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Erika L. Silverman, Jin Zhao, Takayuki Nagasaki; Centriole Position As A Directional Marker Of Homeostatic Corneal Epithelial Cell Movements. Invest. Ophthalmol. Vis. Sci. 2012;53(14):1823.
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Corneal epithelial cells exhibit continuous centripetal movements at a rate of about 30 µm per day, but the driving force is not known. The direction of cell movements - mostly centripetal - can be determined by a time-lapse recording with multiple time points, but existing techniques are not adequate to do so with only one time point in both live and histological specimens. To facilitate the investigation of homeostatic cell movements, we examined if the intracellular position of a centriole relative to the nucleus can be used as a directional marker of epithelial cell movements in the mouse cornea.
Ubiquitous GFP mice and C57BL/6J mice were used. Direction of cell movements was determined by a time-lapse recording of GFP-positive epithelial cells in live mice; it was estimated in fixed specimens from a pattern of sub-epithelial nerve fibers, stained with either beta-tubulin or Thy-1, and a pattern of epithelial GFP stripes. The position of centrioles was determined by gamma-tubulin immunohistology with whole-mount corneal specimens. The centriole position relative to the nucleus, stained with DAPI or Hoechst, was determined in relation to the direction of cell movements.
Epithelial cell movements in a normal cornea are generally in the centripetal direction from the limbus to the central cornea. However, the trail of movements is not always a straight radial line, and it frequently forms a curved path. In the central cornea, cell movements roughly follow a shape of a logarithmic spiral and converge in the center. When we determined the position of centrioles in areas of a cornea where cell movements follow generally along a radial path, there were significantly more cells, about 60% of total cells, that contained a centriole in front of a nucleus. On the other hand, in the central cornea where cells exhibit a spiral pattern of movements, there was no statistical difference as to whether a centriole was in front or back of the nucleus. The rate of cell movements in this area is considerably slower, which may be related to the centriole positioning.
Centrioles tend to be positioned in front of the nucleus in corneal basal epithelial cells when they are moving centripetally at a steady rate along the corneal radius. Although the direction of individual cells cannot be determined (only at a 60% precision), the direction of a group of cells can be statistically predicted from this data. Since epithelial cells move together as a group in a cornea, this technique may be suitable for predicting the direction of a given group of cells in both live eyes and histological specimens without resorting to a time-lapse recording.
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