A Cell Culture Approach to Optimized Human Corneal Endothelial Cell Function
| Autor: | Alena Bartakova, Jeffrey L. Goldberg, Noelia J. Kunzevitzky, Karen Alvarez-Delfin, Olga Kuzmenko |
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| Rok vydání: | 2018 |
| Předmět: |
Male
0301 basic medicine Corneal endothelium Cell Survival medicine.medical_treatment Cell Cell Culture Techniques corneal cell therapy Specimen Handling Tight Junctions Flow cytometry Cornea 03 medical and health sciences 0302 clinical medicine In vivo Cadaver endothelial-mesenchymal transition medicine Humans Cell Proliferation Cryopreservation medicine.diagnostic_test Tight junction Chemistry Growth factor Endothelium Corneal Endothelial Cells human corneal endothelial cells CD56 Antigen eye diseases In vitro Culture Media Cell biology 030104 developmental biology medicine.anatomical_structure in vitro production Cell culture 030221 ophthalmology & optometry Female sense organs Biomarkers |
| Zdroj: | Investigative Ophthalmology & Visual Science |
| ISSN: | 1552-5783 |
| Popis: | Purpose Cell-based therapies to replace corneal endothelium depend on culture methods to optimize human corneal endothelial cell (HCEC) function and minimize endothelial-mesenchymal transition (EnMT). Here we explore contribution of low-mitogenic media on stabilization of phenotypes in vitro that mimic those of HCECs in vivo. Methods HCECs were isolated from cadaveric donor corneas and expanded in vitro, comparing continuous presence of exogenous growth factors ("proliferative media") to media without those factors ("stabilizing media"). Identity based on canonical morphology and expression of surface marker CD56, and function based on formation of tight junction barriers measured by trans-endothelial electrical resistance assays (TEER) were assessed. Results Primary HCECs cultured in proliferative media underwent EnMT after three to four passages, becoming increasingly fibroblastic. Stabilizing the cells before each passage by switching them to a media low in mitogenic growth factors and serum preserved canonical morphology and yielded a higher number of cells. HCECs cultured in stabilizing media increased both expression of the identity marker CD56 and also tight junction monolayer integrity compared to cells cultured without stabilization. Conclusions HCECs isolated from donor corneas and expanded in vitro with a low-mitogenic media stabilizing step before each passage demonstrate more canonical structural and functional features and defer EnMT, increasing the number of passages and total canonical cell yield. This approach may facilitate development of HCEC-based cell therapies. |
| Databáze: | OpenAIRE |
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