基于miR⁃224⁃5P调控的IL6ST/JAK/STAT信号通路在糖尿病视网膜病变发生发展中的作用  

Effect of micro⁃ribonucleic acid⁃224⁃5P in the occurrence and development of diabetes retinopathy via regulating interleukin⁃6 signal transducer/Janus⁃ac⁃tivated kinase/signal transducer and activator of transcription signaling path⁃way

作　　者：赵鑫 杨静[1] 辛向阳[2] ZHAO Xin;YANG Jing;XIN Xiangyang(School of Basic Medicine and Forensic Medicine,Inner Mongolia University of Science and Technology Baotou Medical College,Baotou 014040,Inner Mongolia,China;Department of Ophthalmology,Inner Mongolia Baogang Hospital,Baotou 014010,Inner Mongolia,China)

机构地区：[1]内蒙古科技大学包头医学院基础医学与法医学院,内蒙古自治区包头市014040 [2]内蒙古包钢医院眼科,内蒙古自治区包头市014010

出　　处：《眼科新进展》2023年第5期357-362,共6页Recent Advances in Ophthalmology

基　　金：内蒙古自治区卫生健康科技计划项目(编号:202201522)。

摘　　要：目的探讨miR⁃224⁃5P在糖尿病视网膜病变(DR)患者中的表达变化及其在高糖诱导的人视网膜色素上皮细胞(ARPE⁃19)中的作用和机制。方法抽取DR患者及健康人群各6名的外周血,RT⁃qPCR检测miR⁃224⁃5P相对表达量。将正常培养的ARPE⁃19细胞随机分成高糖组(给予30 mmol·L^(-1)高糖)、高糖+miR⁃224⁃5P mimics组(转染100 nmol·L^(-1) miR⁃224⁃5P mimics后给予30 mmol·L^(-1)高糖)、高糖+miR⁃224⁃5P inhibitor组(转染100 nmol·L^(-1) miR⁃224⁃5P inhibitor后给予30 mmol·L^(-1)高糖)、高糖+OE⁃IL6ST组(转染100 nmol·L^(-1) OE⁃IL6ST后给予30 mmol·L^(-1)高糖)和高糖+miR⁃224⁃5P mimics+OE⁃IL6ST组(转染100 nmol·L^(-1) miR⁃224⁃5P mimics和100 nmol·L^(-1) OE⁃IL6ST后给予30 mmol·L^(-1)高糖),CCK⁃8检测细胞活力,Transwell检测细胞迁移率,双荧光素酶实验验证miR⁃224⁃5P与IL6ST的靶向关系,Western blot检测IL6ST、P⁃JAK及P⁃STAT蛋白表达,ELISA检测细胞中IL⁃1β、IL⁃6及TNF⁃α蛋白表达水平。通过过表达IL6ST进一步验证miR⁃224⁃5P是否通过IL6ST/JAK/STAT通路发挥作用。结果DR患者外周血中miR⁃224⁃5P mRNA相对表达量显著低于健康人群(P<0.001)。与高糖组相比,高糖+miR⁃224⁃5P mimics组细胞活力、细胞迁移率增高,高糖+miR⁃224⁃5P inhibitor组细胞活力、细胞迁移率降低(均为P<0.001)。双荧光素酶实验检测结果证实IL6ST是miR⁃224⁃5P调控的靶基因。与高糖组相比,高糖+miR⁃224⁃5P mimics组细胞的IL6ST蛋白表达水平降低,P⁃JAK和P⁃STAT蛋白表达水平升高(均为P<0.05);高糖+miR⁃224⁃5P inhibitor组细胞的IL6ST蛋白表达水平升高(P<0.05),P⁃JAK和P⁃STAT蛋白表达水平降低(均为P<0.05)。与高糖组相比,高糖+miR⁃224⁃5P mimics组细胞的IL⁃1β、IL⁃6及TNF⁃α蛋白表达水平均下降(均为P<0.01),高糖+miR⁃224⁃5P inhibitor组细胞的IL⁃1β、IL⁃6及TNF⁃α蛋白表达水平均显著上升(均为P<0.Objective To investigate the expression changes of micro⁃ribonucleic acid⁃224⁃5P(miR⁃224⁃5P)in dia⁃betes retinopathy(DR)patients and its effect and mechanism in high glucose⁃induced adult retinal pigment epithelial cell⁃19(ARPE⁃19 cells).Methods The peripheral blood was extracted from 6 DR patients and healthy people,respectively.Re⁃verse transcription⁃quantitative polymerase chain reaction was used to detect the relative expression of miR⁃224⁃5P.The normally cultured ARPE⁃19 cells were randomly divided into the high glucose group(administered with 30 mmol·L^(-1) high glucose),high glucose+miR⁃224⁃5P mimics group(administered with 30 mmol·L^(-1) high glucose after transfection with 100 nmol·L^(-1) miR⁃224⁃5P mimics),high glucose+miR⁃224⁃5P inhibitor group(administered with 30 mmol·L^(-1) high glu⁃cose after transfection with 100 nmol·L^(-1) miR⁃224⁃5P inhibitor),high glucose+OE⁃IL6ST group(administered with 30 mmol·L^(-1) high glucose after transfection with 100 nmol·L^(-1) OE⁃IL6ST),and high glucose+miR⁃224⁃5P mimics+OE⁃IL6ST group(administered with 30 mmol·L^(-1) high glucose after transfection with 100 nmol·L^(-1) miR⁃224⁃5P mimics and 100 nmol·L^(-1) OE⁃IL6ST).Cell viability was analyzed by Cell Counting Kit⁃8;cell migration rate was detected by Tran⁃swell;targeting relationship between miR⁃224⁃5P and interleukin⁃6 signal transducer(IL6ST)was verified by dual⁃luciferase assay;Western blot was used to determine the protein expressions of IL6ST,phosphorylated Janus⁃activated kinase(P⁃JAK)and phosphorylated signal transducer and activator of transcription(P⁃STAT);the protein expressions of interleukin(IL)⁃1β,IL⁃6 and tumor necrosis factor⁃α(TNF⁃α)were detected by enzyme⁃linked immunosorbent assay.Additionally,the IL6ST was overexpressed to confirm whether miR⁃224⁃5P functioned through IL6ST/JAK/STAT signaling pathway.Re⁃sults The relative messenger ribonucleic acid(mRNA)expression of miR⁃224⁃5P in the

关 键 词：糖尿病视网膜病变 miR⁃224⁃5P IL6ST/JAK/STAT通路 

分 类 号：R774.1[医药卫生—眼科]