He Guang-Hui, Zhang Wei, Ma Ying-Xue, Yang Jing, Chen Li, Song Jian, Chen Song
Tianjin Eye Hospital; Tianjin Key Lab of Ophthalmology and Visual Science; Tianjin Eye Institute; Clinical College of Ophthalmology, Tianjin Medical University, Tianjin 300020, China.
Int J Ophthalmol. 2018 Apr 18;11(4):559-566. doi: 10.18240/ijo.2018.04.04. eCollection 2018.
To observe the effect of exosomes derived from human umbilical cord blood mesenchymal stem cells (hUCMSCs) on the expression of vascular endothelial growth factor-A (VEGF-A) in blue light injured human retinal pigment epithelial (RPE) cells and laser-induced choroidal neovascularization (CNV) in rats.
Exosomes were isolated from hUCMSCs and characterized by transmission electron microscope and Western blot. MSCs-derived exosomes were cultured with RPE cells exposed to blue light. The mRNA and protein expression of VEGF-A were determined by real time-polymerase chain reaction (PCR) and Western blot, respectively. Immunofluorescence assay was used for the detection of the expression level of VEGF-A. We injected different doses of MSCs-derived exosomes intravitreally to observe and compare their effects in a mouse model of laser-induced retinal injury. The histological structure of CNV in rats was inspected by hematoxylin-eosin (HE) staining and fundus fluorescein angiography. The expression of VEGF-A was detected by immunohistochemistry.
Exosomes exhibited the typical characteristic morphology (cup-shaped) and size (diameter between 50 and 150 nm). The exosomes marker, CD63, and hUCMSCs marker, CD90, showed a robust presence. , MSCs-derived exosomes downregulated the mRNA(Exo-L: =6.485, 7.959, 9.286; Exo-M: =7.517, 10.170, 13.413; Exo-H: =10.317, 12.234, 14.592, <0.05) and protein (Exo-L: =2.945, 4.477, 6.657; Exo-M: =4.713, 6.421, 8.836; Exo-H: =6.539, 12.194, 12.783; <0.05) expression of VEGF-A in RPE cells after blue light stimulation. , we found that the MSCs-derived exosomes reduced damage, distinctly downregulated VEGF-A (Exo-H: =0.957, 1.382; <0.05), and gradually improved the histological structures of CNV for a better visual function (Exo-L: 0.346, Exo-M: 3.382, Exo-H: 8.571; <0.05).
MSCs-derived exosomes ameliorate blue light stimulation in RPE cells and laser-induced retinal injury downregulation of VEGF-A.
观察人脐带血间充质干细胞（hUCMSCs）来源的外泌体对蓝光损伤的人视网膜色素上皮（RPE）细胞中血管内皮生长因子-A（VEGF-A）表达及大鼠激光诱导脉络膜新生血管（CNV）的影响。
从hUCMSCs中分离外泌体，通过透射电子显微镜和蛋白质免疫印迹进行鉴定。将MSCs来源的外泌体与暴露于蓝光的RPE细胞共培养。分别通过实时聚合酶链反应（PCR）和蛋白质免疫印迹法测定VEGF-A的mRNA和蛋白表达。采用免疫荧光法检测VEGF-A的表达水平。向玻璃体内注射不同剂量的MSCs来源的外泌体，在激光诱导的视网膜损伤小鼠模型中观察并比较其效果。通过苏木精-伊红（HE）染色和眼底荧光血管造影检查大鼠CNV的组织结构。采用免疫组织化学法检测VEGF-A的表达。
外泌体呈现典型的特征形态（杯状）和大小（直径在50至150nm之间）。外泌体标志物CD63和hUCMSCs标志物CD90均有显著表达。MSCs来源的外泌体下调了蓝光刺激后RPE细胞中VEGF-A的mRNA（Exo-L：=6.485，7.959，9.286；Exo-M：=7.517，10.170，13.413；Exo-H：=10.317，12.234，14.592，<0.05）和蛋白（Exo-L：=2.945，4.477，6.657；Exo-M：=4.713，6.421，8.836；Exo-H：=6.539，12.194，12.783；<0.05）表达。我们发现MSCs来源的外泌体减少了损伤，显著下调了VEGF-A（Exo-H：=0.957，1.382；<0.05），并逐渐改善了CNV的组织结构，以获得更好的视觉功能（Exo-L：0.346，Exo-M：3.382，Exo-H：8.571；<0.05）。
MSCs来源的外泌体改善了RPE细胞中的蓝光刺激和激光诱导的视网膜损伤——VEGF-A的下调。
