autophagy was determined by monodansylcadaverine staining and apoptosis by caspase 3 activity.Damage to cell structures was observed under light and by transmission electron microscopy.The release of lactate dehydrogenase was detected by the commercial kit.Reactive oxygen species were measured by flow cytometry.Glutathione peroxidase activity,superoxide dismutase activity and malondialdehyde level were detected by the appropriate commercial kit.Prostaglandin peroxidase synthase 2 and glutathione peroxidase 4 gene expression was detected by real-time quantitative polymerase chain reaction.Glutathione peroxidase 4 and nuclear factor erythroid-derived-like 2 protein expression was detected by western blot analysis.Results showed that ferrostatin-1 can significantly counter the effects of glutamate on HT-22 cells,improving the survival rate,reducing the release of lactate dehydrogenase and reducing the damage to mitochondrial ultrastructure.However,it did not affect the caspase-3 expression and monodansylcadaverine-positive staining in glutamate-injured HT-22 cells.Ferrostatin-1 reduced the levels of reactive oxygen species and malondialdehyde and enhanced superoxide dismutase activity.It decreased gene expression of prostaglandin peroxidase synthase 2 and increased gene expression of glutathione peroxidase 4 and protein expressions of glutathione peroxidase 4 and nuclear factor(erythroid-derived)-like 2 in glutamate-injured HT-22 cells.Treatment of cultured cells with the apoptosis inhibitor Z-Val-Ala-Asp(OMe)-fluoromethyl ketone(2–8μM),autophagy inhibitor 3-methyladenine(100–400μM)or necrosis inhibitor necrostatin-1(10–40μM)had no effect on glutamate induced cell damage.However,the iron chelator deferoxamine mesylate salt inhibited glutamate induced cell death.Thus,the results suggested that ferroptosis is caused by glutamate-induced toxicity and that ferrostatin-1 protects HT-22 cells from glutamate-induced oxidative toxicity by inhibiting the oxidative stress.展开更多
Oxidative stress-mediated cell death in cardiomyocytes contributes to the development of atrial fibrillation.However,the detailed mechanisms are still unclear.In the present study,we established atrial fibrillation mo...Oxidative stress-mediated cell death in cardiomyocytes contributes to the development of atrial fibrillation.However,the detailed mechanisms are still unclear.In the present study,we established atrial fibrillation models in mice.The cardiomyocytes were isolated from atrial fibrillation mice and normal mice and were cultured in vitro,respectively.The results showed that cell proliferation and viability in cardiomyocytes with atrial fibrillation were significantly lower than the cells from the normal mice.Consistently,atrial fibrillation cardiomyocytes were prone to suffer from apoptotic cell death.Also,the oxidative stress and ferroptosis-associated signatures were significantly increased in atrial fibrillation cardiomyocytes compared to normal cardiomyocytes,and ferroptosis inhibitor and NAC rescued cell viability in atrial fibrillation cardiomyocytes during in vitro cell culture.In addition,low-expressed miR-143-3p was observed in atrial fibrillation cardiomyocytes compared to normal cardiomyocytes,and overexpression of miR-143-3p increased cell proliferation and inhibited cell death in atrial fibrillation cardiomyocytes.Furthermore,glutamic-oxaloacetic transaminase 1 could be negatively regulated by miR-143-3p in normal cardiomyocytes,and miR-143-3p overexpression inhibited cell ferroptosis in atrial fibrillation cardiomyocytes by sponging glutamic-oxaloacetic transaminase 1.Collectively,overexpression of miR-143-3p increased cell viability and promoted cell proliferation in cardiomyocytes with atrial fibrillation by inhibiting glutamic-oxaloacetic transaminase 1 mediated oxidative damages and cell ferroptosis.展开更多
