Serum-free and xenobiotic-free preservation of cultured human limbal epithelial cells

PloS One
Oeygunn UtheimTor P Utheim

Abstract

To develop a one-week storage method, without serum and xenobiotics, that would maintain cell viability, morphology, and phenotype of cultured human limbal epithelial sheets. Human limbal explants were cultured on intact human amniotic membranes for two weeks. The sheets were stored in a hermetically sealed container at 23°C in either a serum-free medium with selected animal serum-derived compounds (Quantum 286) or a xenobiotic-free medium (Minimal Essential Medium) for 4 and 7 days. Stored and non-stored cultures were analyzed for cell viability, amniotic membrane and epithelial sheet thickness, and a panel of immunohistochemical markers for immature cells (ΔNp63α, p63, Bmi-1, C/EBP∂, ABCG2 and K19), differentiated cells (K3 and Cx43), proliferation (PCNA), and apoptosis (Caspase-3). The cell viability of the cultures was 98 ± 1% and remained high after storage. Mean central thickness of non-stored limbal epithelial sheets was 23 ± 3 μm, and no substantial loss of cells was observed after storage. The non-stored epithelial sheets expressed a predominantly immature phenotype with ΔNp63α positivity of more than 3% in 9 of 13 cultures. After storage, the expression of ABCG2 and C/EBP∂ was reduced for the 7 day Quantum 286-storage...Continue Reading

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Citations

Nov 13, 2018·Biomaterials Science·Soyoung HongChangmo Hwang
Mar 25, 2018·Stem Cells Translational Medicine·Tor Paaske UtheimAmer Sehic
Aug 29, 2020·International Journal of Molecular Sciences·Raquel Hernáez-MoyaNoelia Andollo
Sep 7, 2021·Frontiers in Medicine·Ayyad Zartasht KhanJon Roger Eidet

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