Annexin V binding to rabbit corneal epithelial cells following overnight contact lens wear or eyelid closure.
OBJECTIVE
To determine the effects of open or closed eye and overnight contact lens wear on rabbit corneal epithelial surface cell death, detected by annexin V binding to cell surface phosphatidylserine and propidium iodide (PI) double-labeling.


METHODS
New Zealand white rabbits (n = 42) weighing 2.5 to 3.5 kg were divided into 7 study groups: hyper Dk/t rigid gas-permeable (RGP) lens; high Dk/t RGP lens; low Dk/t RGP lens; hyper Dk/t soft lens; high Dk/t soft lens; eyelid closure; and nictitating membranectomy study group (n = 6 rabbits for each group). Each rabbit was randomly chosen to have either one eye fitted with a test contact lens or both eyelids sutured closed; the contralateral eye served as a control. Rabbits were humanely sacrificed after 24 hours. Corneal buttons including the limbus were excised and stained with annexin V-FITC and PI to identify the number of nonviable epithelial surface cells. A series of sequential microscopic adjacent fields (200 microm X560 microm) from the inferior limbus to the central cornea were evaluated using epifluorescence microscopy, and the total number of cells stained with annexin V or PI, or both, was counted in each field.


RESULTS
Nonviable cells (cells with positive staining) were lowest in density at the limbus and gradually increased in numbers towards the central cornea under normal open eye conditions (P<0.05). Eyelid closure (no lens) caused a significant decrease in the numbers of nonviable cells in the peripheral and central cornea (P<0.05) but not at the limbus (P>0.05). Overnight wear of hyper Dk/t or high Dk/t RGP soft contact test lenses caused significant decreases in the numbers of nonviable cells on the central corneal surface (P<0.05). Hyper Dk/t and high Dk/t soft lenses had similar suppressive effects on the number of surface nonviable cells, independent of lens oxygen transmissibility. By contrast, the number of nonviable cells was dependent upon RGP lens oxygen transmissibility; hyper Dk/t RGP lens wear produced significantly less suppression of the number of nonviable cells in the central cornea than high Dk/t RGP lens wear (P<0.05). Wear of a nonphysiologic low oxygen transmissible RGP test lens however, produced a significant increase in nonviable cells in the central cornea. The number of nonviable epithelial surface cells was not affected by nictitating membranectomy under open eye conditions (P>0.05).


CONCLUSIONS
This study revealed in the rabbit model a gradient of nonviable surface epithelial cells increasing towards the central cornea under open eye conditions and with central suppression of surface cell death following closed eye (no lens) or physiologic contact lens wear. Taken together, the results suggest that eyelid closure or contact lens wear may protect the corneal epithelial cells against the shear stress forces exerted by eyelid blinking, which are believed to drive central epithelial surface cell death and exfoliation. However, under very low oxygen tensions combined with lens effect, such as in low Dk/t RGP lens wear, surface cell death may be accelerated.
Li
Ling
L
Department of Ophthalmology, The University of Texas Southwestern Medical Center at Dallas, 75390-9057, USA.
Ren
David H
DH
Ladage
Patrick M
PM
Yamamoto
Kazuaki
K
Petroll
W Matthew
WM
Jester
James V
JV
Cavanagh
H Dwight
HD
D000818:Animals; D017304:Annexin A5 / Q000378:metabolism*; D017209:Apoptosis; D001767:Blinking; D002452:Cell Count; D003262:Contact Lenses, Extended-Wear; D019573:Epithelium, Corneal / Q000166:cytology / Q000378:metabolism*; D005143:Eyelids / Q000601:surgery*; D016650:Fluorescein-5-isothiocyanate; D010718:Phosphatidylserines / Q000378:metabolism*; D011419:Propidium; D011817:Rabbits; D012683:Sensory Deprivation
D017304:Annexin A5; D010718:Phosphatidylserines; D011419:Propidium; D016650:Fluorescein-5-isothiocyanate