Cloning and sequencing of some relevant drug metabolism genes in cattle: the Programma Galileo 2003 summary report

Objective. Liver biotransformations play a critical role in the individual response to harmful substances and increasing importance has been recently given to drug metabolising enzymes (DMEs) gene expression analysis. In cattle, little is known about the expression of DMEs and their transcription factors (i.e. nuclear receptors, NRs). Therefore, some relevant bovine liver DMEs and NRs genes have been cloned and sequenced. Method. Liver samples from control or phenobarbital-induced cattle were snap-frozen in liquid nitrogen and stored at –80°C. Total mRNA was extracted, purified and quantified. Specific primer pairs were designed from highly conserved regions among different mammalian species. Single band PCR products were cloned and plasmids containing the expected PCR fragment submitted to confirmatory enzyme restriction analysis and resolution on agarose gels. Therefore, they were amplified, purified and sequenced. Results. Eleven cattle genes (CAR, CYP1A1, CYP2B, CYP2C, CYP4A, GRα, HNF4α, PgP, PXR, RXRα and UDPGT) were submitted to GenBank. Some of them were then successfully used for liver gene expression analysis (northern blotting) in cattle treated, in vivo, with corticosteroids alone or in combination with steroids and β-agonists. Discussion. Cattle are economically important worldwide, but little is known about the expression of liver DMEs and NRs. Their molecular characterization is important for understanding mechanisms of detoxification, or bioactivation, in basic, pharmacological and environmental research. Particularly, these genes might be used in the future to define some biomarkers of effect or dose response in cattle administered with illicit drugs.