Characterization of testosterone metabolism and 7-hydroxycoumarin conjugation by rat and human liver slices after storage in liquid nitrogen for 1 h up to 6 months.
1. Slices of human and rat liver were cryopreserved in 18% dimethyl sulphoxide (DMSO) and subsequently stored in liquid nitrogen for periods up to as long as 6 months. After thawing, the metabolism of testosterone to hydroxylated products and conjugation of 7-hydroxycoumarin were investigated. 2. Rat liver slices stored in liquid nitrogen for 6 months exhibited rates of formation of 7alpha-, 6beta- 16alpha- and 2alpha-hydroxytestosterone, and of androstenedione that did not differ significantly from those observed with fresh slices. 3. No formation of 2alpha-hydroxytestosterone was detected with slices of human liver. However, in contrast with the rat, human slices produced 2beta-hydroxytestosterone. The rates of formation of 7alpha-, 6beta-, 16alpha- and 2beta-hydroxytestosterone and of androstenedione by human liver slices after 6 months of storage in liquid nitrogen were 82, 71, 236, 66 and 92%, respectively, of the corresponding rates by fresh slices. 4. The rates of sulphation and glucuronidation of 7-hydroxycoumarin by slices from rat liver were 97 and 119%, respectively, of the corresponding fresh values after 6 months of storage in liquid nitrogen. 5. 7-Hydroxycoumarin glucuronidation by human liver slices was 53% of the corresponding fresh values after 6 months of storage. However, human slices showed little or no capacity to conjugate 7-hydroxycoumarin with sulphate. 6. It was demonstrated that slices of both human and rat liver can be cryopreserved and stored in liquid nitrogen for at least 6 months without major changes in their rates of metabolism of testosterone to its hydroxylated products and of 7-hydroxycoumarin conjugation. These findings further emphasize that cryopreservation of liver slices can be an effective tool in the use of biological material of limited availability.
Granhall
C
C
Research DMPK, AstraZeneca R&D Södertälje, S-151 85 Södertälje, Sweden.
Floby
E
E
Nordmark
A
A
Orzechowski
A
A
Thörne
A
A
Tybring
G
G
Sohlenius-Sternbeck
A-K
AK
D000735:Androstenedione / Q000378:metabolism; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D004396:Coloring Agents / Q000494:pharmacology; D015925:Cryopreservation / Q000379:methods*; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D005615:Freezing; D006801:Humans; D008099:Liver / Q000187:drug effects* / Q000378:metabolism*; D008297:Male; D009584:Nitrogen / Q000494:pharmacology; D010078:Oxazines; D051381:Rats; D017207:Rats, Sprague-Dawley; D013048:Specimen Handling; D013739:Testosterone / Q000378:metabolism*; D013997:Time Factors; D014468:Umbelliferones / Q000378:metabolism*; D014966:Xanthenes
D004396:Coloring Agents; D010078:Oxazines; D014468:Umbelliferones; D014966:Xanthenes; C005843:resazurin; D013739:Testosterone; D000735:Androstenedione; C031477:7-hydroxycoumarin; D009584:Nitrogen; D004121:Dimethyl Sulfoxide