Decellularized pig corneal stroma seeded with limbal cells

Abstract

Event Abstract Back to Event Decellularized pig corneal stroma seeded with limbal cells Wendolyne Lara Daz1, Beatriz Hernandez Tellez1, Judith Alvarez-Perez1, Katia Jarquín-Yanez1, Gabriela Piñón-Zarate1, Sandra Acevedo Nava1, Miguel A. Herrera-Enríquez1 and Andrés Castell-Rodriguez1 1 UNIVERSIDAD NACIONAL AUTÓNOMA DE MÉXICO, Cell and tissue biology, Medicine School, Mexico Introduction: The cornea is a highly specialized avascular organ in the anterior surface of the eye. It provides two thirds of the refractive capacity thereof while shielding structures acting as a barrier to infectious agents and mechanical damage. The cornea consists of five layers: anterior epithelium, Bowman's membrane, stroma, Descemet's membrane and posterior epithelium (or corneal endothelium). In the transition zone between the cornea and sclera (limbus) resides a subpopulation of corneal stem cells has been observed, exhibit a high proliferative potential and have the ability to differentiate into corneal epithelial cells, called limbal epithelial cells. Various diseases can damage the cornea leading to corneal opacity and once this happens transplantation is the predominant choice to correct visual impairment. Tissue engineering allows the creation of tissue equivalents with cultured cells in an extracellular matrix and growth factors. Corneal cells have been cultivated in different types of three-dimensional matrices as fibrin, collagen and synthetic polymers, however they present several disadvantages as rapid degradation of the scaffold and inflammation. While devitalized stroma not only possess adequate formed, but show binding sites integrins located in the limbal epithelial cells, which are an alternative to the development of a corneal equivalent it may be used as a potential source of xenografts viable cornea for transplantation. Objective: The aime of this study was to induce the growth of limbal epithelial cells in a porcine decellularized corneal stroma in vitro. Materials and Methods: Anterior and posterior corneal limbal epithelial cells were isolated and expanded, corneal stromas were obtained and devitalized using Triton X100 solution. The latter was histologically analyzed to check absence of cells and α-gal epitope. Epithelial cells were seeded and cultured in the anterior portion and the posterior endothelial cells in decellularized portion stroma. Further histological slices were obtained and processed to detect the presence of cytokeratin immunohistochemistry. Results: Histological analysis of devitalized the absence of stroma cells were found within the stroma and α-gal. Limbal epithelial cells proliferated on the anterior surface of the stroma decellularized and formed a confluent layer of cells two or three layers thick, whereas endothelial cells and later joined epithelium expanded on the stromal surface to confluence and form a monolayer. Immunohistochemical analysis revealed the presence of cytokeratins in the epithelium. Conclusions: limbal epithelial cells seeded on the decellularized porcine stroma were able to proliferate and differentiate to give rise to the generation of an earlier and a later epithelium. It is important to conduct more studies in this type of constructs for the future may be used in people with corneal opacities. Acknowledgements: DGAPA PAPIIT IN218315. DGAPA- PAPIIT IN218315