目的:研究铁抑素-1(Fer-1)对血红素诱导的小鼠海马神经元铁死亡的影响。方法:小鼠海马神经元细胞系HT22细胞分为对照组(control)、血红素处理组(hemin)以及血红素和铁抑素-1联合处理组(hemin+Fer1)。利用CCK-8试剂盒检测各组细胞的活性...目的:研究铁抑素-1(Fer-1)对血红素诱导的小鼠海马神经元铁死亡的影响。方法:小鼠海马神经元细胞系HT22细胞分为对照组(control)、血红素处理组(hemin)以及血红素和铁抑素-1联合处理组(hemin+Fer1)。利用CCK-8试剂盒检测各组细胞的活性,利用活性氧荧光探针(DCFH-DA)法检测各组细胞中反应性活性氧(ROS)的含量,利用商品化试剂盒检测谷胱甘肽(GSH)的含量,利用JC-1试剂盒检测线粒体膜电位(MMP)变化,利用Western Blot和real time RT-PCR检测细胞中谷胱甘肽过氧化物酶4(GPX4)的表达。结果:血红素处理可降低HT22细胞活性,诱发氧化应激,破坏线粒体膜电位(MMP),并下调GPX4表达而诱导铁死亡。相反,Fer-1可以使细胞活性恢复,降低氧化应激反应,从而抑制HT22细胞铁死亡。结论:血红素中诱发了HT22细胞内脂质过氧化反应,引起线粒体损伤及铁死亡;铁死亡抑制剂Fer-1可以阻断血红素这种细胞毒性。展开更多
BACKGROUND Ferroptosis is involved in developing inflammatory diseases;yet,its role in acute hypertriglyceridemic pancreatitis(HTGP)remains unclear.AIM To explore whether ferroptosis is involved in the process of HTGP...BACKGROUND Ferroptosis is involved in developing inflammatory diseases;yet,its role in acute hypertriglyceridemic pancreatitis(HTGP)remains unclear.AIM To explore whether ferroptosis is involved in the process of HTGP and elucidate its potential mechanisms.METHODS An HTGP mouse model was induced using intraperitoneal injection of P-407 and caerulein(CAE).Then,pancreatic tissues from the model animals were subjected to proteome sequencing analysis.The pathological changes and scores of the pancreas,lung,and kidney were determined using hematoxylin-eosin staining.The levels of serum amylase(AMY),triglyceride,and total cholesterol were measured with an automatic blood cell analyzer.Additionally,the serum levels of tumor necrosis factor(TNF)-α,interleukin(IL)-6,and IL-1βwere determined by enzyme linked immunosorbent assay.Malonaldehyde(MDA),glutathione(GSH),and Fe^(2+)were detected in the pancreas.Finally,immunohistochemistry was performed to assess the expression of ferroptosis-related proteins.RESULTS Proteome sequencing revealed that ferroptosis was involved in the process of HTGP and that NADPH oxidase(NOX)2 may participate in ferroptosis regulation.Moreover,the levels of serum AMY,TNF-α,IL-6,and IL-1βwere significantly increased,MDA and Fe^(2+)were upregulated,GSH and ferroptosis-related proteins were reduced,and the injury of the pancreas,lung,and kidney were aggravated in the P407+CAE group compared to CAE and wild type groups(all P<0.05).Notably,the inhibition of ferroptosis and NOX2 attenuated the pathological damage and the release of TNF-α,IL-6,and IL-1βin the serum of the mice.CONCLUSION Ferroptosis was found to have an important role in HTGP and may be considered a potential target for clinical treatment.展开更多
目的:验证心肌细胞缺氧/复氧(H/R)损伤中是否有铁死亡的发生,以及探讨铁死亡抑制剂Ferrostatin-1(Fer-1)对心肌细胞H/R损伤的作用及其机制。方法:新生1~3 d SD乳鼠,提取原代心肌细胞,随机分为正常对照组(Control)、H/R组和H/R+Fer-1组。...目的:验证心肌细胞缺氧/复氧(H/R)损伤中是否有铁死亡的发生,以及探讨铁死亡抑制剂Ferrostatin-1(Fer-1)对心肌细胞H/R损伤的作用及其机制。方法:新生1~3 d SD乳鼠,提取原代心肌细胞,随机分为正常对照组(Control)、H/R组和H/R+Fer-1组。H/R组细胞培养52 h后,加入4 mmol/L Na 2 S 2 O 4溶液,缺氧1 h后,用含10%小牛血清的DMEM培养液复氧培养3 h。H/R+Fer-1组经Fer-1(2μmol/L)预处理24 h后再进行缺氧复氧处理。各组细胞应用紫外分光光度法检测乳酸脱氢酶(LDH)释放率,CCK-8法检测细胞存活率,黄嘌呤氧化酶法检测超氧化物歧化酶(SOD),化学比色法检测丙二醛(MDA),免疫荧光观测线粒体膜电位、活性氧(ROS)改变,Western blot检测铁死亡关键蛋白ACSL4、GPX4的表达。结果:与control组比较,H/R组细胞活性、SOD释放量和MMP水平均显著降低(P<0.05),LDH、MDA、ROS释放量均显著增加(P<0.05),ACSL4蛋白表达显著升高(P<0.05)、GPX4蛋白表达显著下降(P<0.05)。与H/R组比较,H/R+Fer-1组细胞活性、SOD释放量和MMP水平均显著升高(P<0.05),LDH、MDA、ROS释放量均显著降低(P<0.05),ACSL4蛋白表达显著下降(P<0.05)、GPX4蛋白表达显著升高(P<0.05)。结论:心肌细胞H/R损伤中有铁死亡的发生,Fer-1可通过调控ACSL4和GPX4抑制细胞内ROS的产生,从而减轻铁死亡引起的原代心肌细胞缺氧复氧损伤。展开更多
