QUANTITATIVE ASSESSMENT OF ANTEROPOSTERIOR KERATOCYTE DENSITY IN THE NORMAL RABBIT CORNEA

The anteroposterior keratocyte density distribution in the rabbit corn ea was measured. Unsectioned tissue blocks from the central cornea of five rabbits were stained with propidium iodide and imaged using a Lei ca laser scanning confocal microscope. A z-series of images was acquir ed in each sample, from anterior to posterior stroma in either 3- or 8 -mu m steps. Software was developed to allow interactive marking of th e keratocyte nuclei within each section of the z-series and for calcul ating cell density. For convenience, cell density was expressed as the number of cells per corneal volume element (CVE), where CVE is a newl y defined volume unit with x, y, and z dimensions of 250, 250, and 10 mu m, respectively. The calculated keratocyte density was 20.2 +/- 1.0 cells/CVE (n = 5), which is equivalent to 32,360 +/- 1,660 cells/mm(3 ). The greatest density was underneath the epithelium (26.3 +/- 2.5 ce lls/CVE), the density then decreased linearly with depth to 15.2 +/- 1 .4 cells/CVE; there was a slight increase in density pre-Descemets mem brane to 18.5 +/- 3.5 cells/CVE. A 30% decrease in cell density over t he entire anteroposterior stromal thickness was observed. To facilitat e statistical analysis, the cell density was averaged over 5% thicknes s intervals from anterior to posterior cornea. A significant differenc e in mean cell density of these intervals was found (ANOVA, n = 20, p < 0.01). To further assess the density distribution, linear regression analysis was performed. A significant correlation was found between k eratocyte density and stromal depth (R = -0.94, n = 20, p < 0.05). We conclude that in the rabbit, keratocyte cell density is maximal underl ying the epithelium and progressively decreases from anterior to poste rior cornea.