人巩膜成纤维细胞中miR-184表达与近视发生、发展的关系及其对PI3K-Akt信号通路的调控作用  

On the micro ribonucleic acid 184 expression in human scleral fibroblasts’correlation with the occurrence and development of myopia as well as its regulation on phosphatidylinositol 3-kinase/protein kinase B signaling pathway

作　　者：张悦之 邓燕[1] 殷小龙[1] 熊伟伟[1] 于春红[1] ZHANG Yuezhi;DENG Yan;YIN Xiaolong;XIONG Weiwei;YU Chunhong(Eye Center of the Second Affiliated Hospital of Nanchang University,Nanchang 330006,Jiangxi Province,China)

机构地区：[1]南昌大学第二附属医院眼科中心,江西省南昌市330006

出　　处：《眼科新进展》2023年第6期444-448,共5页Recent Advances in Ophthalmology

基　　金：江西省自然科学基金项目(编号:20212BAB206049)。

摘　　要：目的探讨人巩膜成纤维细胞中miR-184表达与近视发生、发展的关系及其对磷脂酰肌醇3-激酶/蛋白激酶B(PI3K-Akt)信号通路的调控作用。方法利用形觉剥夺法建立豚鼠左眼(实验眼)近视模型,右眼为对照眼,采用实时定量PCR法检测豚鼠实验眼巩膜组织中miR-184以及PI3K-Akt信号通路相关mRNA的表达水平,采用Pearson相关分析豚鼠实验眼巩膜组织中miR-184与PI3K、Akt、雷帕霉素机械靶蛋白(mTOR)mRNA表达水平的相关性。体外培养人巩膜成纤维细胞,并分为对照组(mimics对照组)、高表达组(转染miR-184 mimic)、低表达组(转染anti-miR-184)和PI-103组(加入PI3K通路抑制剂PI-103)。采用Western blot检测各组人巩膜成纤维细胞中PI3K-Akt信号通路相关蛋白表达水平,流式细胞检测技术检测细胞增殖,Annexin V-FITC染色观察细胞凋亡。结果豚鼠实验眼巩膜组织中miR-184与PI3K、Akt、mTOR mRNA表达水平均明显低于对照眼(均为P<0.05),且实验眼巩膜组织中miR-184与PI3K、Akt、mTOR mRNA表达水平均呈正相关性(均为P<0.05)。体外实验中,培养24 h、48 h、72 h、96 h时,高表达组人巩膜成纤维细胞增殖倍数>对照组>低表达组>PI-103组;培养24 h时,4组人巩膜成纤维细胞增殖倍数比较差异无统计学意义(P>0.05);培养48 h、72 h、96 h时,4组人巩膜成纤维细胞增殖倍数整体比较以及组间两两比较差异均有统计学意义(均为P<0.05)。培养24 h时,4组人巩膜成纤维细胞凋亡率差异有统计学意义(P<0.05),PI-103组>低表达组>对照组>高表达组,组间两两比较差异均有统计学意义(均为P<0.05)。培养72 h时,4组人巩膜成纤维细胞中PI3K、Akt、mTOR蛋白表达水平差异均有统计学意义(均为P<0.05),高表达组人巩膜成纤维细胞中PI3K、Akt、mTOR蛋白表达水平>对照组>低表达组>PI-103组,组间两两比较差异均有统计学意义(均为P<0.05)。结论近视眼的发生、发展与巩膜组织中miR-184表达下�Objective To investigate the relationship between the expression of micro ribonucleic acid 184(miR-184)in human scleral fibroblasts and the occurrence and development of myopia,as well as the regulatory effect of miR-184 on the phosphatidylinositol 3-kinase/protein kinase B(PI3K-Akt)signaling pathway.Methods The form deprivation method was used to establish the myopia model in the left eye(experimental eye)of guinea pigs,and the right eye was taken as the control eye.The expression levels of miR-184 and messenger RNA(mRNA)of PI3K-Akt signal pathway in the scleral tissues of experimental eyes were detected by quantitative PCR.Pearson correlation analysis was used to analyze the correlation between miR-184 and the mRNA expressions of PI3K,Akt and mammalian target of rapamycin(mTOR)in the scleral tissues of experimental eyes.Human scleral fibroblasts were cultured in vitro and divided into the control group(mimics control group),miR-184 high expression group(transfected with miR-184 mimic),miR-184 low expression group(transfected with anti-miR-184),and PI-103 group(added with PI-103,which is a PI3K signaling pathway inhibitor).Western blot was used to detect the expression of PI3K-Akt signaling pathway-related proteins in the transfected human scleral fibroblasts.Flow cytometry was adopted to detect cell proliferation.Annexin V-FITC staining was used to observe cell apoptosis.Results The expression of miR-184 and mRNA expressions of PI3K,Akt and mTOR in the scleral tissues of the experimental eyes were significantly lower than those of the control eyes(all P<0.05).The expression of miR-184 was positively correlated with the mRNA expressions of PI3K,Akt and mTOR in the scleral tissues of the experimental eyes(all P<0.05).The in vitro experiments showed that the proliferation rate of human scleral fibroblasts at 24 h,48 h,72 h and 96 h after cultivation decreased sequentially in the miR-184 high expression group,control group,miR-184 low expression group and PI-103 group.At 24 h after cultivation,there was no signific

关 键 词：近视 巩膜组织 人巩膜成纤维细胞 磷脂酰肌醇3-激酶/蛋白激酶B 细胞凋亡 

分 类 号：R778.1[医药卫生—眼科]