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J. Oh, M. Kim, J. Ko, H. Lee, J. Lee, W. Wee; Development of Porcine Cornea for Human Corneal Substitute. Invest. Ophthalmol. Vis. Sci. 2009;50(13):603.
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© ARVO (1962-2015); The Authors (2016-present)
In order to investigate the propriety of porcine cornea as a source for lamellar corneal xenografts and to develop the novel processing method to reduce the immunogenicity of porcine corneal stroma.
To investigate the immune-related susceptibility of porcine keratocytes in a xeno-related rejection in humans, we evaluated the deposition of complement, inhibition of cell proliferation, and cytotoxicity of porcine keratocytes after mixing cultured porcine keratocytes with human sera or PBMCs. To develop the novel methods to reduce the immunogenicity from porcine corneal stroma, we treated porcine corneas with (i) freezing, (ii) three freezing-thawing, (iii) hypertonic saline, (iv) hyperosmolar glycerol, (v) trypsin/sodium dodecyl sulfate/Dispase, (vi) DNase/RNase, (vii) three freezing-thawing with hypertonic saline. After processing, we examined the cells and collagen structures of the decellularized corneas using hematoxylin-eosin staining, TUNEL assay, and transmission electron microscopy, live/dead cell staining and organ culture. Moreover, RNA was isolated from cultured porcine keratocytes with or without processing, and gene expression was comparatively analyzed with microarray method. To investigate the efficacy of processed porcine corneas in xenotransplantation, we performed the anterior lamellar transplantation using processed porcine corneas in rabbits and nonhuman primates. Graft survival and integration were assessed. The grafted corneas were histologically examined sequentially after transplantation.
A significant amount of complement was deposited and marked inhibition of proliferation and cytotoxicity were observed in porcine keratocytes when mixed with human sera or PBMCs. Hypertonic saline-based treatment preserved the collagen structure of the porcine cornea and removed cells without producing much apoptotic debris. The treated porcine corneal stroma remained free of inflammation after anterior lamellar xenotransplantation, even in the absence of immunosuppression. These treatments also allowed for integration of the transplant into the adjacent host tissue by providing a conductive template for cell ingrowth.
Porcine corneal lamellar grafts after processing have the potential for use in humans as non-immunogenic donor sources for corneal transplantation, particularly in subjects with functional corneal endothelium or in subjects with corneal perforation who require a tectonic patch.
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