Effect of supercooling and cell volume on intracellular ice formation
| Autor: | Locksley E. McGann, Leah A. Marquez-Curtis, Janet A.W. Elliott, Richelle C. Prickett |
|---|---|
| Jazyk: | angličtina |
| Předmět: |
Cell Membrane Permeability
Cryoprotectant Biology General Biochemistry Genetics and Molecular Biology Cryopreservation 03 medical and health sciences Cryoprotective Agents 0302 clinical medicine Freezing Human Umbilical Vein Endothelial Cells Extracellular Humans Supercooling Cells Cultured Cell Size Medicine(all) Saline Solution Hypertonic 030219 obstetrics & reproductive medicine Agricultural and Biological Sciences(all) Cell Death Biochemistry Genetics and Molecular Biology(all) Cryomicroscopy Ice Osmolar Concentration 0402 animal and dairy science Membrane integrity IIf 04 agricultural and veterinary sciences General Medicine Freezing point depression 040201 dairy & animal science Biochemistry Biophysics Ice nucleus Human umbilical vein endothelial cells (HUVECs) Tonicity Isotonic Solutions General Agricultural and Biological Sciences Intracellular |
| Zdroj: | Cryobiology. (2):156-163 |
| ISSN: | 0011-2240 |
| DOI: | 10.1016/j.cryobiol.2015.02.002 |
| Popis: | Intracellular ice formation (IIF) has been linked to death of cells cryopreserved in suspension. It has been assumed that cells can be supercooled by 2 to 10°C before IIF occurs, but measurements of the degree of supercooling that cells can tolerate are often confounded by changing extracellular temperature and solutions of different osmolality (which affect the cell volume). The purpose of this study was to examine how the incidence of IIF in the absence of cryoprotectants is affected by the degree of supercooling and cell volume. Human umbilical vein endothelial cells were suspended in isotonic (300mOsm) and hypertonic (∼600 to 700mOsm) solutions and exposed to supercooling ranging from 2 to 10°C before extracellular ice was nucleated. The number of cells undergoing IIF was examined in a cryostage (based on the darkening of cells upon intracellular freezing (“flashing”)) as a function of the degree of supercooling, and cell survival post-thaw was assessed using a membrane integrity assay. We found that while the incidence of IIF increased with supercooling in both isotonic and hypertonic solutions, it was higher in the isotonic solution at any given degree of supercooling. Since cells in hypertonic solution were shrunken due to water efflux, we hypothesized that the difference in IIF behavior could be attributed to the decreased volume of cells in the hypertonic solution. Our results confirm that cells with a smaller diameter before extracellular ice nucleation have a decreased probability of IIF and suggest that cell volume could play a more significant role in the incidence of IIF than the extracellular ice nucleation temperature. |
| Databáze: | OpenAIRE |
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