Desarrollo de un modelo de experimentación tridimensional de cicatrización cornealfacilitando el estudio de nuevos fármacos

Abstract

In this doctoral thesis, an ex vivo porcine corneal wound healing model is presented. Currently, it is necessary to develop this type of models to reduce the number of laboratory animals used in research and particularly in visual science research. First, a study of the porcine OS was carried out from the histopathological point of view to confirm that the findings derived from the thesis could be extrapolated to humans. To do so, we used ocular exenterations with their eyelids from animals that had been sacrificed for human consumption. Next, a protocol to damage the porcine corneal epithelium and to culture the sclerocorneal rings for enough time to produce a full corneal epithelial healing was defined. Epithelial wounds were exposed to different molecules to evaluate the performance of the model from a functional point of view measuring changes in the healing rate. One of the molecules evaluated was melatonin, which had previously been shown to promote corneal wound healing in a rabbit corneal damage model. Prior to evaluate the impact of melatonin exposure, we performed a complete study of the distribution of its main receptors (MT1 and MT2) and the precursor molecule AANAT in porcine OS. The ex vivo model of corneal healing was then exposed to melatonin, both in solution and encapsulated in novel lecithin and lecithin-chitosan based NP formulations. Our results confirmed 1) the similarity between porcine and human OS, 2) the presence and regional distribution of melatonin receptors and AANAT in different tissues of porcine OS, 4) the usefulness of the model to test active molecules, and 3) the successful melatonin encapsulation in the NP formulation, although we could not find differences in the percentage of epithelial healing between encapsulated melatonin and its controls.