SOX9通过转化生长因子β信号通路调节角膜内皮损伤后内皮-间质转化过程

SOX9 Regulates Endothelial-mesenchymal Transition After Corneal Endothelial Injury Through Transforming Growth Factor-βSignaling Pathway

目的探究性别决定区Y框蛋白9(sex-determining region Y-box protein 9,SOX9)是否会调控角膜内皮细胞损伤后的角膜上皮-间质转化(endothelial-to-mesenchymal transition,EndMT)过程及具体机制。方法转染小干扰RNA(small interfering RNA,siRNA)敲低人角膜内皮细胞B4G12中SOX9的表达,使用甲萘醌构建体外B4G12细胞损伤模型,设4个分组:si-NC组(转染siRNA-阴性对照)、si-SOX9组(转染siRNA-SOX9)、si-NC+甲萘醌组(转染siRNA-阴性对照后添加外源性甲萘醌做损伤处理)、si-SOX9+甲萘醌组(转染siRNA-SOX9后添加外源性甲萘醌做损伤处理)。通过实时荧光逆转录聚合酶链反应(real-time reverse transcription polymerase chain reaction,RT-PCR)和Western blot检测各组细胞中EndMT关键因子Snail家族转录抑制因子2(snail family transcriptional repressor 2,SNAIL2)及相关信号通路关键因子表达变化,阐明SOX9调控EndMT过程的功能和机制。结果转染siRNA-SOX9敲低细胞SOX9表达后,给予甲萘醌细胞损伤处理,观察到细胞中SNAIL2的表达会随SOX9的敲低而降低,同时观察到转化生长因子β(transforming growth factor beta,TGF-β)信号通路中SNAIL2的上游关键因子Smad2/Smad3的表达也随着SOX9的敲低而降低。结论SOX9通过TGF-β信号通路调控角膜内皮损伤后EndMT过程。

Objective To investigate whether sex-determining region Y-box protein 9(SOX9)regulates endotheli-al-to-mesenchymal transition(EndMT)after corneal endothelial cell injury and its specific mechanism.Methods Small interfering RNA(siRNA)was transfected to knock down SOX9 expression in human corneal endothelial cells B4G12,and menadione was used to construct B4G12 cell injury model in vitro,4 groups were set up:si-NC group(transfection of siRNA-negative control),si-SOX9 group(transfection of siRNA-SOX9),si-NC+menadione group(after transfection of siRNA-negative control,exogenous menadione was added for injury treatment);si-SOX9+menadione group(after transfection of siRNA-SOX9,exogenous menadione was added for injury treatment).The expression changes of snail family transcriptional repressor 2(SNAIL2)and related signaling pathway key factors of EndMT in each group of B4G12 cells were detected by real-time reverse transcription polymerase chain reaction(RT-PCR)and Western blot to clarify the function and mechanism of SOX9 regulating EndMT process.Results After transfection of siRNA-SOX9 to knock down the expression of SOX9,it was given menadione cell damage treatment,the expression of SNAIL2 in the cells decreased along with the knockdown of SOX9 after injury,at the same time,the expression of Smadt2/Smad3,which were upstream of SNAIL2 in the transforming growth factor beta(TGF-β)signaling pathway,was also decreased in response to SOX9 knockdown.Conclusion SOX9 regulates the EndMT process after corneal endothelial injury through TGF-βsignaling pathway.