Gurdal Mehmet, Korkmaz Ilayda, Kocamanoglu Meltem, Arici Mesut, Yildirim Nuri, Barut Selver Ozlem
Limbustem R&D Medical Products Ltd., Ege University Technopark, 35100, Izmir, Türkiye.
Ocular Surface Research Laboratory, Faculty of Medicine, Ege University, 35100, Izmir, Türkiye.
Cell Tissue Bank. 2025 Sep 4;26(3):39. doi: 10.1007/s10561-025-10190-3.
Although many preclinical and clinical studies are ongoing on amniotic membrane extract (AME), an amniotic membrane-derived product developed to support ocular surface healing, the effect of AME on the basic cellular functions and properties of human corneal epithelial cells (hCECs) has not been clearly defined. In this study, we aimed to evaluate the effect of AME supplementation to the culture media, on basic cellular functions of hCECs and on expression of specific cell markers of hCECs, as well as to determine its effectiveness in an experimental in vitro wound model. hCECs were seeded with the constant cell density in 6, 24 and 48 well plates. The next day, the media was refreshed with 0 mg/ml, 0.75 mg/ml, 1.5 mg/ml and 3 mg/ml final concentration of AME supplemented complete growth medium. Cellular morphology, viability, metabolic activity, proliferation assessments and immunocytochemistry were conducted on the time points at day 3, 6 and 9. Then, in vitro wound healing assay was performed on hCECs under the effect of AME. AME did not affect cellular morphology, viability, metabolic activity and proliferation. AME supplementation induced CK3 and CK12 expression of hCECs significantly (p < 0.05) higher than without AME group. In vitro wound healing assay revealed that while control and AME-treated cultures both exhibited healing, AME-treated mechanical abrasions closed at a greater rate compared to control. Our study shows that AME promotes in vitro wound healing and cell characteristics in terms of CK3 and CK12 protein expressions, while preserving basic cellular functions of corneal epithelial cells.
尽管针对羊膜提取物（AME）正在进行许多临床前和临床研究，AME是一种为支持眼表愈合而研发的羊膜衍生产品，但AME对人角膜上皮细胞（hCECs）基本细胞功能和特性的影响尚未明确界定。在本研究中，我们旨在评估向培养基中添加AME对hCECs基本细胞功能以及hCECs特异性细胞标志物表达的影响，并确定其在体外实验性伤口模型中的有效性。将hCECs以恒定细胞密度接种于6孔、24孔和48孔板中。次日，用终浓度分别为0 mg/ml、0.75 mg/ml、1.5 mg/ml和3 mg/ml的添加AME的完全生长培养基更换培养基。在第3天、第6天和第9天的时间点进行细胞形态、活力、代谢活性、增殖评估和免疫细胞化学检测。然后，在AME作用下对hCECs进行体外伤口愈合试验。AME不影响细胞形态、活力、代谢活性和增殖。添加AME显著诱导hCECs的CK3和CK12表达（p < 0.05），高于未添加AME的组。体外伤口愈合试验显示，虽然对照组和AME处理组的培养物均表现出愈合，但与对照组相比，AME处理的机械擦伤愈合速度更快。我们的研究表明，AME在促进体外伤口愈合以及在CK3和CK12蛋白表达方面的细胞特性，同时保留角膜上皮细胞的基本细胞功能。
