Purpose: Our previous in vivo confocal microscopic (CM) studies identified quantifiable differences in the corneal epitl elium and stroma for surfactants of different irritancy. This study characterized tbe relationship of area and depth of the initial corneal changes measured by in vivo CM to cell death as measured by ex vivo, Live/Dead Assay (L/D, Molecular Probes) Methods; For 4 groups of rabbits (8 animals each), 10 pi of a surfactant known to produce slight, mild, moderate, or severe irritation was placed on the central cornea of one eye; 2 untreated rabbits served as controls. In vivo CM measurement of groip total mean epithelial thickness and depth of keratocyte loss of 4 corneal regions weie made in 4 rabbits of each group and 2 controls at 3 hr and in the remaining rabbits at 3 hr and 1 day. Corneas were then removed, regions excised and placed in culture media containing 2 pM calcein AM and 4 uM ethidium homodimer. Using las>:r scanning CM, number of dead epithelial or stromal cells in a 300x300x170 (W> (x>z) volume of the cornea was determined. Results: (All p<0.05) By in vivo CM, the slight irritant resulted in epithelial thinning at 3 hr (41.0 ±3.7 pm vs. 45.6 ±2.5 urn fo control, n=4 and 2), while on day 1 mild, moderate and severe irritants caused complete epithelial loss and keratocyte disappearance to a depth of 26.4 ±15.5 u.m, 44.1 ±21.6 urn and 796.0 ±172.3 urn, respectively. At 3 hr, L/D assay detected more dead epithelial cells in slightly irritated eyes (59.8 ±19.5 vs. 14.9 11,6 cells for control); significantly correlating with the in vivo CM measurement of epithelial thinning (r=0.89). At day 1, mild and moderate irritants showed increasing stromal cell death from 9.8 ±16.2 to 36.4 ±31.2 cells which significantly correlated with in vivo CM keratocyte loss (r=0.78). No surviving keratocytes were detected in severely irritated eyes. Conclusions The data supports our hypothesis that differences in surfactant induced ocular irritation are directly related to area and depth of acute corneal injury.
|Original language||English (US)|
|Journal||Investigative Ophthalmology and Visual Science|
|State||Published - Dec 1 1997|
ASJC Scopus subject areas
- Sensory Systems
- Cellular and Molecular Neuroscience