Analysis of p63 and cytokeratin expression in a cultivated limbal autograft used in the treatment of limbal stem cell deficiency.
OBJECTIVE
To investigate the expression of p63 and cytokeratins throughout the course of producing a cultivated autograft of limbal epithelial cells.


METHODS
A 75 year old male with a severe alkali burn to his right eye received two cultivated autografts of limbal epithelial cells on amniotic membrane followed by a corneal allograft. Immunostaining for p63 and cytokeratins was performed during ex vivo expansion with 3T3 fibroblasts, following subcultivation on amniotic membrane, and on the excised corneal button.


RESULTS
Cultures grown in the presence of 3T3 fibroblasts or on amniotic membrane displayed positive staining for keratins 14 and 19, and p63, but poor staining for keratin 3 (K3). The excised corneal button possessed a stratified epithelium of K3 positive cells residing on amniotic membrane.


CONCLUSIONS
Our results document for the first time the co-expression of cytokeratins 14 and 19 with p63 in a cultivated limbal graft. These data support the conclusion that cultivated grafts of limbal epithelium contain predominantly undifferentiated cells with the potential to regenerate a normal corneal epithelium.
Harkin
D G
DG
Tissue BioRegeneration and Integration Program, Institute for Health and Biomedical Innovation, Queensland University of Technology, 2 George Street, Brisbane 4000, Australia. d.harkin@qut.edu.au
Barnard
Z
Z
Gillies
P
P
Ainscough
S L
SL
Apel
A J G
AJ
D000368:Aged; D002057:Burns, Chemical / Q000378:metabolism / Q000601:surgery*; D002478:Cells, Cultured; D016039:Corneal Transplantation / Q000379:methods; D004268:DNA-Binding Proteins; D004847:Epithelial Cells / Q000737:chemistry* / Q000637:transplantation; D019573:Epithelium, Corneal / Q000737:chemistry* / Q000637:transplantation; D005126:Eye Burns / Q000378:metabolism / Q000601:surgery*; D016147:Genes, Tumor Suppressor; D006801:Humans; D053547:Keratin-14; D053558:Keratin-3; D007633:Keratins / Q000032:analysis*; D016850:Limbus Corneae / Q000473:pathology*; D008297:Male; D010641:Phenotype; D010750:Phosphoproteins / Q000032:analysis*; D033581:Stem Cell Transplantation / Q000379:methods; D015534:Trans-Activators / Q000032:analysis*; D014157:Transcription Factors; D014182:Transplantation, Autologous; D025521:Tumor Suppressor Proteins
D004268:DNA-Binding Proteins; C507540:KRT14 protein, human; C479360:KRT3 protein, human; D053547:Keratin-14; D053558:Keratin-3; D010750:Phosphoproteins; C115051:TP63 protein, human; D015534:Trans-Activators; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; D007633:Keratins