Nuzzi Raffaele, Buono Lola, Scalabrin Simona, De Iuliis Marco, Bussolati Benedetta
Eye Clinic, Department of Surgical Sciences, University of Turin, AOU Città della Salute e della Scienza, Turin, Italy.
Department of Biotechnology and Health Sciences, University of Turin, Turin, Italy.
Stem Cells Int. 2021 Jan 16;2021:6644463. doi: 10.1155/2021/6644463. eCollection 2021.
Human corneal endothelial cells (HCECs) are essential to visual function; however, since they have limited proliferative capacity , they are prone to corneal endothelial dysfunction. At present, the only treatment is a corneal transplantation from donor cadavers. Also, due to a global shortage of donor corneas, it is important to find alternative strategies. Recent studies highlight that stem cell-derived extracellular vesicles (EVs) play a relevant role in stem cell-induced regeneration by reprogramming injured cells and inducing proregenerative pathways. The aim of this work is to evaluate whether EVs derived from mesenchymal stem cells (MSC-EVs) are able to promote regeneration of damaged HCECs.
We isolated HCECs from discarded corneas in patients undergoing corneal transplantation or enucleation ( = 23 patients). Bone marrow mesenchymal stem cells (MSCs) were obtained from Lonza, cultured, and characterized. MSC-EVs were obtained from supernatants of MSCs. In order to establish a valid damage model to test the regenerative potential of EVs on HCECs, we evaluated the proliferation rate and the apoptosis after exposing the cells to serum-deprived medium at different concentrations for 24 hours. We then evaluated the HCEC migration through a wound healing assay.
In the selected serum deprivation damage conditions, the treatment with different doses of MSC-EVs resulted in a significantly higher proliferation rate of HCECs at all the tested concentrations of EVs (5-20 × 10 MSC-EV/cell). MSC-EVs/cell induced a significant decrease in number of total apoptotic cells after 24 hours of serum deprivation. Finally, the wound healing assay showed a significantly faster repair of the wound after HCEC treatment with MSC-EVs.
Results highlight the already well-known proregenerative potential of MSC-EVs in a totally new biological model, the endothelium of the cornea. MSC-EVs, indeed, induced proliferation and survival of HCECs, promoting the migration of HCECs .
人角膜内皮细胞（HCECs）对视觉功能至关重要；然而，由于它们的增殖能力有限，容易发生角膜内皮功能障碍。目前，唯一的治疗方法是使用供体尸体的角膜移植。此外，由于全球供体角膜短缺，寻找替代策略很重要。最近的研究表明，干细胞衍生的细胞外囊泡（EVs）通过重编程受损细胞和诱导促再生途径，在干细胞诱导的再生中发挥相关作用。这项工作的目的是评估间充质干细胞衍生的细胞外囊泡（MSC-EVs）是否能够促进受损HCECs的再生。
我们从接受角膜移植或眼球摘除术的患者（n = 23例）废弃的角膜中分离出HCECs。从Lonza公司获得骨髓间充质干细胞（MSCs），进行培养和鉴定。从MSCs的上清液中获得MSC-EVs。为了建立一个有效的损伤模型来测试EVs对HCECs的再生潜力，我们评估了细胞在不同浓度的血清饥饿培养基中暴露24小时后的增殖率和凋亡情况。然后，我们通过伤口愈合试验评估HCEC的迁移。
在选定的血清饥饿损伤条件下，用不同剂量的MSC-EVs处理后，在所有测试的EVs浓度（5 - 20×10个MSC-EV/细胞）下，HCECs的增殖率均显著更高。在血清饥饿24小时后，MSC-EVs/细胞诱导总凋亡细胞数量显著减少。最后，伤口愈合试验表明，用MSC-EVs处理HCECs后，伤口修复明显加快。
结果突出了MSC-EVs在一个全新的生物学模型——角膜内皮中早已为人所知的促再生潜力。事实上，MSC-EVs诱导了HCECs的增殖和存活，促进了HCECs的迁移。
