Ph.D. Proposal Presentation by Shannon L. Stott
Wednesday, August 25, 2004

(Dr. Jens O. M. Karlsson , Chair)

"Effects of Cell-Substrate and Cell-Cell Interactions on Intracellular Ice Formation in Tissue"

Abstract

The long-term goal of this research is to develop optimal freezing procedures for tissue containing one or more cell types. The proposed approach is to elucidate the mechanisms of damage associated with the freezing process, and to develop theoretical models of these mechanisms, towards the rational design and computer-aided optimization of techniques for cryopreservation or cryosurgery. Although this approach has previously been successful in the development of cryopreservation protocols for suspensions of isolated cells, the response of intact tissue is known to be fundamentally different from that of cell suspensions. In particular, the probability of intracellular ice formation (IIF), a major cause of cryoinjury, is significantly larger during tissue freezing than during freezing of cell suspensions. Because the reasons for these differences are not well known, the immediate objectives of the proposed research are to elucidate the dominant mechanisms of IIF in tissue constructs, to develop theoretical models of these mechanisms, and to test the predictions of the models. The underlying hypothesis of the proposed research is that the probability of IIF is enhanced in tissue as a result of cell-substrate and cell-cell interactions; in particular, cell-substrate interactions are hypothesized to predispose cell membranes to mechanical damage caused by extracellular ice, while cell-cell interactions are hypothesized to enable propagation of intracellular ice. To validate this hypothesis, a series of well defined cryomicroscopy experiments using micropatterned cells will be proposed. Specifically, the aims of this proposal are: (i) to determine the effects of cell adhesion on cryoinjury, (ii) to elucidate the mechanism of intercellular ice propagation in primary cells, and (iii) to predict the kinetics and distribution of IIF in tissue constructs containing one or more cell types. The results of this proposed work will remove the ambiguity surrounding the effects of cell-substrate and cell-cell interactions on IIF, enabling the creation of mathematical models that accurately predict the freezing kinetics for both homogeneous and heterogeneous tissues.