骨形态发生蛋白7模拟肽THR123对PVR发生及RPE细胞EMT的调控作用及机制  

Effects and mechanisms of BMP7 mimetic peptide THR123 on regulating proliferative vitreoretinopathy and retinal pigment epithelial-mesenchymal transition

作　　者：姚海佩 王方 王于蓝[1] Yao Haipei;Wang Fang;Wang Yulan(Department of Ophthalmology,Shanghai Eye Diseases Prevention&Treatment Center/Shanghai Eye Hospital,School of Medicine,Tongji University,National Clinical Research Center for Eye Diseases,Shanghai Engineering Research Center of Precise Diagnosis and Treatment of Eye Diseases,Shanghai 200040,China;Shanghai Bright Eye Hospital,Shanghai 200050,China)

机构地区：[1]上海市眼病防治中心/同济大学附属眼科医院眼科、国家眼部疾病临床医学研究中心、上海市眼科疾病精准诊疗工程技术研究中心,上海200040 [2]上海普瑞眼科医院,上海200050

出　　处：《中华实验眼科杂志》2025年第1期18-26,共9页Chinese Journal Of Experimental Ophthalmology

基　　金：上海市“科技创新行动计划”自然科学基金(21ZR1459300)。

摘　　要：目的探讨骨形态发生蛋白(BMP)7模拟肽THR123对增生性玻璃体视网膜病变(PVR)发生及视网膜色素上皮(RPE)细胞上皮-间充质转化(EMT)的调控作用及机制。方法取分离自供体眼球的人原代RPE细胞,将其分为阴性对照组、转化生长因子β2(TGF-β2)刺激组和THR123治疗组,分别用常规培养基、含10 ng/ml TGF-β2培养基、含10 ng/ml TGF-β2和5μg/ml THR123培养基培养48 h。采用Western blot法检测各组EMT标志物E-钙粘素(E-Cadherin)、纤连蛋白(FN)、α平滑肌肌动蛋白(α-SMA)和BMP受体激活素受体样激酶2(ALK2)及转录因子锌指蛋白转录因子1(Snail1)表达;采用凝胶收缩实验检测各组RPE收缩功能差异;采用Transwell实验检测各组RPE细胞迁移能力;采用跨上皮电阻检测RPE细胞上皮极性变化。取成年青紫蓝兔30只,采用玻璃体腔注射RPE细胞及血小板源性生长因子建立兔PVR模型,按照随机数字表法分为PVR组、0.5μg/ml THR123组和5μg/ml THR123组分别在造模日及造模后7 d进行右眼相应浓度THR123玻璃体腔注射,采用检眼镜每7 d进行1次眼底观察,采用B型超声确认有无视网膜脱离,分别于造模后14和28 d进行PVR分级,造模后28 d空气栓塞法处死模型兔后取眼球行苏木精-伊红染色及α-SMA免疫荧光染色,比较各组PVR进展差异。原代RPE细胞中加入BMP受体抑制剂LDN193189作为LDN抑制组,检测阴性对照组和LDN抑制组RPE细胞EMT标志物及功能差异;在RPE细胞中分别转染对照和Snail1的小干扰RNA作为siNC+TGF-β2组和siSnail1+TGF-β2组,检测各组RPE细胞EMT标志物及功能差异。结果阴性对照组、TGF-β2刺激组和THR123治疗组凝胶收缩比例、跨上皮电阻值及E-Cadherin、FN和α-SMA蛋白相对表达量总体比较,差异均有统计学意义(F=28.38、136.30、38.50、2.53、53.54,均P<0.01)。与阴性对照组和THR123治疗组比较,TGF-β2刺激组凝胶收缩比例、跨上皮电阻值、E-Cadherin相对表达量降低,α-SMA�Objective To investigate the role of bone morphogenetic protein(BMP)7 peptidomimetics THR123 in the regulation of proliferative vitreoretinopathy(PVR)and epithelial-mesenchymal transition(EMT)in retinal pigment epithelial(RPE)cells.Methods Primary human RPE cells were isolated from donor eyes.The RPE cells was cultured for 48 hours with conventional medium,medium containing 10 ng/ml transforming growth factor-β2(TGF-β2),medium containing 10 ng/ml TGF-β2 and 5μg/ml THR123,respectively,serving as negative control group,TGF-β2 stimulation group and THR123-treated group.The relative protein expression levels of E-cadherin,α-smooth muscle actin(α-SMA),fibronectin(FN)activin receptor-like kinase-2(ALK2)and Snail family zinc finger transcription factor 1,Snail1)were detected by Western blot analysis in each group.RPE cell contractile function was detected by collagen gel contraction assay.RPE cell migration function was detected by transwell assay.RPE cell polarity was detected by transepithelial resistance.For the in vivo model,the rabbit PVR model was established by intravitreal injection of RPE cells and cytokine platelet-derived growth factor.The PVR rabbits were randomly divide into PVR group,0.5μg/ml THR123 group and 5μg/ml THR123 group,and were intravitreally injected with corresponding concentration of THR123 on the day of modelling and 7 days after modelling.The fundus conditions of rabbits were observed every 7 days.Retinal detachment was checked by B-scan ultrasonography every 7 days,and PVR grading was performed on 14 and 28 days after modelling.On day 28 after modelling,the rabbit was sacrificed by air embolism and the eyeball was enucleated.Hematoxylin-eosin(HE)staining andα-SMA immunofluorescence staining were performed to compare the differences in PVR progression among groups.In the LDN inhibition group,BMP receptor inhibitor LDN193189 was added to primary RPE cells and the expression of EMT markers and cell functional differences of RPE cells were compared between negative control group and L

关 键 词：骨形态发生蛋白 增生性玻璃体视网膜病变 视网膜色素上皮 上皮-间充质转化 

分 类 号：R774.1[医药卫生—眼科]