Introduction. Bovine CYP3A cluster (chromosome 25) comprises at least three CYP3A genes, i.e. CYP3A28, 3A38, and 3A48. Recently, thirteen exonic missense variants have been identified, and their potential functional impact in terms of catalytic activity towards marker substrates (testosterone, nifedipine) has been evaluated in a heterologous expression system (V79 cells). As genetic variants may significantly contribute to inter-individual variability in drug metabolism and disposition, the aim of the present study was to estimate the incidence of aforementioned CYP3A variants in a representative population of Piedmontese (PDM) and Limousine (LIM) cattle breed. Materials and Methods. Three hundred liver aliquots from PDM cattle, collected at the slaughterhouse, and 215 blood samples from LIM cattle sampled by official veterinarians, were used. DNA was extracted using a commercial kit, and each CYP3As individual genotype was determined using melting curve genotyping assays (FRET probes). Results and Conclusions. Heterologous expression studies proved that only 5 out of 13 missense exonic variants resulted in differences in CYP3A-dependent catalytic activity. The Rs384467435 variant (here identified with CYP3A28.7), already present in the Hereford breed database, was the only single nucleotide polymorphism (SNP) for which both heterozygous (HET; 18% and 24% for PDM and LIM, respectively) and homozygous mutated genotypes (MUT; ~4% in both breeds) were found. CYP3A28.11 variant was present only in heterozygosis, and more frequently in LIM cattle (13% vs 5%), while CYP3A28.10 mutation was never found in the two cattle populations. CYP3A38.11A appeared in heterozygosis in only one PDM individual out of 300. Finally, CYP3A48.8B SNP was evidenced in both heterozygosis (25%) and homozygosis (4%) in PDM breed, while in LIM the genotyping was unsuccessful owing to the presence of additional silent mutations in strict nearness. This study proves that CYP3A polymorphisms are present in cattle breeds, and these ones might result in differences either in drug kinetics than in the presence of harmful xenobiotic residues. Acknowledgements Project supported by MIUR (2009ZE5HJP) and Regione del Veneto (DGR 2080, 30.12.2015).
Cytochrome P450 3As (CYP3As) Genotyping In Piedmontese and Limousine Cattle Breeds
Mery GiantinConceptualization
;Roberta TolosiInvestigation
;Mauro Dacasto
Project Administration
2018
Abstract
Introduction. Bovine CYP3A cluster (chromosome 25) comprises at least three CYP3A genes, i.e. CYP3A28, 3A38, and 3A48. Recently, thirteen exonic missense variants have been identified, and their potential functional impact in terms of catalytic activity towards marker substrates (testosterone, nifedipine) has been evaluated in a heterologous expression system (V79 cells). As genetic variants may significantly contribute to inter-individual variability in drug metabolism and disposition, the aim of the present study was to estimate the incidence of aforementioned CYP3A variants in a representative population of Piedmontese (PDM) and Limousine (LIM) cattle breed. Materials and Methods. Three hundred liver aliquots from PDM cattle, collected at the slaughterhouse, and 215 blood samples from LIM cattle sampled by official veterinarians, were used. DNA was extracted using a commercial kit, and each CYP3As individual genotype was determined using melting curve genotyping assays (FRET probes). Results and Conclusions. Heterologous expression studies proved that only 5 out of 13 missense exonic variants resulted in differences in CYP3A-dependent catalytic activity. The Rs384467435 variant (here identified with CYP3A28.7), already present in the Hereford breed database, was the only single nucleotide polymorphism (SNP) for which both heterozygous (HET; 18% and 24% for PDM and LIM, respectively) and homozygous mutated genotypes (MUT; ~4% in both breeds) were found. CYP3A28.11 variant was present only in heterozygosis, and more frequently in LIM cattle (13% vs 5%), while CYP3A28.10 mutation was never found in the two cattle populations. CYP3A38.11A appeared in heterozygosis in only one PDM individual out of 300. Finally, CYP3A48.8B SNP was evidenced in both heterozygosis (25%) and homozygosis (4%) in PDM breed, while in LIM the genotyping was unsuccessful owing to the presence of additional silent mutations in strict nearness. This study proves that CYP3A polymorphisms are present in cattle breeds, and these ones might result in differences either in drug kinetics than in the presence of harmful xenobiotic residues. Acknowledgements Project supported by MIUR (2009ZE5HJP) and Regione del Veneto (DGR 2080, 30.12.2015).
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