Effects of cell-cell and cell-substrate interactions on ice formation in micropatterned tissue constructs

The preservation of biological materials by means of cryogenic temperatures is the preferred method today for preserving reproductive cells and other types of isolated cells. The new challenge for cryopreservation comes from the increasing demand for organs and tissues for transplantation, and the lack of a satisfactory protocol for preserving tissues over long periods of time. Extending the time between the moment an organ becomes available, from a donor, or through tissue engineering, and the moment when it is needed by a patient, would improve the availability of biological materials. The main obstacle in preserving tissues is that our knowledge of the cryobiology of isolated cells can not simply be extrapolated to multicellular structures, because the low temperature behavior of cells in tissues is different from that of isolated cells. In particular, the process of intracellular ice formation, the major mode of cell damage during cryopreservation, is known to be different in isolated and cultured cells.; Simple tissue engineered constructs, involving a limited number of cells, were used for qualitative and quantitative studies of the intracellular ice formation in tissues. By using a micropatterning technique, cell geometry, time in culture, and the cell-cell interaction, were independently controlled. This has allowed the separation of the cell-cell interaction and cell geometry effects on the probability of intracellular ice formation. Through mathematical modeling, the influence of each of these factors on the ice formation process was quantified, and details of the cell-cell interaction, e.g. the role played by gap junctions, and the influence of cell shape and cell-substrate interactions on the behavior of cells during freezing, could be elucidated. The quantitative knowledge gained from simple cellular systems can be used to make predictions about the behavior of more complex systems at low temperatures, to better design freezing protocols for the preservation of tissues, and to model the ice formation in tumor tissues, with relevance for cryosurgery.