[Ultrastructural and immunohistochemical study of 3-dimensional cultures of human keratinocytes on a collagen gel].
OBJECTIVE
To investigate the ultrastructural and immunochemical features of three-dimensional cultures of human keratocytes in collagen gel matrix.


METHODS
Human keratocytes were obtained from primary cultures of stromal explains. They were cultured in bovine type I collagen gel matrix for 6 weeks. Keratocyte-populated gels were analyzed by means of immunochemistry and transmission electron microscopy.


RESULTS
Human keratocytes cultured in collagen gel matrix developed processes and formed networks of connecting cells. They showed positive staining for vimentin, collagen I, V, and VI, and connexin. Electron microscopy showed elongated cells with processes and gap junctions. Keratocytes synthesized collagen fibrils and filaments. No fibrils' organization similar to that observed in the normal human corneal stroma (i.e. parallel bundles of collagen fibrils) was observed.


CONCLUSIONS
Ultrastructure and immunochemical phenotype of three-dimensional cultures of human keratocytes in collagen gel matrix are similar to those observed in situ. These cultures represent a useful in vitro model to study the different corneal stroma components.
Banque de Cornées Saint-Antoine, ETS de l'Hôpital Saint-Antoine, Paris.
N
N
Mourra
V
V
Borderie
L
L
Laroche
D000818:Animals; D002417:Cattle; D018929:Cell Culture Techniques / Q000379:methods*; D002478:Cells, Cultured; D003094:Collagen; D017630:Connexins / Q000032:analysis; D003319:Corneal Stroma / Q000166:cytology*; D003893:Desmin / Q000032:analysis; D016326:Extracellular Matrix Proteins / Q000032:analysis; D017629:Gap Junctions / Q000648:ultrastructure; D005782:Gels; D006801:Humans; D007124:Immunoenzyme Techniques; D015603:Keratinocytes / Q000737:chemistry / Q000276:immunology / Q000648:ultrastructure*; D014746:Vimentin / Q000032:analysis
D017630:Connexins; D003893:Desmin; D016326:Extracellular Matrix Proteins; D005782:Gels; D014746:Vimentin; D003094:Collagen
Etude ultrastructurale et immunohistochimique des cultures tridimensionnelles de kératocytes humains dans un gel de collagène.