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Analysis of p63 and cytokeratin expression in a cultivated limbal autograft used in the treatment of limbal stem cell deficiency

¹ Tissue BioRegeneration and Integration Program, Institute for Health and Biomedical Innovation, Queensland University of Technology, Brisbane, Queensland, Australia
² School of Biomolecular & Biomedical Science, Griffith University, Brisbane, Queensland, Australia
³ Queensland Skin Bank, Australian Red Cross Blood Service, Brisbane, Queensland, Australia
⁴ Princess Alexandra Hospital, Brisbane, Queensland, Australia

Correspondence to:
Correspondence to:
Dr D G Harkin
Tissue BioRegeneration and Integration Program, Institute for Health and Biomedical Innovation, Queensland University of Technology, 2 George Street, Brisbane 4000, Australia; d.harkin{at}qut.edu.au

Aim: 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.

Abbreviations: EDTA, ethylene diamine tetra-acetic acid; FBS, fetal bovine serum; K3, keratin 3; PBS, phosphate buffered saline

Keywords: corneal stem cell deficiency; cultivated limbal graft; amniotic membrane; p63; cytokeratins

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