Orthotopic corneal transplantation in the mouse - A new surgical technique with minimal endothelial cell loss

Background: The murine model of orthotopic performing keratoplasty is important for studying the allograft reaction, but the small dimensions cause technical difficulties. Methods: The anterior chamber of the eve of the BALB/c mouse was measured with the confocal microscope and with histological methods. Ten C3H mouse donor corneas each were separated by the conventional technique and by the newly developed ''underwater'' technique, where the opened donor eye did not lose its shape under water. The corneal endothelium was stained with trypan blue and alizarin red S. Ten BALB/c (H-2d) mice received a corneal graft taken from a C3H (H-2k) mouse by the underwater technique. Results: The 3.7-mm eye of the BALB/c mouse has a corneal diameter of 3.5 mm. The cornea has a central thickness of 170 mu m, the epithelium comprising 30% and the stroma 70%. While none of the corneas separated by the new ''underwater'' technique evidenced endothelial damage, a 28 +/- 17.0% defect of the endothelial surface was found with the conventional technique. All transplanted corneas were clear when the lids were opened on the 2 postoperative day and clouded between the 7th and 30th days (mean 16.5 days) due to an allograft reaction. Conclusion: The newly developed ''underwater'' technique is superior to the conventional technique, since floating of the very thin donor cornea during the separation procedure prevents endothelial defects by guarding against folds. By enabling reliable keratoplasty in the mouse, this technique facilitates studies on the experimental allograft reaction.