Ex vivo produced human conjunctiva and oral mucosa equivalents grown in a serum-free culture system.
OBJECTIVE
We sought to develop full-thickness ex vivo produced human conjunctiva and oral mucosa equivalents using a serum-free culture system without a feeder layer and to compare conjunctiva and oral mucosa equivalents to assess their suitability as graft materials for eyelid reconstruction.


METHODS
Human conjunctival and oral mucosal keratinocytes were cultured, expanded, and seeded onto AlloDerm (LifeCell Corp, Branchburg, NJ), a cadaveric, acellular dermis, to produce ex vivo produced full-thickness mucosa equivalents. Histology of equivalents and their expression of immunoreactive Ki-67, a proliferation marker, and GLUT1, a membrane antigen seen in barrier tissues, were examined at 4, 11, and 18 days after seeding onto AlloDerm.


RESULTS
Progressive epithelial stratification was observed on day 4, 11, and 18 conjunctiva and oral mucosa equivalents. Ki-67 immunoreactivity progressively increased with cultured time in both types of equivalent, indicating the continued presence of actively proliferating cells. GLUT1 immunoreactivity, concentrated in the basal keratinocytes of stratified epithelia of both types of equivalents, mimicked native tissue and indicated a high glycolytic state of the basal cells.


CONCLUSIONS
Conjunctival and oral mucosal equivalents are similar to native tissue and demonstrate high proliferative and glycolytic states. Due to the similarity to conjunctiva, oral mucosal equivalents may be useful for eyelid reconstruction. Their advantages for surgical reconstruction include 1) ease of obtaining autogenous oral epithelium for expansion in vitro without the possibility of contaminating cellular- or serum-borne biologic agents, 2) growth of intact, confluent epithelia on rigid, transplantable human allogeneic dermis that may be surgically transplanted, and 3) reduced donor site morbidity and surgical time.
Yoshizawa
Michiko
M
Department of Oral and Maxillofacial Surgery, University of Michigan Medical Center, Ann Arbor, MI 48109, USA.
Feinberg
Stephen E
SE
Marcelo
Cynthia L
CL
Elner
Victor M
VM
D001672:Biocompatible Materials; D002455:Cell Division / Q000502:physiology; D002478:Cells, Cultured; D003094:Collagen; D003228:Conjunctiva / Q000166:cytology*; D016895:Culture Media, Serum-Free; D004847:Epithelial Cells / Q000166:cytology; D005143:Eyelids / Q000601:surgery; D051272:Glucose Transporter Type 1; D006019:Glycolysis / Q000502:physiology; D006801:Humans; D015603:Keratinocytes / Q000166:cytology* / Q000637:transplantation; D019394:Ki-67 Antigen / Q000032:analysis; D009004:Monosaccharide Transport Proteins / Q000032:analysis; D009061:Mouth Mucosa / Q000166:cytology*; D013997:Time Factors; D014019:Tissue Donors; D023822:Tissue Engineering
C440568:Alloderm; D001672:Biocompatible Materials; D016895:Culture Media, Serum-Free; D051272:Glucose Transporter Type 1; D019394:Ki-67 Antigen; D009004:Monosaccharide Transport Proteins; C495677:SLC2A1 protein, human; D003094:Collagen