Effects of Amniotic Membrane-Derived Fibroblast Supernatant on Corneal Epithelium

• Published in: Investigative ophthalmology & visual science 2019-09, Vol.60 (12), p.3718-3726
• Main Authors: Higa, Kazunari, Higuchi, Junko, Kimoto, Reona, Satake, Yoshiyuki, Yamaguchi, Takefumi, Tomida, Daisuke, Shimazaki, Jun
• Format: Article
• Language: English
• Subjects: Amnion - cytology; Animals; Biological Factors - pharmacology; Biomarkers - metabolism; Cell Differentiation - drug effects; Cell Division - drug effects; Cells, Cultured; Coculture Techniques; Corneal Injuries - drug therapy; Corneal Injuries - metabolism; Corneal Injuries - pathology; Epithelium, Corneal - drug effects; Epithelium, Corneal - metabolism; Epithelium, Corneal - pathology; Female; Fibroblasts - physiology; Flow Cytometry; Fluorescent Antibody Technique, Indirect; Humans; Limbus Corneae - cytology; Mice; NIH 3T3 Cells; Phenotype; Rabbits; Reverse Transcriptase Polymerase Chain Reaction; Wound Healing - physiology
• ISSN: 1552-5783; 1552-5783
• DOI: 10.1167/iovs.19-27041

To evaluate the effects of human amniotic membrane-derived fibroblast (AMF) cell supernatant (AMF-sup) on corneal epithelium. The phenotype of AMF cells was analyzed by flow cytometry using cell-surface markers. AMF cells were also induced to form osteoblasts and neural cells, and cell phenotypes were observed by staining and RT-PCR. Cultivated human corneal limbal epithelial sheets generated using AMF-sup were analyzed using immunohistochemistry and colony-forming efficiency, and the wound healing of epithelial defects was observed using a tissue-punch method. The effects of instillation of each supernatant in a rabbit corneal epithelial wound healing model were compared. Mesenchymal stem cell (CD29, CD44, CD73, and CD90) and neural crest (CD49d and CD56) markers were expressed on the AMF cell surface. Following induction of differentiation, isolated AMF cells showed characteristics of osteoblasts and neural cells. Application of AMF-sup resulted in maintenance of the limbal epithelial phenotype and immature state, and significantly promoted wound healing in cultivated human corneal limbal epithelial sheets (P < 0.05) and rabbit corneal epithelium (P < 0.05) compared with the control. These data suggest that AMF cells have multi-differentiation potential, and that AMF-sup is effective in maintaining the limbal epithelial phenotype and promoting corneal epithelial wound healing, which may be of value in ocular surface reconstruction.