DRUG-METABOLIZING ENZYME-ACTIVITIES AND SUPEROXIDE FORMATION IN PRIMARY AND IMMORTALIZED RAT-BRAIN ENDOTHELIAL-CELLS

The activities of several enzymes involved in drug metabolism, NADPH-c ytochrome P450 reductase, cytochrome P450 isoforms CYP1A and CYP2B, an d uridine diphosphate glucuronosyltransferase (UGT) have been measured in primary cultures of rat cerebrovascular endothelial cells and in t he immortalized rat brain endothelial cell line RBE4. These drug metab olizing activities were similar in the microsomes prepared from both c ell types, even after 20 passages for RBE4 cells. These results were c onfirmed by Western immunoblotting analysis, using polyclonal antibodi es raised against rat liver enzymes. The superoxide production observe d during NADPH-cytochrome P450 reductase-dependent monoelectronic redu ction of four xenobiotics, menadione, anthraquinone, nitrofurazone and diquat, was also investigated in these cultured cells at confluence. The rates of radical production were concentration-dependent. The supe roxide formation induced by quinone metabolism was comparable in both cell cultures, and amounts of superoxide radicals were produced even a fter 20 passages of RBE4 cells. On the other hand, nitrofurazone and d iquat metabolism produced weak amounts of superoxide radicals in both cell types. Taken together, these results suggest that RBE4 cell line seems to constitute a valuable in vitro model for studies on the activ ity of some enzymatic systems involved in drug metabolism at the blood -brain barrier and the functional consequences of their activity.