文摘Ferroptosis is a type of programmed cell death dependent on iron.It is different from other forms of cell death such as apoptosis,classic necrosis and autophagy.Ferroptosis is involved in many neurodegenerative diseases.The role of ferroptosis in glutamate-induced neuronal toxicity is not fully understood.To test its toxicity,glutamate(1.25–20 mM)was applied to HT-22 cells for 12 to 48 hours.The optimal experimental conditions occurred at 12 hours after incubation with 5 mM glutamate.Cells were cultured with 3–12μM ferrostatin-1,an inhibitor of ferroptosis,for 12 hours before exposure to glutamate.The cell viability was detected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay.Autophagy was determined by monodansylcadaverine staining and apoptosis by caspase 3 activity.Damage to cell structures was observed under light and by transmission electron microscopy.The release of lactate dehydrogenase was detected by the commercial kit.Reactive oxygen species were measured by flow cytometry.Glutathione peroxidase activity,superoxide dismutase activity and malondialdehyde level were detected by the appropriate commercial kit.Prostaglandin peroxidase synthase 2 and glutathione peroxidase 4 gene expression was detected by real-time quantitative polymerase chain reaction.Glutathione peroxidase 4 and nuclear factor erythroid-derived-like 2 protein expression was detected by western blot analysis.Results showed that ferrostatin-1 can significantly counter the effects of glutamate on HT-22 cells,improving the survival rate,reducing the release of lactate dehydrogenase and reducing the damage to mitochondrial ultrastructure.However,it did not affect the caspase-3 expression and monodansylcadaverine-positive staining in glutamate-injured HT-22 cells.Ferrostatin-1 reduced the levels of reactive oxygen species and malondialdehyde and enhanced superoxide dismutase activity.It decreased gene expression of prostaglandin peroxidase synthase 2 and increased gene expression of glutathione peroxidase 4 and protein expressions of glutathione peroxidase 4 and nuclear factor(erythroid-derived)-like 2 in glutamate-injured HT-22 cells.Treatment of cultured cells with the apoptosis inhibitor Z-Val-Ala-Asp(OMe)-fluoromethyl ketone(2–8μM),autophagy inhibitor 3-methyladenine(100–400μM)or necrosis inhibitor necrostatin-1(10–40μM)had no effect on glutamate induced cell damage.However,the iron chelator deferoxamine mesylate salt inhibited glutamate induced cell death.Thus,the results suggested that ferroptosis is caused by glutamate-induced toxicity and that ferrostatin-1 protects HT-22 cells from glutamate-induced oxidative toxicity by inhibiting the oxidative stress.
基金financially supported by the Youth Science and Innovation Research Fund of Xinjiang Province(No.WJWY-201966).
文摘Oxidative stress-mediated cell death in cardiomyocytes contributes to the development of atrial fibrillation.However,the detailed mechanisms are still unclear.In the present study,we established atrial fibrillation models in mice.The cardiomyocytes were isolated from atrial fibrillation mice and normal mice and were cultured in vitro,respectively.The results showed that cell proliferation and viability in cardiomyocytes with atrial fibrillation were significantly lower than the cells from the normal mice.Consistently,atrial fibrillation cardiomyocytes were prone to suffer from apoptotic cell death.Also,the oxidative stress and ferroptosis-associated signatures were significantly increased in atrial fibrillation cardiomyocytes compared to normal cardiomyocytes,and ferroptosis inhibitor and NAC rescued cell viability in atrial fibrillation cardiomyocytes during in vitro cell culture.In addition,low-expressed miR-143-3p was observed in atrial fibrillation cardiomyocytes compared to normal cardiomyocytes,and overexpression of miR-143-3p increased cell proliferation and inhibited cell death in atrial fibrillation cardiomyocytes.Furthermore,glutamic-oxaloacetic transaminase 1 could be negatively regulated by miR-143-3p in normal cardiomyocytes,and miR-143-3p overexpression inhibited cell ferroptosis in atrial fibrillation cardiomyocytes by sponging glutamic-oxaloacetic transaminase 1.Collectively,overexpression of miR-143-3p increased cell viability and promoted cell proliferation in cardiomyocytes with atrial fibrillation by inhibiting glutamic-oxaloacetic transaminase 1 mediated oxidative damages and cell ferroptosis.
