Objective:Age-relate cataract(ARC)is a disease of the eyes with no effective drugs to prevent or treat patients.The aim of the present study is to determine whether histone H3,αA-crystallin(CRYAA),β-galactosidase(GL...Objective:Age-relate cataract(ARC)is a disease of the eyes with no effective drugs to prevent or treat patients.The aim of the present study is to determine whether histone H3,αA-crystallin(CRYAA),β-galactosidase(GLB1),and p53 are involved in the pathogenesis of ARC.Methods:A total of 99 anterior lens capsules(ALCs)of patients with ARC of various nuclear grades,ultraviolet models of ALCs,and two human lens epithelial cell lines(FHL-124 and SRA01/04)were used,and the expression of histone H3,CRYAA,GLB1,and p53 were detected by immunoblotting and reverse transcription and real time-quantitative polymerase chain reaction.The association between CRYAA with histone H3,GLB1,and p53 was assessed in FHL-124 and SRA01/04 cells following CRYAA overexpression.Results:Histone H3 and p53 in ALCs of patients with ARC were up-regulated in a grade-dependent manner,and the expression of CRYAA showed a positive association with histone H3,p53,and GLB1.In UV models of ALCs and human lens epithelial cell lines,the expression levels of histone H3,cell apoptosis factors(Bax/Bcl-2,cleaved caspase-3),and inflammation factors(interleukin-6,tumor necrosis factor-α)were all up-regulated.Furthermore,transfection of CRYAA in FHL-124 cells induced overexpression of histone H3.Conclusion:CRYAA-mediated upregulation of histone H3 may be involved in the pathogenesis of ARC.p53 may also have a role in ARC development,but not via the CRYAA-histone H3 axis.The results of the present study may assist in improving our understanding of the pathogenesis of ARC and in identifying potential targets for treatment.展开更多
Background:Cataracts are the leading cause of reversible blindness worldwide.Diabetic cataract(DC),a prevalent complication of diabetes mellitus,is characterized by its high occurrence,rapid progression,and severe imp...Background:Cataracts are the leading cause of reversible blindness worldwide.Diabetic cataract(DC),a prevalent complication of diabetes mellitus,is characterized by its high occurrence,rapid progression,and severe impact.The prevalence of diabetes varies greatly between the northern and southern regions,with higher rates observed among northern residents.DC-induced lens opacity is mainly attributed to oxidative stress.However,it remains unclear whether ferroptosis,a form of regulated cell death,occurs in crystalline epithelial cells during the pathogenesis,which may represent a novel mechanism contributing to DC.Methods:Transmission electron microscopy,quantitative assays for iron levels and reactive oxygen species(ROS),real-time quantitative polymerase chain reaction(RT-qPCR),western blotting,immunofluorescence,and immunohistochemistry were used to detect ferroptosis.Gene editing techniques were utilized to study the regulatory relationships among lipocalin 2(LCN2),glutathione peroxidase 4(GPX4),and ferritin heavy chain(FTH).Local knockdown of the LCN2 gene in B-3 cells and the eyes of Sprague Dawley(SD)rats was performed to verify and further explore the role and regulatory mechanisms of LCN2 in DC-associated ferroptosis.Results:An in vitro model using high glucose levels and an in vivo model with streptozotocin-induced diabetes in SD rats were successfully established.Ferroptosis was observed in both in vitro and in vivo experiments.LCN2 protein was normally expressed in human and rat lens epithelial cells,but its expression significantly increased during ferroptosis.The ferroptosis inhibitor,ferrostatin-1(Fer-1)effectively inhibited ferroptosis and reduced LCN2 protein expression.Notably,local knockdown of LCN2 via gene editing protected lens epithelial cells from ferroptosis in vitro and slowed the progression of DC in SD rats in vivo.Conclusion:Our findings underscore the significant role of ferroptosis in the pathogenesis of DC,suggesting that selectively targeting LCN2 activation and enhancing ferroptosis resistance may offer a novel therapeutic approach for treating DC.展开更多
基金This work was supported by the Nature Science Foundation of China(81470618)the Scientific Research Foundation of First Affiliated Hospital of Harbin Medical University(2017B013).
