香青兰总黄酮(TFDM)对高糖诱导的视网膜神经节细胞氧化损伤的影响及其机制  

Effect of total flavones of Dracocephalum moldavica L.on high-glucose induced oxidative damage to retinal ganglion cells and its mechanism

作　　者：谷李影 杨胜富 张启明 王书新 GU Liying;YANG Shengfu;ZHANG Qiming;WANG Shuxin(Ophthalmology Department of Wuhan Sixth Hospital,Jianghan University Affiliated Hospital,Wuhan 430014,Hubei Province,China;Ophthalmology Department of China Guihang Group 302 Hospital,Anshun 561000,Guizhou Province,China)

机构地区：[1]江汉大学附属医院武汉市第六医院眼科,湖北省武汉市430014 [2]中国贵航集团三〇二医院眼科,贵州省安顺市561000

出　　处：《眼科新进展》2024年第12期937-942,共6页Recent Advances in Ophthalmology

基　　金：贵州省卫生健康委科学技术基金项目(编号:gzwkj2022-300)。

摘　　要：目的探讨香青兰总黄酮(TFDM)对高糖诱导的视网膜神经节细胞(RGCs)氧化损伤的影响及其机制。方法本研究实验对象为小鼠RGCs,将RGCs接种于24孔板中(每孔2.5×10^(4)个),分为对照组(用含体积分数10%胎牛血清的培养基培养48 h)、高糖组(用含30 mmol·L^(-1)葡萄糖的培养基培养48 h)、高糖+TFDM-L组(用含30 mmol·L^(-1)葡萄糖、25 mg·L^(-1) TFDM的培养基共同培养48 h)、高糖+TFDM-M组(用含30 mmol·L^(-1)葡萄糖、50 mg·L^(-1) TFDM的培养基共同培养48 h)、高糖+TFDM-H组(用含30 mmol·L^(-1)葡萄糖、100 mg·L^(-1) TFDM的培养基共同培养48 h)、miR-NC组(细胞先转染miR-NC后用含30 mmol·L^(-1)葡萄糖的培养基培养48 h)、miR-93-5p组(细胞先转染miR-93-5p mimics后用含30 mmol·L^(-1)葡萄糖的培养基培养48 h)、anti-miR-NC组(细胞先转染anti-miR-NC后用含100 mg·L^(-1) TFDM、30 mmol·L^(-1)葡萄糖的培养基共同培养48 h)、anti-miR-93-5p组(细胞先转染anti-miR-93-5p后用含100 mg·L^(-1) TFDM、30 mmol·L^(-1)葡萄糖的培养基共同培养48 h)。依据试剂盒说明书检测各组RGCs氧化应激指标水平,采用硫代巴比妥酸法检测丙二醛(MDA)含量,比色法检测过氧化氢酶(CAT)、8-羟基脱氧鸟苷酸(8-OHdG)含量,流式细胞术检测细胞凋亡率,实时荧光定量PCR检测mRNA表达情况,双荧光素酶报告实验验证miR-93-5p、E2F转录因子1(E2F1)靶向调控作用,Western blot检测蛋白表达情况。对各组数据进行统计学分析。结果高糖组RGCs MDA、8-OHdG含量及细胞凋亡率、Bax蛋白表达水平均高于对照组(均为P<0.05);CAT含量、Bcl-2蛋白表达水平均低于对照组(均为P<0.05);高糖+TFDM-L组、高糖+TFDM-M组、高糖+TFDM-H组MDA、8-OHdG含量及细胞凋亡率、Bax蛋白表达水平均低于高糖组(均为P<0.05);CAT含量、Bcl-2蛋白表达水平均高于高糖组,且高糖+TFDM-L组、高糖+TFDM-M组、高糖+TFDM-H组随着TFDM浓度的增高各指标逐渐向对照组恢复(�Objective To investigate the effects of total flavones of Dracocephalum moldavica L.(TFDM)on oxidative damage to retinal ganglion cells(RGCs)induced by high glucose(HG)and its mechanism.Methods RGCs of mice were taken as the research subjects.RGCs were inoculated in 24-well plates(with 2.5×10^(4) cells in each well)and divided into control group(cultured with medium containing 10%fetal bovine serum for 48 h),HG group(cultured with medium containing 30 mmol·L^(-1) glucose for 48 h),HG+TFDM-L group(cultured with medium containing 30 mmol·L^(-1) glucose and 25 mg·L^(-1) TFDM for 48 h),HG+TFDM-M group(cultured with medium containing 30 mmol·L^(-1) glucose and 50 mg·L^(-1) TFDM for 48 h),HG+TFDM-H group(cultured with medium containing 30 mmol·L^(-1) glucose and 100 mg·L^(-1) TFDM for 48 h),miR-NC group(transfected with miR-NC and then cultured with medium containing 30 mmol·L^(-1) glucose for 48 h),miR-93-5p group(transfected with miR-93-5p mimics and then cultured with medium containing 30 mmol·L^(-1) glucose for 48 h),anti-miR-NC group(transfected with anti-miR-NC and then cultured with medium containing 100 mg·L^(-1) TFDM and 30 mmol·L^(-1) glucose for 48 h),and anti-miR-93-5p group(transfected with anti-miR-93-5p and then cultured with medium containing 100 mg·L^(-1) TFDM and 30 mmol·L^(-1) glucose for 48 h).Levels of RGCs oxidative stress indexes in each group were detected according to the kit instructions.The thiobarbituric acid method was used to measure the level of malondialdehyde(MDA),the colorimetric method was adopted to detect the levels of catalase(CAT)and 8-hydroxydeoxyguanosine(8-OHdG),apoptosis rate was detected by flow cytometry,the messenger ribonucleic acid(mRNA)expressions were analyzed by real-time quantitative polymerase chain reaction,the targeting regulation of miR-93-5p and E2F transcription factor 1(E2F1)were verified by dual luciferase reporter assay,and the protein expressions were detected by Western blot.Statistical analysis was conducted on the data of each group.Results

关 键 词：香青兰总黄酮 微小RNA-93-5p E2F转录因子1 高糖 视网膜神经节细胞 细胞凋亡 氧化应激 

分 类 号：R774[医药卫生—眼科]