Ultrastructural evaluation of human keratinocyte growth and differentiation on a fibrin substrate.
OBJECTIVE
In order to circumvent several difficulties that have been met in the routine use of the in vitro keratinocyte cultures using the standard procedure described by Rheinwald and Green, and obtain a more resilient and the least possible immunogeneic skin substitute for a future clinical application, this work studied a new keratinocyte culture system, which envisages the utilization of a fibrin substrate in association with high densities of human keratinocytes.


METHODS
Through light and transmission electron microscopy and immunohistochemical assays, long-term proliferative and differentiative characteristics of keratinocytes cultured onto a fibrin gel under immerse and air-liquid interface culture conditions were evaluated.


RESULTS
Despite the absence of a dermal substitute, the results demonstrated that the proposed composite was constituted of a transparent and elastic fibrin film covered by a well-attached, multistratified epithelium with morphological characteristics that resemble human epidermis, including the neoformation, albeit incomplete, of the basement membrane.


CONCLUSIONS
Increased mechanical resistance due to the presence of an easy handling substrate, the delivery of nonclonfluent keratinocytes as well as the removal of animal-derived cells from the culture system suggest its potential use for future transplantation purposes.
Department of Surgery, USP, Sao Paulo, Brazil. danielatanikawa@gmail.com
Daniela Yukie Sakai
DY
Tanikawa
Nivaldo
N
Alonso
Marisa Roma
MR
Herson
Monica Beatriz
MB
Mathor
Elia Garcia
EG
Caldini
Silvia Vanessa
SV
Lourenço
Marcus Castro
MC
Ferreira
D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation / Q000502:physiology*; D049109:Cell Proliferation; D005337:Fibrin / Q000737:chemistry*; D005782:Gels / Q000737:chemistry*; D006801:Humans; D015603:Keratinocytes / Q000166:cytology / Q000648:ultrastructure*
D005782:Gels; D005337:Fibrin