文摘Objective:Age-relate cataract(ARC)is a disease of the eyes with no effective drugs to prevent or treat patients.The aim of the present study is to determine whether histone H3,αA-crystallin(CRYAA),β-galactosidase(GLB1),and p53 are involved in the pathogenesis of ARC.Methods:A total of 99 anterior lens capsules(ALCs)of patients with ARC of various nuclear grades,ultraviolet models of ALCs,and two human lens epithelial cell lines(FHL-124 and SRA01/04)were used,and the expression of histone H3,CRYAA,GLB1,and p53 were detected by immunoblotting and reverse transcription and real time-quantitative polymerase chain reaction.The association between CRYAA with histone H3,GLB1,and p53 was assessed in FHL-124 and SRA01/04 cells following CRYAA overexpression.Results:Histone H3 and p53 in ALCs of patients with ARC were up-regulated in a grade-dependent manner,and the expression of CRYAA showed a positive association with histone H3,p53,and GLB1.In UV models of ALCs and human lens epithelial cell lines,the expression levels of histone H3,cell apoptosis factors(Bax/Bcl-2,cleaved caspase-3),and inflammation factors(interleukin-6,tumor necrosis factor-α)were all up-regulated.Furthermore,transfection of CRYAA in FHL-124 cells induced overexpression of histone H3.Conclusion:CRYAA-mediated upregulation of histone H3 may be involved in the pathogenesis of ARC.p53 may also have a role in ARC development,but not via the CRYAA-histone H3 axis.The results of the present study may assist in improving our understanding of the pathogenesis of ARC and in identifying potential targets for treatment.
基金approved by the Institutional Review Board(or Ethics Committee)of the First Affiliated Hospital of Harbin Medical University(IACUC:2022020 and date of approval:April 8,2022).
文摘Background:Cataracts are the leading cause of reversible blindness worldwide.Diabetic cataract(DC),a prevalent complication of diabetes mellitus,is characterized by its high occurrence,rapid progression,and severe impact.The prevalence of diabetes varies greatly between the northern and southern regions,with higher rates observed among northern residents.DC-induced lens opacity is mainly attributed to oxidative stress.However,it remains unclear whether ferroptosis,a form of regulated cell death,occurs in crystalline epithelial cells during the pathogenesis,which may represent a novel mechanism contributing to DC.Methods:Transmission electron microscopy,quantitative assays for iron levels and reactive oxygen species(ROS),real-time quantitative polymerase chain reaction(RT-qPCR),western blotting,immunofluorescence,and immunohistochemistry were used to detect ferroptosis.Gene editing techniques were utilized to study the regulatory relationships among lipocalin 2(LCN2),glutathione peroxidase 4(GPX4),and ferritin heavy chain(FTH).Local knockdown of the LCN2 gene in B-3 cells and the eyes of Sprague Dawley(SD)rats was performed to verify and further explore the role and regulatory mechanisms of LCN2 in DC-associated ferroptosis.Results:An in vitro model using high glucose levels and an in vivo model with streptozotocin-induced diabetes in SD rats were successfully established.Ferroptosis was observed in both in vitro and in vivo experiments.LCN2 protein was normally expressed in human and rat lens epithelial cells,but its expression significantly increased during ferroptosis.The ferroptosis inhibitor,ferrostatin-1(Fer-1)effectively inhibited ferroptosis and reduced LCN2 protein expression.Notably,local knockdown of LCN2 via gene editing protected lens epithelial cells from ferroptosis in vitro and slowed the progression of DC in SD rats in vivo.Conclusion:Our findings underscore the significant role of ferroptosis in the pathogenesis of DC,suggesting that selectively targeting LCN2 activation and enhancing ferroptosis resistance may offer a novel therapeutic approach for treating DC.
