Effects of flumequine on cytochrome P450 enzymes in allogynogenetic silver crucian carp, Carassius auratus gibelio

Abstract

The effects of flumequine on activities of several microsomal P450 monooxygenases of allogynogenetic silver crucian carp (Carassius auratus gibelio) were determined and the induction of fluemquine on CYP1A was further explored at the level of protein expression and mRNA transcription.After 24 h following a single intraperitoneal injection at a dose of 35 mg/kg,the ethoxyresorufin O-deethylase (EROD) activity in the liver of the Crucian carp was 54.33 pmol/(mg·min),which was significantly higher than that of the control carps [34.00 pmol/(mg·min)](P0.01).It was revealed that flumequine could significantly induce EROD activity(P0.01).However,the other P450 monooxygenases,such as erythromycin N-demethylase (ERND) [177.98 pmol/(mg·min)],aminopyrine N-demethylase [934.40 pmol/(mg·min)] and ethoxycoumarin O-deethylase (ECOD) [9.84pmol/(mg·min)]were not significantly affected by flumequine,comparing with these activities in control [140.90 pmol/(mg·min),850.71 pmol/(mg·min) and 8.93 pmol/(mg·min)],respectively.Except that the ERND activity in kidney tissue was higher than that in liver,the activities of APD,EROD and ECOD were the highest in liver tissue.An immunoblot analysis with rabbit anti-rat P4501A polyclonal antibody showed that CYP 1A protein expression in flumequine-treated carps was markedly higher than that in control carps,which was consistent with the trend in EROD activities.These findings suggest that P450 1A isoform,constitutively expressed in crucian carp,was particularly susceptible to activation by flumequine.Semiquantitative RT-PCR indicated that CYP1A mRNA was not affected by flumequine.In vitro assay,microsomes incubating with various concentrations of flumequine demonstrated that this fluoroquinolone antibiotic had no induction or inhibition of CYP1A.On the basis of the above,liver CYP1A induction of flumequine on carp occurs at the post-translation level,which may enhance protein stability.