Metabolic activity is unstable in primary hepatocyte cultures and is influenced by the matrix composition.The effect of incorporating 20% chondroitin-6-sulphate(Ch6SO4), a glycosaminoglycan, into collagen films and gels(0.3%w/v), and crosslinking the films and gels with 1,6-diaminohexane(DAH)on the glucuronidation of kaempferol by primary rat hepatocytes cultured for 48h and 7 days was investigated. Hepatocytes isolated from male Sprague-Dawley rats by collagenase perfusion were cultured (3x106/60mm Petri dish)in Chee's medium with 5% v/v foetal calf serum. Cells were incubated with 100mM kaempferol for 1h at 378C and the glucuronides(K1 and K2) were measured by HPLC. Cells cultured on collagen films formed only the K2 metabolite after 48h in culture. The addition of Ch6SO4 or DAH significantly increased the formation of this glucuronide. However, cells seeded on gels showed no metabolism after 48h in culture. By 7 days in culture, both K1 and K2 glucuronides were formed in cells on all the different films and gels. The formation of K1 glucuronide was significantly higher with the addition of Ch6SO4 to the films. K2 glucuronide was significantly higher in all of the crosslinked films compared to the plain films. Modification of collagen based substrates may improve cultured hepatocyte phenotype.
|Journal||Biochemical Society Transactions|
|Publication status||Published - 2002|
- hepatocyte cultures
- intracellular reduced glutathione
- confocal laser scanning microscopy
- InGaN laser source
- cultured hepatocytes