An improved and rapid method to construct skin equivalents from human hairfollicles and fibroblasts

To produce sufficient amounts of high quality skin equivalents (SE), either allogenic for dermatopharmacological and dermatotoxicological studies or a utologous for transplantation purposes, we established a rapid, easy and co st effective three-dimensional SE model on the basis of human dermal fibrob lasts. collagen and freshly plucked hair follicles. Acidic liquid collagen was polymerized with sodium hydroxide in the presence of fibroblasts to for m a dermal equivalent (DE) resembling normal human dermis. At 24 h later. f reshly plucked hair follicles were implanted into the surface of these DEs after cutting their bulbs off. Another 48 h later, the surface of the SEs w as lifted to the air-liquid interface. Fourteen days after implantation, ou tgrowing keratinocytes from the outer root sheath of the hair follicles com pletely covered the surface of. the SE and built a fully developed, multi-l ayered and cornified epidermis. Histology and immunofluorescence studies wi th specific antibodies directed against components of keratinocytes, fibrob lasts, cell-adhesion molecules, different extracellular matrix and basement membrane proteins revealed the similarity of our three-dimensional SEs to the in vivo situation in normal human skin. Using autologous cell sources a nd cell culture media enriched with serum from the respective cell donor, i t will be possible to use these SEs for autologous transplantation, thereby reducing the risk of transplant rejection.