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A. H. Kakazu, J. He, H. E. P. Bazan; Aspirin-Triggered Lipoxin A4 (epi-LXA4) Increases the Preservation Time of Human Corneal Endothelium in Optisol-GS. Invest. Ophthalmol. Vis. Sci. 2010;51(13):2824.
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© ARVO (1962-2015); The Authors (2016-present)
Epi-LXA4 is an anti-inflammatory bioactive lipid formed when aspirin acetylate cyclooxygenase-2 (COX-2). In previous studies we have shown that low concentrations of epi-LXA4 stimulate wound closure in rabbit corneal endothelial cells (ARVO 2008 and 2009). Optisol-GS (Bausch & Lomb) is considered an excellent medium to store corneas for transplant; however, clinical studies indicate that the time of storage should not exceed 7 days. The purpose of the current study is to investigate if epi-LXA4 increases the integrity of the human corneal endothelium after prolonged storage in Optisol-GS.
Human and porcine corneal endothelial cells (HCEC and PCEC) were cultured in DMEM/F12 supplemented with 8-15% FBS. LXA4 receptor was detected by immunoestaining and Western blot. To assess cell migration, confluent HCEC and PCEC were wounded by a linear scraping in the center of the well and incubated in the presence of 5 mM hydroxyurea for 24 h with or without 100 nM epi-LXA4. Cell proliferation was evaluated with Ki-67 antibody. Pairs of human corneas (ages ranging from 35 to 88 years old) were purchased from NDRI (Philadelphia, PA). Corneas were organ-cultured in DMEN/F12 with or without 100 nM epi-LXA4 for 24 h at 37ºC, transferred to Optisol- GS, and stored at 4ºC for 12 days. Corneal endothelial integrity was evaluated by combined staining with the vital stain trypan blue and the intercellular stain alizarin red S. Endothelial cell damage was defined by the percentage of cells stained with trypan blue, areas devoid of cells, and areas of cellular disruption.
HCEC and PCEC expressed the LXA4 receptor. There was a 3-fold increase in cell proliferation when HCEC were incubated with epi-LXA4 for 24 h. For PCEC, the increase in cell proliferation was 60%. Cell migration was not affected. When the human corneas were incubated in DMEN/F12 in the presence of epi-LXA4 and then stored in Optisol-GS for 12 days, there was a 25 to 50% decrease in endothelial damage compared to storage in Optisol-GS alone.
Epi-LXA4 increases the length of time that donor tissue can be stored in Optisol-GS without loss of endothelial integrity. The bioactive lipid seems to stimulate the proliferation when the corneas are pre-incubated for 24 h before storage.
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