Effect of different culture media and deswelling agents on survival of human corneal endothelial and epithelial cells in vitro
Research output: Contribution to journal › Research article › Contributed › peer-review
Contributors
Abstract
To examine the effects of media and deswelling agents on human corneal endothelial and epithelial cell viability using a previously developed screening system.
The human corneal endothelial cell line HCEC-12 and the human corneal epithelial cell line HCE-T were cultured in four different corneal organ culture media (serum-supplemented: MEM +2 % FCS, CorneaMaxA (R)/CorneaJetA (R), serum-free: Human Endothelial-SFM, Stemalpha-2 and -3) with and without 6 % dextran T500 or 7 % HES 130/0.4. Standard growth media F99(HCEC) and DMEM/F12(HCE-T) served as controls. In additional controls, the stress inducers staurosporine or hydrogen peroxide were added. After 5 days in the test media, cell viability was assessed by flow cytometrically quantifying apoptotic and necrotic cells (sub-G1 DNA content, vital staining with YO-PRO-1A (R) and propidium iodide) and intracellular reactive oxygen species (ROS).
The MEM-based media were unable to support HCEC-12 and HCE-T survival under stress conditions, resulting in significantly increased numbers of apoptotic and necrotic cells. HCEC-12 survival was markedly improved in SFM-based media even under staurosporine or hydrogen peroxide. Likewise, HCE-T survival was improved in SFM with or without dextran. The media CorneaMaxA (R), CorneaJetA (R), and CorneaMaxA (R) with HES supported HCEC-12 survival better than MEM-based media, but less well than SFM-based media. HCE-T viability was also supported by CorneaJetA (R), but not by CorneaMaxA (R) with or without HES. Stemalpha-based media were not suitable for maintaining viability of HCEC-12 or HCE-T in the applied cell culture system.
The use of serum-supplemented MEM-based media for corneal organ culture should be discontinued in favour of serum-free media like SFM.
Details
Original language | English |
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Pages (from-to) | 285-295 |
Number of pages | 11 |
Journal | Graefe's Archive for Clinical and Experimental Ophthalmology |
Volume | 254 |
Issue number | 2 |
Publication status | Published - Feb 2016 |
Peer-reviewed | Yes |
External IDs
WOS | 000370004300010 |
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Scopus | 84955651863 |
ORCID | /0000-0003-3205-1876/work/142256437 |
Keywords
Keywords
- Corneal organ cultivation, Serum-free medium, Deswelling substance, Cell survival, Human corneal endothelium, Human corneal epithelium, SERUM-FREE MEDIUM, ORGAN-CULTURE, DONOR CORNEAS, STORAGE, DEXTRAN, APOPTOSIS, NECROSIS, DENSITY