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Martyna M. Urbanowicz, Jin Zhao, Takayuki Nagasaki; Spatial Distribution Of Cell Divisions In The Basal Epithelium Of Mouse Cornea. Invest. Ophthalmol. Vis. Sci. 2011;52(14):320.
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© ARVO (1962-2015); The Authors (2016-present)
Corneal epithelium is a dynamic tissue where its homeostasis is maintained by a delicate balance of cell division, cell movements, and cell loss. As such, spatial distribution of dividing cells is a critical parameter but there have been disagreements whether cell division is more frequent in the peripheral cornea or in the central cornea. To resolve the issue, spatial distribution of cell division was determined in the basal epithelium of mouse cornea, along the entire length of a diameter from the limbus to the limbus.
Cell division rates were determined by in vivo and histological methods. For the former, either 5-bromo-2-deoxyuridine (BrdU) or 5-ethynyl-2'-deoxyuridine (EdU) was delivered systemically with a subcutaneous osmotic pump, and their incorporation in dividing cells over a 24-hour period was determined in live mice. For the latter, distribution of cells expressing Ki-67 antigen or Ser10-phosphorylated histone H3 was determined by immunohistochemistry. For both methods, digitized microscopic images of whole-mount specimens were used to count cells that expressed a marker of interest in the basal layer of the epithelium. Every cell was scored within two rectangular strips of the cornea, 50 to 100 µm wide, along the length of a diameter from the limbus to the limbus, not including the limbus, one in the superior-inferior and the other in the nasal-temporal direction. The total cell number in the same area was determined by counting the number of nuclei that were labeled with either 4',6-diamidino-2-phenylindole (DAPI) or Hoechst 33258.
Nuclear labeling with DAPI or Hoechst revealed that the density of basal epithelial cells was about 15,000 cells/mm2, which was nearly uniform in the entire cornea. The average density of BrdU-positive cells was about 7,000 cells/mm2, indicating that nearly a half of all basal cells were in an S-phase of the cell cycle during a 24-hour period. The proportion of BrdU-positive cells was at least 20% less in a zone of central 20% than in the rest of the cornea. There was no significant difference between a superior-inferior strip and a nasal-temporal strip of the same cornea. Similar results were obtained with Edu labeling and phosphorylated histone H3 staining.
We determined the rates of basal epithelial cell division in the entire diameter of the mouse cornea from the limbus to the limbus. A cell division rate is clearly reduced in the central cornea although its biological meaning is not clear at this time. This finding can now be used to develop a quantitative mathematical model of epithelial homeostasis, in combination with the data of cell movements and cell loss.
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