文摘目的:研究铁抑素-1(Fer-1)对血红素诱导的小鼠海马神经元铁死亡的影响。方法:小鼠海马神经元细胞系HT22细胞分为对照组(control)、血红素处理组(hemin)以及血红素和铁抑素-1联合处理组(hemin+Fer1)。利用CCK-8试剂盒检测各组细胞的活性,利用活性氧荧光探针(DCFH-DA)法检测各组细胞中反应性活性氧(ROS)的含量,利用商品化试剂盒检测谷胱甘肽(GSH)的含量,利用JC-1试剂盒检测线粒体膜电位(MMP)变化,利用Western Blot和real time RT-PCR检测细胞中谷胱甘肽过氧化物酶4(GPX4)的表达。结果:血红素处理可降低HT22细胞活性,诱发氧化应激,破坏线粒体膜电位(MMP),并下调GPX4表达而诱导铁死亡。相反,Fer-1可以使细胞活性恢复,降低氧化应激反应,从而抑制HT22细胞铁死亡。结论:血红素中诱发了HT22细胞内脂质过氧化反应,引起线粒体损伤及铁死亡;铁死亡抑制剂Fer-1可以阻断血红素这种细胞毒性。
基金Supported by the National Natural Science Foundation of Shandong Province,No.ZR2021MH032.
文摘BACKGROUND Ferroptosis is involved in developing inflammatory diseases;yet,its role in acute hypertriglyceridemic pancreatitis(HTGP)remains unclear.AIM To explore whether ferroptosis is involved in the process of HTGP and elucidate its potential mechanisms.METHODS An HTGP mouse model was induced using intraperitoneal injection of P-407 and caerulein(CAE).Then,pancreatic tissues from the model animals were subjected to proteome sequencing analysis.The pathological changes and scores of the pancreas,lung,and kidney were determined using hematoxylin-eosin staining.The levels of serum amylase(AMY),triglyceride,and total cholesterol were measured with an automatic blood cell analyzer.Additionally,the serum levels of tumor necrosis factor(TNF)-α,interleukin(IL)-6,and IL-1βwere determined by enzyme linked immunosorbent assay.Malonaldehyde(MDA),glutathione(GSH),and Fe^(2+)were detected in the pancreas.Finally,immunohistochemistry was performed to assess the expression of ferroptosis-related proteins.RESULTS Proteome sequencing revealed that ferroptosis was involved in the process of HTGP and that NADPH oxidase(NOX)2 may participate in ferroptosis regulation.Moreover,the levels of serum AMY,TNF-α,IL-6,and IL-1βwere significantly increased,MDA and Fe^(2+)were upregulated,GSH and ferroptosis-related proteins were reduced,and the injury of the pancreas,lung,and kidney were aggravated in the P407+CAE group compared to CAE and wild type groups(all P<0.05).Notably,the inhibition of ferroptosis and NOX2 attenuated the pathological damage and the release of TNF-α,IL-6,and IL-1βin the serum of the mice.CONCLUSION Ferroptosis was found to have an important role in HTGP and may be considered a potential target for clinical treatment.
文摘目的:验证心肌细胞缺氧/复氧(H/R)损伤中是否有铁死亡的发生,以及探讨铁死亡抑制剂Ferrostatin-1(Fer-1)对心肌细胞H/R损伤的作用及其机制。方法:新生1~3 d SD乳鼠,提取原代心肌细胞,随机分为正常对照组(Control)、H/R组和H/R+Fer-1组。H/R组细胞培养52 h后,加入4 mmol/L Na 2 S 2 O 4溶液,缺氧1 h后,用含10%小牛血清的DMEM培养液复氧培养3 h。H/R+Fer-1组经Fer-1(2μmol/L)预处理24 h后再进行缺氧复氧处理。各组细胞应用紫外分光光度法检测乳酸脱氢酶(LDH)释放率,CCK-8法检测细胞存活率,黄嘌呤氧化酶法检测超氧化物歧化酶(SOD),化学比色法检测丙二醛(MDA),免疫荧光观测线粒体膜电位、活性氧(ROS)改变,Western blot检测铁死亡关键蛋白ACSL4、GPX4的表达。结果:与control组比较,H/R组细胞活性、SOD释放量和MMP水平均显著降低(P<0.05),LDH、MDA、ROS释放量均显著增加(P<0.05),ACSL4蛋白表达显著升高(P<0.05)、GPX4蛋白表达显著下降(P<0.05)。与H/R组比较,H/R+Fer-1组细胞活性、SOD释放量和MMP水平均显著升高(P<0.05),LDH、MDA、ROS释放量均显著降低(P<0.05),ACSL4蛋白表达显著下降(P<0.05)、GPX4蛋白表达显著升高(P<0.05)。结论:心肌细胞H/R损伤中有铁死亡的发生,Fer-1可通过调控ACSL4和GPX4抑制细胞内ROS的产生,从而减轻铁死亡引起的原代心肌细胞缺氧复氧损伤。
