m6A去甲基化酶ALKBH5在形觉剥夺性近视豚鼠中的表达变化及其意义  

Changes in m6A demethylase ALKBH5 expression in form-deprivation myopic guinea pigs and its significance

作　　者：向小玲 廖萱[1] 孟茄玉 周桂梅 周红 周羡媚 Xiang Xiaoling;Liao Xuan;Meng Jiayu;Zhou Guimei;Zhou Hong;Zhou Xianmei(Department of Ophthalmology,Affiliated Hospital of North Sichuan Medical College,Medical School of Ophthalmology&Optometry,North Sichuan Medical College,Nanchong 637000,China;The Sichuan Provincial Key Laboratory for Human Disease Gene Study,the Center for Medical Genetics,Sichuan Provincial People's Hospital,School of Medicine,University of Electronic Science and Technology of China,Chengdu 611731,China)

机构地区：[1]川北医学院附属医院眼科、川北医学院眼视光医学院,南充637000 [2]电子科技大学医学院、四川省人民医院医学遗传中心、四川省人类疾病基因研究重点实验室,成都611731

出　　处：《中华实验眼科杂志》2024年第4期315-321,共7页Chinese Journal Of Experimental Ophthalmology

基　　金：四川省科技厅自然科学基金(2023NSFSC0595);南充市市校科技战略合作项目(22SXFWDF0003);川北医学院附属医院科研发展计划重点项目(2023ZD010)。

摘　　要：目的探讨形觉剥夺性近视(FDM)豚鼠视网膜中m6A去甲基化酶AlkB同源蛋白5(ALKBH5)表达变化及其参与近视的作用机制。方法采用随机数字表法将30只健康SPF级3周龄三色豚鼠分为正常对照组和实验组,每组15只。实验组中眼罩遮盖右眼作为FDM组,暴露左眼为自身对照组。分别于实验前及实验1、2、3和4周时进行豚鼠眼生物学参数测量。采用带状光检影镜测量屈光度,采用A型超声仪测量眼轴长度。实验4周,通过免疫组织化学染色和免疫荧光染色检测ALKBH5在豚鼠视网膜中的表达分布。采用实时荧光定量PCR和Western blot法检测豚鼠视网膜中ALKBH5 mRNA和蛋白表达情况。结果与正常对照组和自身对照组相比,实验2、3和4周,FDM组豚鼠近视屈光度明显增加,眼轴显著增长,差异均有统计学意义(均P<0.001)。免疫组织化学染色和免疫荧光染色显示,ALKBH5分布在视网膜神经纤维层、视锥视杆细胞层和视网膜色素上皮(RPE)层,其中以神经纤维层和RPE层为主。正常对照组、自身对照组和FDM组豚鼠ALKBH5蛋白相对荧光强度值分别为1.000±0.204、0.874±0.076和0.571±0.053,FDM组视网膜中ALKBH5蛋白荧光强度值明显小于正常对照组和自身对照组,差异均有统计学意义(t=4.069,P=0.006;t=5.176,P=0.014)。造模后4周,FDM组豚鼠视网膜中ALKBH5 mRNA和蛋白相对表达量明显低于正常对照组和自身对照组,差异均有统计学意义(均P<0.01)。结论FDM组豚鼠视网膜中m6A去甲基化酶ALKBH5表达下降,ALKBH5及相关m6A甲基化修饰可能参与了近视的发生和发展。Objective To investigate the alteration of m6A demethylase AlkB homolog 5(ALKBH5)expression and its impact on form-deprivation myopia(FDM)retina in guinea pigs.Methods Thirty normal SPF grade 3-week-old tricolor guinea pigs were randomly divided into normal control group and experimental group,with 15 in each group.In the experimental group,the right eyes were covered as FDM group and the left eyes uncovered were set as self-control group.Ocular biometry was performed at one-week intervals from baseline to week 4 of the experiment.Spherical equivalent was detected by streak retinoscopy and axial length was measured by A-scan ultrasonography.Animals were sacrificed after 4 weeks of modeling.The distribution and expression of ALKBH5 protein in the guinea pig retina was detected by immunohistochemical and immunofluorescence staining.Expression of ALKBH5 mRNA and protein in guinea pig retina was detected by real-time fluorescence quantitative PCR and Western blot,respectively.The use of animals in ophthalmic and vision research followed the tenets of Animal Research in Vision and Ophthalmology,and the study was approved by the Ethics Committee of North Sichuan Medical College(No.2023087).Results At weeks 2,3,and 4 after myopia induction,diopters and axial lengths were significantly higher in the FDM group than in the normal control group and the self-control group(all at P<0.001).Immunohistochemistry and immunofluorescence assays showed that ALKBH5 protein was expressed in the retinal nerve fiber layer,rod/cone photoreceptor cells,and retinal pigment epithelium(RPE)layer,and was highly expressed in the retinal nerve fiber layer and RPE layer.The relative ALKBH5 immunofluorescence intensity in the normal control group,self-control group and FDM group was 1.000±0.204,0.874±0.076 and 0.571±0.053,respectively,which was lower in the FDM group than in the normal control and self-control groups,showing statistically significant differences(t=4.069,P=0.006;t=5.176,P=0.014).After 4 weeks of modeling,ALKBH5 mRNA and protein

关 键 词：近视 豚鼠 AlkB同源蛋白5 RNA去甲基酶 m6A甲基化 发病机制 

分 类 号：R778.11[医药卫生—眼科]