Cell & Tissue Kinetics

The Cell and Tissue Kinetics Core was developed to facilitate skin-related research by assisting the center investigators in isolation and cultivation of primary skin cells in two and three-dimensional cultures and in analysis of genes related to skin diseases by efficient and systematic generation of transgenic tissue models of skin that show either transgene overexpression or suppression.

Construction of Organotypic Skin Cultures
Culturing human keratinocytes at the air-liquid interface and supporting them by a dermal substitute matrix triggers a differentiation cascade more reminiscent of naïve epidermis. This experiment provides an excellent model for human epidermis when structure integrity of the tissue is required. This three-dimensional model system can reveal biological activities of genes (e.g., oncogenes) that are not apparent in submerged culture models. Furthermore, as organotypic cultures are composed of the dermal and the epidermal compartments, epidermal-mesenchymal interactions can be studied by genetic modification in one or both compartment. We will assist investigators in the design of chimeric organotypic rafts, according to the specifics of the proposed projects. The generated tissue will be provided to the investigator for further molecular/biochemical analysis or to Core A for morphological analysis.

Mouse Cell Culture Services
Skin of one-two day old newborn mice is washed twice with Betadine and twice with 70% ethanol. The skin will then be removed and the epidermis and the dermis will be separated following 0.25% trypsin digestion. Both epidermal cells and dermal fibroblasts can be harvested and either freshly harvested keratinocytes or primary cultures (or both) will be provided to the investigator.

Mouse Skin Reconstitution
In this assay, fresh isolated or cultured neonatal mouse epidermal cells are mixed with mouse fresh isolated or cultured dermal fibroblasts, and implanted as a slurry on the mouse fascia under a transplantation chamber to regenerate a well-differentiated epidermis and associated pilosebaceous units. During culture period, either cell population may be transduced with lenti- or retro-viral vectors and reconstructed on the back of a nude mouse. Fully differentiated and normalized skin is formed in 4-6 weeks. The genetically engineered skin will be provided to the investigators for future analysis.

Primary Human Cell Cultures
In this assay, Six-mm punch biopsies from normal or patient skin will be provided by the investigator, using their own IRB approval.  Both keratinocytes and fibroblasts isolated from the given sample can be provided to upon request (cryopreserved vial or in a T-25 tissue culture flask).