A basement membrane-like matrix formed by cell-released proteins at the medium/air interface supports growth of keratinocytes.
Epithelial cells require adherence to a matrix for regular growth. During standard keratinocyte cell culture in serum-free medium, we observed that cell colonies formed not only on the bottom of the culture vessels but also at the medium/air interface. Coomassie blue staining detected a protein membrane that extended up to several centimeters between the colonies of floating cells. Ultrastructural investigation of this membrane revealed structures closely resembling those of basement membranes, and immunochemical staining confirmed the presence of laminins-1 and -5 as well as collagen IV, representative components of basement membranes. Cells attached to the floating membrane proliferated and could be cultivated for up to six months. When keratinocyte-conditioned medium was filtered and transferred to a culture vessel without cells, the protein membrane at the liquid/air interface formed within one week suggesting self-assembly of cell-released proteins. Our findings provide a basis for the production of epidermal basement membranes for potential medical uses.
Eckhart
Leopold
L
Department of Dermatology, University of Vienna Medical School, Vienna, Austria.
Reinisch
Christina
C
Inoue
Sadayuki
S
Messner
Paul
P
Dockal
Michael
M
Mayer
Christoph
C
Tschachler
Erwin
E
D001485:Basement Membrane / Q000166:cytology* / Q000378:metabolism; D002448:Cell Adhesion / Q000502:physiology; D002455:Cell Division / Q000502:physiology; D002478:Cells, Cultured; D024141:Collagen Type IV / Q000378:metabolism*; D016895:Culture Media, Serum-Free / Q000737:chemistry; D000078404:Epidermal Cells; D004817:Epidermis / Q000378:metabolism; D006801:Humans; D015603:Keratinocytes / Q000166:cytology* / Q000378:metabolism; D007797:Laminin / Q000378:metabolism*; D008854:Microscopy, Electron
D024141:Collagen Type IV; D016895:Culture Media, Serum-Free; D007797:Laminin