G J Harocopos1, K M Alvares, A E Kolker, D C Beebe. 1. Department of Ophthalmology and Visual Sciences, Cataract Research Center, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Abstract
PURPOSE: To evaluate the importance of lens epithelial cell death in age-related cataract. To determine whether the large percentage of terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labeling (TUNEL)-positive lens epithelial cells previously reported in human capsulotomy specimens results from apoptosis or necrosis. METHODS: Capsulotomy specimens from patients who had undergone cataract surgery and epithelia from cataractous lenses of eye bank eyes were compared with epithelia from noncataractous lenses of eye bank eyes. DNA fragmentation was assayed using the TUNEL method. Cell membrane integrity was tested using a fluorescent stain for DNA, BOBO-3, that is excluded from living cells. Cell proliferation was assayed by labeling with 5-bromo-2'-deoxyuridine (BrdU). The number of cells in different regions of the lens epithelium was measured by digital imaging and computerized counting of nuclei after staining with methyl green. RESULTS: TUNEL-positive cells were sometimes detected adjacent to denuded regions of capsulotomy specimens, especially when epithelia were not fixed immediately after surgery. TUNEL-stained cells usually stained with BOBO-3, indicating loss of plasma membrane integrity. No BrdU-labeled cells were detected in capsulotomy specimens. Cell density in cataractous lens epithelia was similar to that in normal lens epithelia. In cataractous lenses from eye bank eyes, cell density in the region of the epithelium overlying the cataract was higher than cell density in the region of the epithelium overlying the transparent part of the lens. No correlation was found between cell density and cataract severity or between cell density and age. CONCLUSIONS: TUNEL staining of lens epithelial cells in capsulotomy specimens most likely results from necrotic cell death caused by damage during or soon after cataract surgery. Loss of cells from the lens epithelium, by apoptosis or other mechanisms of cell death, does not seem to play a major role in age-related cataract formation.
PURPOSE: To evaluate the importance of lens epithelial cell death in age-related cataract. To determine whether the large percentage of terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labeling (TUNEL)-positive lens epithelial cells previously reported in human capsulotomy specimens results from apoptosis or necrosis. METHODS: Capsulotomy specimens from patients who had undergone cataract surgery and epithelia from cataractous lenses of eye bank eyes were compared with epithelia from noncataractous lenses of eye bank eyes. DNA fragmentation was assayed using the TUNEL method. Cell membrane integrity was tested using a fluorescent stain for DNA, BOBO-3, that is excluded from living cells. Cell proliferation was assayed by labeling with 5-bromo-2'-deoxyuridine (BrdU). The number of cells in different regions of the lens epithelium was measured by digital imaging and computerized counting of nuclei after staining with methyl green. RESULTS: TUNEL-positive cells were sometimes detected adjacent to denuded regions of capsulotomy specimens, especially when epithelia were not fixed immediately after surgery. TUNEL-stained cells usually stained with BOBO-3, indicating loss of plasma membrane integrity. No BrdU-labeled cells were detected in capsulotomy specimens. Cell density in cataractous lens epithelia was similar to that in normal lens epithelia. In cataractous lenses from eye bank eyes, cell density in the region of the epithelium overlying the cataract was higher than cell density in the region of the epithelium overlying the transparent part of the lens. No correlation was found between cell density and cataract severity or between cell density and age. CONCLUSIONS: TUNEL staining of lens epithelial cells in capsulotomy specimens most likely results from necrotic cell death caused by damage during or soon after cataract surgery. Loss of cells from the lens epithelium, by apoptosis or other mechanisms of cell death, does not seem to play a major role in age-related cataract formation.
Authors: Basil V Worgul; Lubomir Smilenov; David J Brenner; Anna Junk; Wei Zhou; Eric J Hall Journal: Proc Natl Acad Sci U S A Date: 2002-07-15 Impact factor: 11.205