Abstract

This study investigates two destructive biophysical mechanisms during freezing (extensive dehydration and intracellular ice formation) at the cellular level in the rodent ELT-3 uterine fibroid cell-line. The osmotically inactive volume fraction (Vb) of ELT-3 cells was approximated to 0.35 of the initial isotonic cell volume (Vo). The water transport characteristics of this cell-line are such that ELT-3 cells are highly permeable with a strong ability to lose water even at low subzero temperatures. The hydraulic reference permeability, Lpg and activation energy, Elp associated with Lp were found to be 0.13 (μm/min.atm) and 19.0 (kcal/mole) [R2 = 0.86] respectively. Intracellular Ice Formation (IIF) occurs at lower temperatures than many cell-types (i.e. TIIF 50% below −15°C) at cooling rates > 25 °C/min. Darkening IIF, which was assumed to occur by Surface Catalyzed Nucleation (SCN), is governed by kinetic Ωo and thermodynamic κo biophysical parameters, which were found to be 6.1×108(m2.s)−1 and 5.3×109(K5) [R2 = 0.94] respectively. At a cooling rate of 100°C/min, twitching IIF (non-darkening IIF) was observed. Viability data from a separate study (Bischof et. al., 2000) indicated that at cooling rates ≤ 1°C/min and ≥ 50°C/min with an end-temperature of −20°C, extensive damage to cells was observed. The current biophysical study shows that extensive dehydration occurs at 1°C/min while substantial IIF (77%) occurs at 50 °C/min. This data suggests that while biophysics can explain some of the destruction occurring at the investigated temperatures, other effects or mechanisms may be playing a role at lower end-temperatures.

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