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Cryopreservation of rat precision-cut liver slices by ultrarapid freezing: influence on phase I and II metabolism and on cell viability upon incubation for 24 hours. 2001

V P Vanhulle, and G A Martiat, and R K Verbeeck, and Y Horsmans, and P B Calderon, and S L Eeckhoudt, and H S Taper, and N Delzenne
Unit of Pharmacokinetics, Metabolism, Nutrition and Toxicology, Université Catholique de Louvain, Brussels, Belgium.

Several cryopreservation methods for precision-cut rat liver slices (PCLS) have been proposed, allowing a short-term (a few hours) maintainance of viability and functionality upon thawing. The aim of the present study was to test the metabolic capacity of PCLS cryopreserved by an ultrarapid method. The biotransformation of paracetamol to its glucuronide and sulfate conjugates and of midazolam to its hydroxylated metabolites was studied in thawed PCLS incubated for 24 hours at 37 degrees C in Williams' medium E. In addition, protein levels of the key enzymes involved in these metabolic reactions, i.e. UGT1A1, ST1A1, CYP2E1 and CYP3A2 were determinated. In addition, biological markers of cell function (ATP and glycogen levels) and toxicity (LDH leakage in the medium) were also measured. Compared to controls (non cryopreserved PCLS), CYP3A2 activity and content and CYP2E1 content were maintained at the same level all along the incubation, whereas paracetamol glucuronidation and sulfation dropped to 24 and 21% of the control value, respectively, immediately after thawing. Freezing-thawing conditions also modified cell functionality, leading to a lower intracellular ATP and glycogen content, and an increase in cell lysis, as shown by LDH released in the medium. The results of this study suggest that cryopreserved PCLS are able to maintain some phase I activities for 24 hours after thawing whereas some phase II metabolic capacities are not maintained.

UI MeSH Term Description Entries
D007527 Isoenzymes Structurally related forms of an enzyme. Each isoenzyme has the same mechanism and classification, but differs in its chemical, physical, or immunological characteristics. Alloenzyme,Allozyme,Isoenzyme,Isozyme,Isozymes,Alloenzymes,Allozymes
D007770 L-Lactate Dehydrogenase A tetrameric enzyme that, along with the coenzyme NAD+, catalyzes the interconversion of LACTATE and PYRUVATE. In vertebrates, genes for three different subunits (LDH-A, LDH-B and LDH-C) exist. Lactate Dehydrogenase,Dehydrogenase, L-Lactate,Dehydrogenase, Lactate,L Lactate Dehydrogenase
D008099 Liver A large lobed glandular organ in the abdomen of vertebrates that is responsible for detoxification, metabolism, synthesis and storage of various substances. Livers
D008297 Male Males
D008874 Midazolam A short-acting hypnotic-sedative drug with anxiolytic and amnestic properties. It is used in dentistry, cardiac surgery, endoscopic procedures, as preanesthetic medication, and as an adjunct to local anesthesia. The short duration and cardiorespiratory stability makes it useful in poor-risk, elderly, and cardiac patients. It is water-soluble at pH less than 4 and lipid-soluble at physiological pH. Dormicum,Midazolam Hydrochloride,Midazolam Maleate,Ro 21-3981,Versed,Hydrochloride, Midazolam,Maleate, Midazolam,Ro 21 3981,Ro 213981
D009924 Organ Culture Techniques A technique for maintenance or growth of animal organs in vitro. It refers to three-dimensional cultures of undisaggregated tissue retaining some or all of the histological features of the tissue in vivo. (Freshney, Culture of Animal Cells, 3d ed, p1) Organ Culture,Culture Technique, Organ,Culture Techniques, Organ,Organ Culture Technique,Organ Cultures
D009926 Organ Preservation The process by which organs are kept viable outside of the organism from which they were removed (i.e., kept from decay by means of a chemical agent, cooling, or a fluid substitute that mimics the natural state within the organism). Organ Preservations,Preservation, Organ,Preservations, Organ
D002470 Cell Survival The span of viability of a cell characterized by the capacity to perform certain functions such as metabolism, growth, reproduction, some form of responsiveness, and adaptability. Cell Viability,Cell Viabilities,Survival, Cell,Viabilities, Cell,Viability, Cell
D003577 Cytochrome P-450 Enzyme System A superfamily of hundreds of closely related HEMEPROTEINS found throughout the phylogenetic spectrum, from animals, plants, fungi, to bacteria. They include numerous complex monooxygenases (MIXED FUNCTION OXYGENASES). In animals, these P-450 enzymes serve two major functions: (1) biosynthesis of steroids, fatty acids, and bile acids; (2) metabolism of endogenous and a wide variety of exogenous substrates, such as toxins and drugs (BIOTRANSFORMATION). They are classified, according to their sequence similarities rather than functions, into CYP gene families (>40% homology) and subfamilies (>59% homology). For example, enzymes from the CYP1, CYP2, and CYP3 gene families are responsible for most drug metabolism. Cytochrome P-450,Cytochrome P-450 Enzyme,Cytochrome P-450-Dependent Monooxygenase,P-450 Enzyme,P450 Enzyme,CYP450 Family,CYP450 Superfamily,Cytochrome P-450 Enzymes,Cytochrome P-450 Families,Cytochrome P-450 Monooxygenase,Cytochrome P-450 Oxygenase,Cytochrome P-450 Superfamily,Cytochrome P450,Cytochrome P450 Superfamily,Cytochrome p450 Families,P-450 Enzymes,P450 Enzymes,Cytochrome P 450,Cytochrome P 450 Dependent Monooxygenase,Cytochrome P 450 Enzyme,Cytochrome P 450 Enzyme System,Cytochrome P 450 Enzymes,Cytochrome P 450 Families,Cytochrome P 450 Monooxygenase,Cytochrome P 450 Oxygenase,Cytochrome P 450 Superfamily,Enzyme, Cytochrome P-450,Enzyme, P-450,Enzyme, P450,Enzymes, Cytochrome P-450,Enzymes, P-450,Enzymes, P450,Monooxygenase, Cytochrome P-450,Monooxygenase, Cytochrome P-450-Dependent,P 450 Enzyme,P 450 Enzymes,P-450 Enzyme, Cytochrome,P-450 Enzymes, Cytochrome,Superfamily, CYP450,Superfamily, Cytochrome P-450,Superfamily, Cytochrome P450
D005615 Freezing Liquids transforming into solids by the removal of heat. Melting

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