EVIDENCE THAT THE LOSS OF RAT-LIVER CYTOCHROME-P450 INVITRO IS NOT SOLELY ASSOCIATED WITH THE USE OF COLLAGENASE, THE LOSS OF CELL-CELL CONTACTS AND OR THE ABSENCE OF AN EXTRACELLULAR-MATRIX

Two methods avoiding the widespread technique of collagenase perfusion have been employed to study the regulation of total cytochrome P450 content in rat hepatocyte culture. One technique required the perfusion of the liver with the chelating agent EDTA to dissociate the parenchymal cells prior to culture. Over a period of 48 hr, cultured hepatocytes isolated by EDTA perfusion showed comparable losses of cytochrome P450 as cells isolated by perfusion with collagenase. The second technique involved the culture of 210-240-mu-m thick "precision cut" liver slices. The results presented here indicate that the liver slices remain viable for 24 hr of culture, but that liver slices also lose their cytochrome P450 content at a comparable rate to collagenase prepared cells in culture. Collectively the results suggest that there is not a direct causal relationship between the loss of cytochrome P450 and one or a combination of the use of collagenase; the loss of cell-cell contacts and the absence of an extracellular matrix.