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M.K. Gordon, P. Bhatt, R. Song, R.A. Hahn, M. Shakarjian, D.R. Gerecke, M. Koch; Collagen XXIII Facilitates Adhesion of Corneal Epithelial Cells to Type IV Collagen and Matrigel . Invest. Ophthalmol. Vis. Sci. 2005;46(13):869.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose:Type XXIII collagen is a newly described transmembrane molecule that is expressed on the surface of corneal epithelial and endothelial cells, but not stromal cells. As a transmembrane molecule, it may facilitate cell attachment, migration, proliferation or differentiation. To assess the role of collagen XXIII in cell adhesion, recombinant collagen XXIII was added to dishes coated with various substrata, and incubated 1 hour to permit binding. Immortalized human corneal epithelial cells were then plated on the dishes, and their ability to adhere was measured. Methods:cDNA encoding the extracellular region of collagen XXIII was ligated into pCEP vector and used to stably transfect 293 cells. Medium collected contained the expressed ectodomain of the collagen. Medium from empty pCEP vector transfected into 293 cells was used as a control. 12 well plastic culture plates were used either as is, or coated with type I collagen, type IV collagen, or Matrigel. 2 ml of medium from either the transfected 293 cells expressing the ecodomain of collagen XIII, or transfected with empty pCEP vector, was added to the 12 well plates, allowing interaction with the plastic or substratum for 60 minutes at room temp. The medium was then removed, and 40,000 corneal epithelial cells were added to the wells. One set of wells was incubated for 1 hour at 37oC before removing the medium. Adherent cells were washed, fixed and stained with crystal violet. A second set of cells was incubated for 4 hours, and a third set for 24 hours, before removing medium and staining the adherent cells. Attached cells were monitored by photography and by absorbance at 570 nm using a plate reader. Results:At all time points, corneal epithelial cells attached well to plastic, collagen I, collagen IV, and Matrigel coated plates pre–incubated with medium from empty vector–transfected 293 cells. However, fewer corneal epithelial cells adhered to the collagen IV and Matrigel coated dishes when the wells were pre–incubated with medium containing recombinantly expressed collagen XXIII. Conclusions:Exogenously added type XXIII collagen competes with corneal epithelial cell surface collagen XXIII for binding to type IV collagen and Matrigel. Our results suggest that corneal epithelial cells use collagen XXIII to facilitate interactions with the basement membrane, possibly through type IV collagen.
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