Improvement of Storage Medium for Cultured Human Retinal Pigment Epithelial Cells Using Factorial Design

Abstract Storage of human retinal pigment epithelium (hRPE) can contribute to the advancement of cell-based RPE replacement therapies. The present study aimed to improve the quality of stored hRPE cultures by identifying storage medium additives that, alone or in combination, contribute to enhancing...

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Autores principales:	L. Pasovic, T. P. Utheim, S. Reppe, A. Z. Khan, C. J. Jackson, B. Thiede, J. P. Berg, E. B. Messelt, J. R. Eidet
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2018
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/e6f71d30b1a443b3b394d8323bb7c8a6
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spelling	oai:doaj.org-article:e6f71d30b1a443b3b394d8323bb7c8a62021-12-02T15:08:22ZImprovement of Storage Medium for Cultured Human Retinal Pigment Epithelial Cells Using Factorial Design10.1038/s41598-018-24121-82045-2322https://doaj.org/article/e6f71d30b1a443b3b394d8323bb7c8a62018-04-01T00:00:00Zhttps://doi.org/10.1038/s41598-018-24121-8https://doaj.org/toc/2045-2322Abstract Storage of human retinal pigment epithelium (hRPE) can contribute to the advancement of cell-based RPE replacement therapies. The present study aimed to improve the quality of stored hRPE cultures by identifying storage medium additives that, alone or in combination, contribute to enhancing cell viability while preserving morphology and phenotype. hRPE cells were cultured in the presence of the silk protein sericin until pigmentation. Cells were then stored for 10 days in storage medium plus sericin and either one of 46 different additives. Individual effects of each additive on cell viability were assessed using epifluorescence microscopy. Factorial design identified promising additive combinations by extrapolating their individual effects. Supplementing the storage medium with sericin combined with adenosine, L-ascorbic acid and allopurinol resulted in the highest cell viability (98.6 ± 0.5%) after storage for three days, as measured by epifluorescence microscopy. Flow cytometry validated the findings. Proteomics identified 61 upregulated and 65 downregulated proteins in this storage group compared to the unstored control. Transmission electron microscopy demonstrated the presence of melanosomes after storage in the optimized medium. We conclude that the combination of adenosine, L-ascorbic acid, allopurinol and sericin in minimal essential medium preserves RPE pigmentation while maintaining cell viability during storage.L. PasovicT. P. UtheimS. ReppeA. Z. KhanC. J. JacksonB. ThiedeJ. P. BergE. B. MesseltJ. R. EidetNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 8, Iss 1, Pp 1-17 (2018)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q L. Pasovic T. P. Utheim S. Reppe A. Z. Khan C. J. Jackson B. Thiede J. P. Berg E. B. Messelt J. R. Eidet Improvement of Storage Medium for Cultured Human Retinal Pigment Epithelial Cells Using Factorial Design
description	Abstract Storage of human retinal pigment epithelium (hRPE) can contribute to the advancement of cell-based RPE replacement therapies. The present study aimed to improve the quality of stored hRPE cultures by identifying storage medium additives that, alone or in combination, contribute to enhancing cell viability while preserving morphology and phenotype. hRPE cells were cultured in the presence of the silk protein sericin until pigmentation. Cells were then stored for 10 days in storage medium plus sericin and either one of 46 different additives. Individual effects of each additive on cell viability were assessed using epifluorescence microscopy. Factorial design identified promising additive combinations by extrapolating their individual effects. Supplementing the storage medium with sericin combined with adenosine, L-ascorbic acid and allopurinol resulted in the highest cell viability (98.6 ± 0.5%) after storage for three days, as measured by epifluorescence microscopy. Flow cytometry validated the findings. Proteomics identified 61 upregulated and 65 downregulated proteins in this storage group compared to the unstored control. Transmission electron microscopy demonstrated the presence of melanosomes after storage in the optimized medium. We conclude that the combination of adenosine, L-ascorbic acid, allopurinol and sericin in minimal essential medium preserves RPE pigmentation while maintaining cell viability during storage.
format	article
author	L. Pasovic T. P. Utheim S. Reppe A. Z. Khan C. J. Jackson B. Thiede J. P. Berg E. B. Messelt J. R. Eidet
author_facet	L. Pasovic T. P. Utheim S. Reppe A. Z. Khan C. J. Jackson B. Thiede J. P. Berg E. B. Messelt J. R. Eidet
author_sort	L. Pasovic
title	Improvement of Storage Medium for Cultured Human Retinal Pigment Epithelial Cells Using Factorial Design
title_short	Improvement of Storage Medium for Cultured Human Retinal Pigment Epithelial Cells Using Factorial Design
title_full	Improvement of Storage Medium for Cultured Human Retinal Pigment Epithelial Cells Using Factorial Design
title_fullStr	Improvement of Storage Medium for Cultured Human Retinal Pigment Epithelial Cells Using Factorial Design
title_full_unstemmed	Improvement of Storage Medium for Cultured Human Retinal Pigment Epithelial Cells Using Factorial Design
title_sort	improvement of storage medium for cultured human retinal pigment epithelial cells using factorial design
publisher	Nature Portfolio
publishDate	2018
url	https://doaj.org/article/e6f71d30b1a443b3b394d8323bb7c8a6
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Improvement of Storage Medium for Cultured Human Retinal Pigment Epithelial Cells Using Factorial Design

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