Publication: Characterization of organotypic keratinocyte cultures on de-epithelialized bovine tongue mucosa
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Date
2002
Authors
Hildebrand, H.C. ; Häkkinen, L. ; Wiebe, C.B. ; Larjava, H.S.
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Publisher
Murcia : F. Hernández
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DOI
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info:eu-repo/semantics/article
Description
Abstract
Organotypic cultures have been used to study
epithelial cell behavior for many years. The aim of this
study was to develop an organotypic culture method that
better mimics the three-dimensional morphology of
interdigitating rete ridges and connective tissue papillae
and that also conserves the basement membrane zone
(BMZ).
Bovine tongue mucosa connective tissue, separated
from epithelium after 1M NaCl incubation, was used as
o rganotypic culture substratum for different human
keratinocyte cell lines. Organotypic cultures were
characterized by electron and immunofluorescence
microscopy for expression of integrin subunits and
extracellular matrix components. While spontaneously
immortalized mucosal keratinocytes produced highly
irregular stratified organotypic cultures, the normal
human epidermal keratinocytes (NHEK) demonstrated
culture morphology that resembled in vivo epidermis.
However, in this model, the histomorphology, expression
of differentiation markers involucrin, keratin 10 and 14,
and integrins varied significantly between the cell lines.
Some cultures appeared to have an extended survival
since they were maintained up to 40 days without
histological signs of degeneration. The ultrastructure of
the BMZ including hemidesmosomes was similar to the
normal dermo-epidermal junction. Extracellular matrix
molecules, including tenascin, laminin-1 and -5, were
expressed in the cultures demonstrating their secretion
solely by keratinocytes. Distribution and expression of
integrins in NHEK cultures was similar to that seen in
vivo skin with the exception of additional expression of
a5ß1 and avß6 integrins. Organotypic NHEK cultures
show similarities to normal stratified epithelium and are
potentially useful for multiple applications for studies on
epithelial cell behavior in vitro.
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