Abstract：
Diabetic retinopathy (DR) is a highly hazardous and widespread complication of diabetes mellitus (DM). The accumulated reactive oxygen species (ROS) play a central role in DR development. The aim of this research was to examine the impact and mechanisms of mesenchymal stem cell (MSC)-derived small extracellular vesicles (sEV) on regulating ROS and retinal damage in DR. Intravitreal injection of sEV inhibited Cullin3 neddylation, stabilized Nrf2, decreased ROS, reduced retinal inflammation, suppressed Müller gliosis, and mitigated DR. Based on MSC-sEV miRNA sequencing, bioinformatics software, and dual-luciferase reporter assay, miR-143-3p was identified to be the key effector for MSC-sEV\'s role in regulating neural precursor cell expressed developmentally down-regulated 8 (NEDD8)-mediated neddylation. sEV were able to be internalized by Müller cells. Compared to advanced glycation end-products (AGEs)-induced Müller cells, sEV coculture decreased Cullin3 neddylation, activated Nrf2 signal pathway to combat ROS-induced inflammation. The barrier function of endothelial cells was impaired when endothelial cells were treated with the supernatant of AGEs-induced Müller cells, but was restored when treated with supernatant of AGEs-induced Müller cells cocultured with sEV. The protective effect of sEV was, however, compromised when miR-143-3p was inhibited in sEV. Moreover, the protective efficacy of sEV was diminished when NEDD8 was overexpressed in Müller cells. These findings showed MSC-sEV delivered miR-143-3p to inhibit Cullin3 neddylation, stabilizing Nrf2 to counteract ROS-induced inflammation and reducing vascular leakage. Our findings suggest that MSC-sEV may be a potential nanotherapeutic agent for DR, and that Cullin3 neddylation could be a new target for DR therapy.
摘要：
糖尿病视网膜病变（DR）是糖尿病（DM）的高度危险和广泛的并发症。积累的活性氧（ROS）在DR的发展中起着核心作用。这项研究的目的是研究间充质干细胞（MSC）衍生的小细胞外囊泡（sEV）对调节DR中ROS和视网膜损伤的影响和机制。玻璃体内注射sEV抑制Cullin3neddylation，稳定的Nrf2，减少的ROS，减少视网膜炎症，抑制了Müller胶质增生,并减轻了DR。基于MSC-sEVmiRNA测序，生物信息学软件，和双荧光素酶报告分析，miR-143-3p被鉴定为MSC-sEV在调节神经前体细胞发育下调8（NEDD8）介导的Neddylation中的作用的关键效应。sEV能够被Müller细胞内化。与晚期糖基化终产物（AGEs）诱导的Müller细胞相比，sEV共培养减少Cullin3neddylation，激活Nrf2信号通路以对抗ROS诱导的炎症。当用AGEs诱导的Müller细胞的上清液处理内皮细胞时，内皮细胞的屏障功能受损，但用AGEs诱导的Müller细胞与sEV共培养的上清液处理后恢复。sEV的保护作用是，然而,当miR-143-3p在sEV中被抑制时受损。此外，当NEDD8在Müller细胞中过表达时，sEV的保护功效减弱。这些发现显示MSC-sEV递送miR-143-3p以抑制Cullin3neddylation，稳定Nrf2以抵消ROS诱导的炎症并减少血管渗漏。我们的研究结果表明，MSC-sEV可能是DR的潜在纳米治疗剂，Cullin3neddylation可能是DR治疗的新靶点。
