Affiliations: Experimental Fetal Surgery and Tissue Engineering Unit, Department of Pediatric and Adolescent Surgery, Medical University of Graz, Graz, Austria | Biobank Graz, Medical University of Graz, Graz, Austria | Group leader of Angiogenesis and Tissue Engineering; RAFT Institute of Reconstructive Plastic Surgery Research, Mount Vernon Hospital, Northwood, Middlesex, England | Imperial College London, Chelsea Children's Hospital, Chelsea and Westminster NHS Foundation Trust, London, UK
Note: [] Address for correspondence: Amulya K. Saxena, MD, Experimental Fetal Surgery and Tissue Engineering Unit, Department of Pediatric and Adolescent Surgery, Medical University of Graz, Auenbruggerplatz-34, A-8036 Graz, Austria. Tel.: +43 316 385 13762; Fax: +43 316 385 13775; E-mail: [email protected]
Abstract: PURPOSE: The yield of a critical number of basal epithelial cells with high mitotic rates from native tissue is a challenge in the field of tissue engineering. There are many protocols that use enzymatic methods for isolation of epithelial cells with unsatisfactory results for tissue engineering. This study aimed to develop a protocol for isolating a sufficient number of epithelial cells with a high Proliferating Index from ovine esophagus for tissue engineering applications. METHODS: Esophageal mucosa was pretreated with dispase–collagenase solution and plated on collagen-coated culture dishes. Distinction of the various types of epithelial cells and developmental stages was done with specific primary antibodies to Cytokeratins and to Proliferating Cell Nuclear Antigen (PCNA). RESULTS: Up to approximately 8100 epithelial cells/mm2 of mucosa tissue were found after one week of migration. Cytokeratin 14 (CK 14) was positive identified in cells even after 83 days. At the same time the Proliferating Index was 71%. CONCLUSION: Our protocol for isolation of basal epithelial cells was successful to yield sufficient numbers of cells predominantly with proliferative character and without noteworthy negative enzymatic affection. The results at this study offer the possibility of generation critical cell numbers for tissue engineering applications.
