Kuleshova L. The mechanisms of effects of extracellular ice formation and related phenomena on biological objects.

Subject - the process of extracellular ice formation and responds of human and animal cells to physico-chemical factors which occur owing to the phase transition under cryopreservation. Aim - to ascertain mechanisms of initiation and growth of extracellular ice crystals in cryopreserved biological systems and mechanisms of action of physico-chemical factors associated with the phase transition "liquid-crystal" on cells. Methods: - cryomicroscopy, low temperature roentgenography, microscopy, spectrophotometry, 1H- NMR spectroscopy, physico-mathematical modelling. A new method for determination of the isotropic tension modulus of rat hepatocyte membranes was theoretically substantiated. The algorithms for determination of transport characteristics of cell membranes under conditions of extracellular ice formation and for prediction of optimal cooling rates were developed. It was quantitatively proved that the main mechanism of solid phase onset in model biological solutions was heterogeneous nucleation and extracellular ice crystal growth was simultaneously realised through normal and dislocation mechanisms of real crystal growth. It was shown that combination of stepwise preliminary dehydration of cells and saturation of them with a penetrating cryoprotector was necessary to avoid safely intracellular ice formation. The mechanism of post-hypertonic lysis of rat hepatocytes under rehydration conditions at warming stage was ascertained. The results of the work can be used during rapid search of optimal cryopreservation conditions of different cell suspensions.