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Long‐term effects of plasma membrane hyperpolarization on cell‐junction stability of corneal endothelial cells in organ and cell culture
Author(s) -
MARTINEZGARCIA C,
EVANS F,
GALLEGO P,
MERAYOLLOVES J,
CHIFFLET S
Publication year - 2010
Publication title -
acta ophthalmologica
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.534
H-Index - 87
eISSN - 1755-3768
pISSN - 1755-375X
DOI - 10.1111/j.1755-3768.2010.206.x
Subject(s) - microbiology and biotechnology , hyperpolarization (physics) , corneal endothelium , actin cytoskeleton , viability assay , endothelial stem cell , propidium iodide , biology , cytoskeleton , cell , chemistry , endothelium , apoptosis , endocrinology , biochemistry , programmed cell death , organic chemistry , nuclear magnetic resonance spectroscopy , in vitro
Purpose One of the most important challenges in corneal preservation is to maintain the epithelial and endothelial integrity. In previous work (Cell Motil. Cytoskeleton 66:1087, 2009) we found that short term (1 hour) hyperpolarization of the plasma membrane potential (HPMP) produces a reorganization of the actin cytoskeleton and increases the stability of cell‐cell junctions of bovine corneal endothelial (BCE) cells in organ and cell cultures. The purpose of this work is to study the effect of long term (several days) HPMP on cell viability, actin organization and intercellular junction stability in cell and organ culture, with the general objective of designing appropriate solutions to use in the maintenance of corneas. Methods Cells and corneas were grown in culture media supplemented by hyperpolarizing agents (valinomycin, choline chloride, lithium chloride). We studied their effects on plasma membrane potential (oxonol V), cell viability (propidium iodide and Tunel), actin organization (FITC‐phalloidin) and cell junction stability (response to calcium deprivation). Results The results showed that cells maintained up to three days in media supplemented with 0.1 µg/mL valinomycin improved their viability in culture and increased the stability of their intercellular junctions, similarly to that obtained in short‐term HPMP treatments. Accordingly, the corneal epithelium and endothelium of treated corneas in culture were better preserved than control ones Conclusion These results suggest that HPMP solutions might be useful in the design of media for preservation of corneas that will be used in transplants.