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S.L. Ainscough, Z. Barnard, D.G. Harkin, I.R. Schwab; Different Populations of Fibroblasts Exist in the Corneal Stroma, Limbus and Sclera . Invest. Ophthalmol. Vis. Sci. 2006;47(13):2978.
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To determine whether different populations of fibroblasts reside in the stroma of the cornea, limbus and sclera.
Fibroblasts were isolated from the cornea, limbus and sclera of human donor anterior segments using collagenase digestion and subsequently cultured in DMEM + 10% FBS. A series of photographs were taken for morphometry and visual analysis at low cell density and confluency. Morphometrical data was obtained by measuring the area, perimeter and elliptical factor of low cell density photographs of 100 cells each from the cornea, limbus and sclera using the Scion Imaging software. Time lapse movies were composed to observe further changes in morphology between the cell populations. Whole cell lysates from each sample were then subsequently analysed using 2D gel electrophoresis.
Visual analysis suggests that fibroblasts isolated from the cornea, limbus and sclera demonstrate morphological differences. Fibroblasts isolated from the sclera have a random orientation at confluence, are larger and less spindle–shaped than corneal and limbal fibroblasts. Fibroblasts isolated from both the cornea and limbus have a spindle–shaped morphology. However, fibroblasts from the cornea possess a distinct orientation at confluence whereas cells from the limbus are more random. These results were confirmed using morphometrical analysis on the different cultures at subconfluence. Statistically significant differences in area, perimeter and elliptical factor between the three populations demonstrated that fibroblasts isolated from the sclera were less spindle–shaped than those extracted from the limbus and cornea. Time lapse movies indicated that these morphological differences were consistent over time. Proteomic analysis has also demonstrated that the fibroblast populations are distinct. 2D gel electrophoresis shows that protein expression levels vary for multiple proteins and further analysis is being conducted to clarify the extent of these differences.
Distinct populations of fibroblasts can be isolated from the stroma of the cornea, limbus and sclera. Morphometrical and proteomic data indicate that at least two or possibly three different populations of fibroblasts reside in the anterior segment.
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