IN-VITRO PROPAGATION OF PRIMARY AND EXTENDED LIFE-SPAN MURINE CORNEALENDOTHELIAL-CELLS

Purpose. To establish primary and extended life span cell cultures of murine corneal endothelial cells for a model system investigating corn eal endothelial cell replacement and the immunologic features of corne al graft rejection. Methods. The authors have been able to grow cornea l endothelium in culture by isolating adult murine Descemet's membrane and allowing the endothelial cells to proliferate from the explants. To isolate extended life span murine corneal endothelial cells, cells were infected with an adenovirus-SV40 hybrid virus (Ad12-SV40). Result s. The primary cells from adult corneas proliferated to passage 3 befo re growth arrest-senescence was observed. However, the extended life s pan cells remained proliferative through passage 36 an maintained a st ructure similar to the primary cells. Immunohistochemical analysis dem onstrate that the extended life span cells are SV40 large T antigen po sitive, and both the primary and extended life span murine corneal end othelial cells exhibit the common expression of several growth factor receptors and TGF-beta. Conclusions. This is the first report of the i solation and culture of mouse corneal endothelial cells. Additionally, these cells have been infected with a virus carrying th SV40 large T antigen, which yields extended life span mouse corneal endothelial cel ls. These cells will be of interest in establishing a muring model for corneal cell transplantation, and the authors are currently establish ing protocols for the specific introduction and manipulation of these cells in both in vitro and in vivo systems. These types of analyses ma y provide and important animal model specific to in vivo corneal endot helial cell replacement for the treatment of endothelial-related kerat opathies and can be a useful model in delineating the immunologic para meters of corneal graft rejection.