Purpose. We recently showed that, at'te corneal epithelial cells divide, both daughter cells either remain in the basal layer, or, after a variable time period, both cells leave the basal layer together and differentiate. An experiment was designed to lest whether the rate at which daughter cell pairs different ated was affected by continued division in neighboring basal cells. Methods. Adult male Sprague Dawley rats were injected i.p. with 5-bromo-deoxyuridine (BrdU; 200 nig/kg). Twelve hours later. 5/d of 5flurouracil (5-FU, 50 mg/ml) was placed on one eye. 5-FU application was repeated every 12 hours for a total of 72 hours. Animals were sacrificed, eyes were fixed and corneas were dissected, penneabilized and stained with anti-BrdU antibody and a rhodamine-labeled secondary antibody and with TOTO 1 ( 1 fiM). a fluorescent DNA stain. Labeled nuclei in whole mounts were viewed by 3D-confocal microscopy and 3D data sets were analyzed using NIH Image 1.61. Results. 5-FU reduced mitosis by >98% , as determined by counting mitoti : figures stained with TOTO-1 and labeling with BrdU at the end of the treatment period. Mitosis was not inhibited in the contralateral, untreated eye. A similar nu nber of BrdU-labeled daughter cell pairs was present in the most superficial layers of 5-FU-treated and control epithelia. However, fewer BrdU-labelcd cells left the basal la /er during the treatment period in 5-Fl J-treated corneas than in controls. Cell density in the basal layer was similar in 5-FU-treated and contralateral control corneas. Conclusions. The probability that, after mitosis, cells wil1 leave the basal layer and diffère n tiatr is influenced by the rate of mitosis in surrounding basal cells. Although the daughter cells produced by cell division behave identically, their joint commitment to div de or to differentiate is not made until after mitosis. Supported by NIH EY04853, a MEI Core Grant for Vision Research and a Jules and Doris Stein RPB Professorship to DCB. None.
|Journal||Investigative Ophthalmology and Visual Science|
|State||Published - 1997|