RT info:eu-repo/semantics/article
T1 Consecutive Expansion of Limbal Epithelial Stem Cells from a Single Limbal Biopsy
A1 López-Paniagua, Marina
A1 Nieto-Miguel, Teresa
A1 De La Mata Sampedro, Ana
A1 Galindo, Sara
A1 Herreras, José M.
A1 Corrales, Rosa M.
A1 Calonge, Margarita
K1 Limbal stem cells; Corneal epithelium; Limbal stem cell deficiency; Limbal explants; Limbal transplantation.
AB Purpose: Corneal epithelium is maintained by limbal epithelial stem cells (LESCs), the loss of which can be catastrophic for corneal transparency. Effective therapies include the transplantation of cultivated LESCs, requiring optimization of in vitro cultivation protocols. Unfortunately, optimization studies are hampered by the limited number of ocular tissue donors. We investigated the feasibility of obtaining more than one limbal primary culture (LPC) from the same 1-2 mm2 limbal explant (LE). Methods: LEs were plated and maintained until outgrowth surrounded each, being removed at this point. LPCs were allowed to reach confluence (LPC0). The same removed LE was plated again, following the same procedure, obtaining LPC1. This procedure was repeated as often as possible up to 6 times. LPCs from each passage were analysed by real time RT-PCR and immunofluorescence-microscopy. Results: LPCs from LPC0 to LPC2 presented an heterogeneous cell population, with cells positive for LESC markers K14, K15, ABCG2 and p63, differentiated corneal epithelial cell-specific markers K3 and K12, and for the fibroblast marker S100A4. These cells had an epithelial-like morphology. In LPC3-LPC4, elongated cell morphology appeared, and the presence of LESC markers decreased, while the presence of differentiated corneal epithelial-cell and fibroblast markers increased.Conclusion: one LE can be successfully cultivated up to three consecutive times while maintaining the LESC phenotype in the LPC cells. This protocol provides several homologous LPCs for basic research. Additionally, by using a cell-carrier, the resulting LPCs could serve reservoirs for potential autologous expanded LESC transplantations and/or for making correlations between laboratory and clinical outcomes.
PB Taylor & Francis Group
SN 0271-3683
YR 2013
FD 2013
LK https://uvadoc.uva.es/handle/10324/62956
UL https://uvadoc.uva.es/handle/10324/62956
LA eng
NO Current Eye Research. 2013 May;38(5):537-549.
NO Producción Científica
DS UVaDOC
RD 11-jul-2024