Endothelial morphometry by image analysis of corneas organ cultured at 31 degrees C

To determine the factors influencing endothelial morphometry by using image analysis of corneas stored in organ culture to determine the coefficient of variation (CV) in cell area and percentage of hexagonal cells.

The endothelia of 505 of the 559 corneas consecutively stored at the eye bank were routinely analyzed with Sambacornea image-analysis software (ver. 1.2.10; Tribvn, Châtillon, France) on three large-field images of 750 x 1000 microm, obtained after osmotic dilation of the intercellular spaces with 0.9% sodium chloride. Analysis was performed on at least 300 cells. The quality of the three-image set was graded poor, average, or good by an independent observer. The studied parameters were donor age and sex, lens status, storage time, and intrinsic quality of captured images. Statistics were analyzed by nonparametric tests.

Image analysis was possible for 504 of the 505 assessed corneas. Donor age correlated significantly with endothelial cell density (ECD; r = -0.343), CV (r = 0.221), and hexagonality (r = -0.314; P < 0.001 for the three). Image quality significantly influenced these three parameters. ECD and hexagonality decreased parallel to image quality, whereas the CV increased. In the 258 corneas assessed twice (on average, at day [D] +4, then D +14) ECD, CV, and hexagonality decreased during storage.

Despite the sometimes mediocre quality of the transmitted light microscopy images, endothelial parameters supplied by the analyzer were clinically reliable, since variations similar to those long known in specular microscopy were found. Endothelial morphometry (CV and hexagonality) is likely to provide further information on the endothelial function of the graft tissue, perhaps particularly for grafts of borderline ECD, close to the discard threshold.