005 Analyses histologiques et ultra structurales des cornées humaines conservées en Corneamax et déturgées durant 48 heures dans les poloxamers

Aim Poloxamers (P) are various synthetic block copolymers widely used in pharmaceutic industry. We previously described the efficiency of P188, 237, 338 and 407 to deswell human corneas at the end of organ culture (OC) and their ability to reduce the endothelial cell loss compared to 5% T500 Dextran in CorneaMax (with 2% foetal calf serum). We aimed to further assess the cellular effects of 48 hours incubation in P using histology and electron microscopy (EM). Materials and Methods The P were dissolved in CMax (Eurobio, Les Ulis, France) at 350 mOsmol/L. Five paired human corneas with more than 2000 endothelial cells/mm 2 used for each poloxamer. After 3 weeks of OC in CMax, one cornea was incubated for 48 hours in CMax + poloxamer, and the mate cornea in CorneaJet (CMax+ 5% T500 Dextran). Controls: fresh corneal button from keratoconus and lattice dystrophy (for normal unstored endothelium) and organ cultured corneas without deswelling. Corneas were divided into 3 parts for cross sectional histology (epithelial thickness measurement and counting of keratocytes), transmission EM of the posterior stroma and endothelium (cell morphology and interfibrillar spaces measurement) and scanning EM of the endothelium (morphology). Results Cross sectional histology showed differences in epithelial thickness. For both types of macromolecules (poloxamers and dextran) TEM revealed cytoplasmic vacuoles in endothelial cells and keratocytes and SEM confirmed the integrity of endothelial monolayer. Discussion Histological and ultrastructural findings, along with capacity of thinning, transparency improvement and endothelial cell preservation will allow selecting poloxamers to be added to CorneaMax for a future clinical application. Conclusion This complementary validation using light and electron microscopy reasonably confirms the promising use of Poloxamer to replace T500 Dextran to deswell corneas during organ culture.