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Maryam A. Shafiq, Richard A. Gemeinhart, Beatrice Y. Yue, Ali R. Djalilian; Development of an Acellular Human Cornea to Support the Growth of Corneal Epithelial and Stromal Cells. Invest. Ophthalmol. Vis. Sci. 2011;52(14):296.
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© ARVO (1962-2015); The Authors (2016-present)
To decellularize a human cadaver cornea to use as a scaffold for reconstruction of corneal epithelium and a stromal niche.
Human cadaver corneas were decellularized by seven different processing methods including detergent and non-detergent approaches. The success of each method on the removal of cells from the cornea was investigated. The structural integrity of the decellularized corneas was compared with the native cornea by scanning electron microscopy. The integrity of the basement membrane of the epithelium was analyzed by Periodic Acid Shiff (PAS) staining and by the expression of collagen-IV. Finally, the ability of the decellularized corneas to support the growth of human corneal epithelial cells and fibroblasts were assessed in vitro.
Corneas processed using sodium dodecyl sulfate (SDS); Triton X-100; liquid nitrogen; polyethylene glycol; alcohol plus trypsin; and freeze thaw plus NaCl all resulted in incomplete removal of cellular material. Corneas processed with the hypotonic-hypertonic buffers and corneas treated with NaCl followed by DNAse and RNAse both successfully removed all cellular material. Both of these methods also showed the presence of intact basement membrane. However, corneas treated with hypotonic-hypertonic buffers only supported fibroblasts but not epithelial cells. On the other hand, corneas processed with NaCl, DNAse and RNAse supported both fibroblast and epithelial cell growth in vitro.
Decellularized human corneas provide a scaffold which can support the growth of corneal epithelial cells and stromal fibroblasts. This approach may be useful for reconstructing the cornea and limbus using autologous cells.
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