年龄相关性白内障患者miR-138的表达及其对人晶状体上皮细胞凋亡的影响  

Expression of mi R-138 and its effects on human lens epithelial cells apoptosis in age-related cataract patients

作　　者：陆博[1] 马立威[1] 王欣玲[1] 冯莉[1] 江陵峰 王春霞[1] 张劲松[1] 阎启昌[1] LU Bo;MA Li-Wei;WANG Xin-Ling;FENG Li;JIANG Ling-Feng;WANG Chun-Xia;ZHANG Jin-Song;YAN Qi-Chang(Department of Ophthalmology,the Fourth Affiliated Hospital of China Medical University,Key Labora-tory of Lens Research of Liaoning Province,Eye Hospital of China Medical University,Shenyang 110005,Liaoning Province,China)

机构地区：[1]中国医科大学附属第四医院眼科,辽宁省晶状体学重点实验室,中国医科大学眼科医院,辽宁省沈阳市110005

出　　处：《眼科新进展》2018年第2期111-115,共5页Recent Advances in Ophthalmology

基　　金：国家自然科学基金资助(编号:81170836;81570838);辽宁省自然科学基金资助(编号:2015020474)~~

摘　　要：目的检测miR-138在年龄相关性白内障晶状体组织中的表达情况,并探讨miR-138对人晶状体上皮细胞增殖和凋亡的影响及其可能的靶基因。方法采用实时荧光定量PCR(real-time quantitative polymerase chain reaction,RT-qP CR)检测年龄相关性白内障患者(白内障组)与正常对照组中miR-138和预测靶基因沉默信息调节因子1(silent in-formation regulator 1,SIRT1)氉的表达水平。向人晶状体上皮细胞系(SRA01/04)细胞中分别转染miR-138模拟物、模拟物阴性对照物、miR-138抑制物、抑制物阴性对照物,采用RT-qP CR检测SIRT1的mR NA表达,Western blotting检测SIRT1的蛋白表达水平;转染72 h后细胞暴露于200μmol·L-1H2O21 h,Caspase-3活性检测试剂盒检测Caspase-3活性。双荧光素酶报告基因检测验证miR-138与SIRT1的靶向关系。结果与正常对照组相比,白内障组miR-138的表达(3.64±0.19)显著升高(P<0.001),SIRT1的mR NA表达(0.32±0.06)显著下降(P<0.001);相对于模拟物阴性对照组,miR-138模拟物组的SIRT1的mRNA(0.42±0.05)及蛋白(0.46±0.05)表达水平均明显降低,Caspase-3活性(3.24±0.17)明显升高(均为P<0.05);相对于抑制物阴性对照组,miR-138抑制物组的SIRT1的mR NA(2.95±0.13)及蛋白(1.98±0.12)表达水平均明显升高,Caspase-3活性(0.42±0.05)均明显下降(均为P<0.05);双荧光素酶报告基因检测确证SIRT1为miR-138的直接作用靶点。结论 miR-138在年龄相关性白内障患者晶状体囊膜中高表达,miR-138通过靶向性负性调控SIRT1表达,促进晶状体上皮细胞凋亡。Objective To detect the expression of miR-138 in lens tissues of age-related cataract and explore the effects of miR-138 on the proliferation and apoptosis of human lens epithelial cells and its possible target genes.Methods Real-time quantitative PCR(RT-qPCR)was applied for the detection of the expression of miR-138 and prediction of target gene sirtuin(silent information regulator 1)(SIRT1)in patients with age-related cataract(cataract group)and anterior lens capsules(normal control group).Then miR-138 mimics,mimic controls,miR-138 inhibitors and inhibitor controls were transfected into the human lens epithelial cell line(SRA01/04),and the expression of SIRT1 mRNA and protein was detected by RT-qPCR and Western blot,accordingly.At 72 hours after transfection,the cells were exposed to 200μmol·L-1 H 2O 2 for 1 hour,followed by detection of the activity of Caspase-3 by the Caspase-3 activity assay kit,and identification of the targeted relationship between miR-138 and SIRT1 by dual luciferase reporter assays.Results Compared with the normal control group,the expression of miR-138(3.64±0.19)was significantly increased(P<0.001),but the expression of SIRT1 mRNA(0.32±0.06)was significantly decreased(P<0.001)in the cataract group.Moreover,The expression levels of SIRT1 mRNA(0.42±0.05)and protein(0.46±0.05)in cells transfected with miR-138 mimics were significantly decreased,while the activity of Caspase-3(3.24±0.17)was significantly elevated when compared with cells transfected with minic controls(all P<0.05);Compared with cells transfected with inhibitor controls,the expressions of SIRT1 mRNA(2.95±0.13)and protein(1.98±0.12)were significantly upregulated,whereas Caspase-3 activity(0.42±0.05)was significantly decreased in cells transfected with miR-138 inhibitors(all P<0.05).And finally,dual luciferase reporter assays showed the confirmation SIRT1 as a direct target of miR-138.Conclusion miR-138 is highly expressed in the lens capsule of age-related cataract patients,and it can promote the apoptosis of lens e

关 键 词：miR-138 沉默信息调节因子1 年龄相关性白内障 凋亡 增殖 

分 类 号：R776.1[医药卫生—眼科]