Cryopreservation of isolated rat hepatocytes |
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Authors: | Deborah L Novicki Grace P Irons Stephen C Strom Randy Jirtle George Michalopoulos |
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Institution: | (1) Department of Pathology, Duke University Medical Center, 27710 Durham, North Carolina;(2) Department of Radiology, Duke University Medical Center, 27710 Durham, North Carolina |
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Abstract: | Summary Isolated parenchymal hepatocytes from adult rats were frozen in media containing 10% glycerol, 10% dimethylsulfoxide (DMSO),
or 20% DMSO. Three microsome-associated functions were compared in nonfrozen cells and cells frozen in each of the above cryoprotectant
solutions. Freezing in DMSO maintains cytochromes P-450 and b5 and NADPH-cytochrome C reductase at levels nearer to control values than does freezing in glycerol. Cells frozen and subsequently
thawed and cultured for 24 h lose a greater amount of cytochrome P-450 than do nonfrozen cultured cells. The levels of cytochrome
b5 and reductase in frozen-thawed cells remain close to control values. Cell viability (trypan blue dye exclusion and percentage
of attached cells) after freezing is maintained better using DMSO as a cryoprotectant. Dimethylsulfoxide protects the hepatocytes
from freeze-induced damage to the extent that many viable cells attach to collagen-coated petri dishes, survive for at least
24 h, and still maintain significant levels of enzymes of importance to drug and carcinogen metabolism.
This work was supported by Grant CA-30241 from the National Institutes of Health, Bethesda, Maryland. |
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Keywords: | freezing hepatocytes dimethylsulfoxide cell viability cytochrome P-450 |
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