Yield and Viability of Human Limbal Stem Cells From Fresh and Stored Tissue

Mason SL, Stewart RMK, Sheridan CM, Keshtkar F, Rooney P, Austin E, Schlötzer-Schrehardt U, Kruse F, Kaye SB (2016)

Publication Type: Journal article

Publication year: 2016

Journal

Investigative Ophthalmology & Visual Science Association for Research in Vision and Ophthalmology (ARVO)

Book Volume: 57

Pages Range: 3708-13

Journal Issue: 8

DOI: 10.1167/iovs.16-19354

Abstract

We compared cell number, putative stem cell markers, and clonogenic ability in fresh uncultured human limbal epithelial cells to that obtained from stored organ-cultured tissue.Cell suspensions were formed from fresh and organ culture-stored human limbal epithelium. Expression of putative stem cell markers ?Np63 and TrkA was performed using immunofluorescent staining before culture. Colony-forming efficiency (CFE) assays were performed at first passage. The effects of tissue storage, age, and postmortem/culture times were analyzed in a general linear model.Limbal tissue from 94 donors (34 fresh and 60 stored) was compared. Three times more cells were obtained per eye from fresh (35.34 × 104; SD, 17.39) than stored (11.24 × 104; SD, 11.57; P < 0.01) tissue. A higher proportion of cells from fresh tissue were viable (91.9%; SD, 5.7 vs. 85%; SD, 10.8) P < 0.01. Higher total cell expression of ?Np63 (20.19 × 104; SD, 15.5 vs. 3.28 104; SD, 4.33) and TrkA (59.24 × 104; SD, 13.21 vs. 7.65 × 104; SD, 1.05) was observed in fresh than stored tissue (P < 0.01). Colony-forming efficiency was higher for fresh (1.42; SD, 0.12) than stored (0.43; SD, 0.15; P < 0.01) cells. For stored tissue only, there was a significant inverse relationship between donor age and total number of cells isolated (R2 = 0.27, P < 0.001).Storage of corneoscleral discs in organ culture medium leads to significant reduction in limbal epithelial cell number, expression of ?Np63 and TrkA, and viability compared to fresh tissue. There is a smaller basal stem cell population in stored compared to fresh tissue.

Authors with CRIS profile

Ursula Schlötzer-Schrehardt Medizinische Fakultät Friedrich Kruse Lehrstuhl für Augenheilkunde

Involved external institutions

The University of Liverpool GB United Kingdom (GB) NHS Blood and Transplant (NHSBT) GB United Kingdom (GB)

How to cite

APA:

Mason, S.L., Stewart, R.M.K., Sheridan, C.M., Keshtkar, F., Rooney, P., Austin, E.,... Kaye, S.B. (2016). Yield and Viability of Human Limbal Stem Cells From Fresh and Stored Tissue. Investigative Ophthalmology & Visual Science, 57(8), 3708-13. https://dx.doi.org/10.1167/iovs.16-19354

MLA:

Mason, Sharon L., et al. "Yield and Viability of Human Limbal Stem Cells From Fresh and Stored Tissue." Investigative Ophthalmology & Visual Science 57.8 (2016): 3708-13.

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