胃饥饿素对高糖下视网膜微血管内皮细胞氧化应激及铁死亡的抑制作用  

Inhibitory effects of ghrelin on oxidative stress and ferroptosis in retinal microvascular endothelial cells under high glucose

作　　者：李蓉[1] 张敏[2] 燕洁静 Li Rong;Zhang Min;Yan Jiejing(Department of Ophthalmology,The First Affiliated Hospital,Xi'an Medical University,Xi'an 710077,China;Department of Endocrinology,The First Affiliated Hospital,Xi'an Medical University,Xi'an 710077,China;Department of Ophthalmology,the First Affiliated Hospital of Northwest University,Xi'an No.1 Hospital,Shaanxi Ophthalmological Institute,Xi'an 710002,China)

机构地区：[1]西安医学院第一附属医院眼科,西安710077 [2]西安医学院第一附属医院内分泌科,西安710077 [3]西北大学附属第一医院眼科西安市第一医院眼科陕西省眼科研究所,西安710002

出　　处：《中华实验眼科杂志》2024年第11期991-996,共6页Chinese Journal Of Experimental Ophthalmology

基　　金：陕西省重点研发计划(2024SF-YBXM-324)。

摘　　要：目的研究胃饥饿素对高糖下人视网膜微血管内皮细胞(HRMEC)氧化应激及铁死亡的影响。方法将体外培养的HRMEC分为对照组、高糖组、高糖+胃饥饿素组,分别采用常规培养基、含30 mmol/L D-葡萄糖培养基、含30 mmol/L D-葡萄糖+10 nmol/L胃饥饿素培养基培养24 h。采用细胞计数试剂盒8检测细胞增生情况;采用流式细胞术检测细胞活性氧簇(ROS)水平;采用试剂盒检测细胞中氧化应激指标还原型谷胱甘肽(GSH)浓度、丙二醛(MDA)浓度、超氧化物歧化酶(SOD)活性及Fe^(2+)浓度;采用透射电子显微镜观察线粒体结构;采用Western blot法检测HRMEC中铁死亡关键分子谷胱甘肽过氧化合物酶4(GPX4)、溶质载体家族7成员11(SLC7A11)蛋白表达。结果对照组、高糖组、高糖+胃饥饿素组细胞增生率分别为(100.62±3.40)%、(63.74±4.25)%和(88.19±4.65)%,ROS荧光强度分别为15512.20±1347.53、46457.00±1072.65和22220.87±1669.20,GSH浓度分别为(68.52±7.61)、(21.45±1.57)和(55.68±5.15)μmol/L,MDA浓度分别为(0.79±0.10)、(2.47±0.27)和(1.08±0.15)μmol/L,SOD活性分别为(111.67±10.32)、(37.75±5.92)和(97.45±9.12)U/ml,Fe^(2+)浓度分别为(3.02±0.30)、(9.45±0.71)和(4.63±0.32)mmol/mgprot。各组间细胞增生率、ROS荧光强度、GSH浓度、MDA浓度、SOD活性及Fe^(2+)浓度总体比较,差异均有统计学意义(F=61.82、414.59、61.28、67.24、61.64,146.14,均P<0.001);与对照组相比,高糖组细胞增生率、GSH浓度和SOD活性均明显降低,ROS荧光强度、MDA浓度和Fe^(2+)浓度明显升高,差异均有统计学意义(均P<0.05);与高糖组相比,高糖+胃饥饿素组细胞增生率、GSH浓度和SOD活性明显升高,ROS荧光强度、MDA浓度和Fe^(2+)浓度明显降低,差异均有统计学意义(均P<0.05)。与对照组相比,高糖组细胞内线粒体铁死亡现象明显;与高糖组相比,高糖+胃饥饿素组线粒体状态明显改善。各组间细胞中GPX4、SLC7A11蛋白相对表达量总体比Objective To investigate the effects of ghrelin on oxidative stress and ferroptosis in retinal microvascular endothelial cells(HRMEC)under high glucose conditions.Methods HRMEC were divided into control group,high glucose group,high glucose+ghrelin group and cultured with conventional medium,30 mmol/L D-glucose medium,and 30 mmol/L D-glucose+10 nmol/L ghrelin medium in vitro for 24 hours accordingly.The cell proliferation was identified by cell counting kit-8 assay.The reactive oxygen species(ROS)levels were detected by flow cytometry.The oxidative stress indexes glutathione(GSH)concentration,malondialdehyde(MDA)superoxide dismutase(SOD)activity and Fe^(2+)concentration were detected by the corresponding kit.The mitochondrial structure was observed by transmission electron microscopy.The expression levels of glutathione peroxidase 4(GPX4)and recombinant solute carrier family 7 member 11(SLC7A11)proteins were detected by Western blot.Results The cell proliferation rates of control group,high glucose group and high glucose+ghrelin group were(100.62±3.40)%,(63.74±4.25)%and(88.19±4.65)%,respectively.The ROS fluorescence intensity of control group,high glucose group and high glucose+ghrelin group was 15512.20±1347.53,46457.00±1072.65 and 22220.87±1669.20,GSH concentration was(68.52±7.61),(21.45±1.57)and(55.68±5.15)μmol/L,MDA concentration was(0.79±0.10),(2.47±0.27)and(1.08±0.15)μmol/L,SOD activity was(111.67±10.32),(37.75±5.92)and(97.45±9.12)U/ml,Fe^(2+)concentration was(3.02±0.30),(9.45±0.71)and(4.63±0.32)mmol/mgprot,respectively.There were statistically significant differences in cell proliferation rate,ROS fluorescence intensity,GSH,MDA,and Fe^(2+)concentrations and SOD activity among the three groups(F=61.82,414.59,61.28,67.24,61.64,146.14;all at P<0.001).Compared with the control group,the cell proliferation rate,GSH concentration and SOD activity were reduced,ROS fluorescence intensity,MDA and Fe^(2+)concentrations were increased in the high glucose group,with statistically significant differ

关 键 词：胃饥饿素 铁死亡 氧化应激 高糖 人视网膜微血管内皮细胞 细胞增生 

分 类 号：R587.2[医药卫生—内分泌] R774.1[医药卫生—内科学]