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T. Ritter, I. Ecke, J. Yang, M. Hempel, S. Metzner, S. Schu, H.–D. Volk, U. Pleyer; Gene transfer of anti–apoptotic and cytoprotective molecules in human corneal endothelial cells and their influence on prevention of apoptosis . Invest. Ophthalmol. Vis. Sci. 2004;45(13):4794.
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Purpose: Corneal transplantation (keratoplasty) is the most often performed allo–transplantation. The loss of endothelial cells during ex–vivo storage of the cornea before grafting is a major complication. Apoptotic cell death seems to play a role during this process. The aim of our work is to investigate whether gene transfer of anti–apoptotic and cytoprotective genes prevents the induction of apoptosis in human corneal endothelial cells (HCEC). Methods: We used a replication–deficient (E1/E3–deleted) adenovirus (Ad) vector to deliver the anti–apoptotic genes bcl–xL, bag–1 and the cytoprotective gene heme oxygenase–1 (HO–1) to HCEC. The influence of gene transfer on the survival rate of endothelial cells after induction of apoptosis with different stimuli (TNF–alpha (100ng/ml)/Cycloheximid (20µg/ml), Camptothecin (5µM), Staurosporine (0,5µM)) has been examined by two different techniques (detection of Caspase 3, 7 and TUNEL–assay). Results: Adbcl–xL gene transfer is able to protect HCEC from induction of apoptosis in all experiments investigated (p=0.01 vs. non–transduced HCEC). Interestingly AdHO–1 and Adbag–1 gene transfer only protects the cells from Camptothecin induced apoptosis but not from TNF–alpha/Cycloheximid and Staurosporine induced apoptosis. Conclusions: Our results indicate that Bcl–xL is a very effective inhibitor of apoptosis in HCEC and therefore is a promising candidate to improve the survival rate of corneal cells during storage and subsequent transplantation.
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