MECHANISMS OF CRYOINJURY AND CRYOPROTECTION IN SPLIT-THICKNESS SKIN

Successful cryopreservation of tissues will ultimately require a more detailed understanding of how the in situ environment modifies cell re sponses during cooling and warming. Low-temperature responses of porci ne split-thickness skin and isolated basal keratinocytes were compared after various cooling protocols and in the presence and absence of cr yoprotectants. Recovery was assessed by measuring oxygen consumption k inetics in skin and tetrazolium reduction in isolated cells. Freeze su bstitution was used to visualize ice nucleation and growth in skin. Th e results indicated that the lime required for diffusion of water in s plit-thickness skin delayed osmotic responses in the basal keratinocyt es and resulted in increased intracellular and intercellular ice forma tion. Rapid cooling (-200 degrees C/min) in the presence of cryoprotec tants resulted in a reduction in the number of cells containing ice an d the size of the intercellular ice crystals and an increase in tissue recovery. These observations support other reports which suggest that cell-to-cell and cell-to-substrate interactions are sensitive sites f or cryoinjury. A practical recommendation from this study is that high recovery of split-thickness skin may be achieved with protocols using high cooling rates. (C) 1996 Academic Press, Inc.