This study aimed to provide further insights into the mechanism of in vivo regulation of cytochrome P450 (CYP450) 1A, 2A and 2E1 activities in pigs with different levels of testicular steroids. Hepatic mRNA and protein expression and enzymatic activity of CYP1A, CYP2A and CYP2E1 were compared between entire male and castrated pigs. Castration was performed either surgically or immunologically. The pigs were divided into four groups. In the first group, piglets were surgically castrated without anaesthesia. Immunological castration was performed by vaccination with Improvac R (Pfizer Ltd). Vaccinated pigs were subdivided into two groups according to the vaccination regimen: early and standard vaccination. Pigs in the early vaccination group were vaccinated when aged 11 and 15 weeks. Pigs in the standard vaccination group were vaccinated when aged 17 and 21 weeks. In the control group, pigs remained intact throughout the study. Hepatic CYP450 mRNA expression, measured by real-time RT-PCR, differed significantly between groups for all isoforms measured: CYP1A2 (P 5 0.002), 2A (P 5 0.000) and 2E1 (P 5 0.002). Lower CYP450 mRNA in entire male pigs suggests suppression of CYP1A2, CYP2A and CYP2E1 by testicular steroids at the transcriptional level. However, this suppression was not always reflected in decreased protein expression and activities of these isoforms, suggesting that at least some CYP450s (e.g. CYP2E1) are regulated by a post-transcriptional mechanism.Keywords: male pigs, testicular steroids, liver, cytochrome P450 ImplicationsWe studied in vivo regulation of cytochrome P450 (CYP450) 1A, 2A and 2E1 activities in uncastrated male pigs with high concentrations of testicular steroids, and surgically and immunologically castrated pigs with low steroid concentrations. We confirmed an association between steroids and CYP450 expression. We suggest that this regulation occurs at the transcriptional level, as indicated by suppressed CYP450 mRNA in uncastrated pigs. This suppression was not always reflected in decreased activities, suggesting that at least some CYP450s (e.g. CYP2E1) are regulated by a post-transcriptional mechanism. These results contribute to the knowledge on porcine CYP450 and its in vivo regulation by testicular steroids.