Initiation and characterization of keratinocyte cultures from biopsies of normal human conjunctiva.
The purpose of the present study was to establish and characterize serum-free epithelial cultures of normal human conjunctiva using fresh biopsy tissue. To this end, small pieces of normal conjunctiva were biopsied from patients undergoing routine cataract surgery. Fragments of the tissue were placed in explant culture in medium containing fetal bovine serum for approximately 1 week to promote epithelial cell outgrowth. Cultures were then passaged multiple times into serum-free medium. Cultures generated in this way were at least 95% keratinocytes and exhibited a typical epithelial morphology, which was dependent on the extracellular Ca(2+)concentration. Immunocytochemical staining revealed that E-cadherin, P-cadherin, and involucrin were present in the cultures, with distributions consistent with their in vivo localization patterns. Distribution of keratins 19, 3, and 4 in conjunctival epithelial cultures were also consistent with in vivo patterns and distinctly different from patterns observed in epithelial cultures similarly generated from cornea and foreskin. Hence, conjunctival keratinocyte cultures retain some tissue-specific markers and do not revert to a generic, culture phenotype.
Risse Marsh
Barbara C
BC
Department of Dermatology, University of Pennsylvania, School of Medicine, Philadelphia, PA, 19104, USA.
Massaro-Giordano
Mina
M
Marshall
Christine M
CM
Lavker
Robert M
RM
Jensen
Pamela J
PJ
D001706:Biopsy; D002118:Calcium / Q000494:pharmacology; D018929:Cell Culture Techniques / Q000379:methods; D003228:Conjunctiva / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D016895:Culture Media, Serum-Free; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D006801:Humans; D015603:Keratinocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D007633:Keratins / Q000378:metabolism
D016895:Culture Media, Serum-Free; D007633:Keratins; D002118:Calcium