Introduction Actually, the obtainment of cattle hepatocyte primary cultures to be used for drug metabolism studies is still troublesome; therefore, a reliable hepatic in vitro model is needed for cattle. To this purpose, we preliminarly characterised a novel bovine foetal hepatocyte-derived (BFH12) cell line, measuring gene expression profiles of principal drug metabolizing enzymes (DMEs) and drug transporters (DTs) either in control conditions than following the incubation with common DMEs inducers. Materials and methods After the definition of the best conditions for cell culturing, BFH12 cells at different passages were seeded and cultivated either at standard conditions or in presence of some prototypical DMEs inducers, i.e., β-naphthoflavone (βNF), PCB126, phenobarbital, CITCO, FL81, dexamethasone, rifampicin, pregnenolone 16α-carbonitrile (PCN), SR12813 and RU486. Total RNA was extracted, and mRNA levels of main cytochromes P450 (CYPs), glucuronosyl- and glutathione-transferase (UGT and GST, respectively) as well as of nuclear receptors, i.e. aryl hydrocarbon receptor (AhR), pregnane X receptor (PXR), constitutive androstan receptor (CAR), retinoid X receptor alpha (RXRα), and hepatocyte nuclear factor (HNF) 1α, 1β and 4α were measured by using quantitative Real time PCR (qPCR) assays. Results and conclusions BFH12 cells expressed detectable amounts of AhR, PXR, CAR, RXRα, HNF-1α, -1β and 4α mRNAs as well as main mixed function oxidases, i.e. CYP1A1, 1A2, 1B1, 2B22, 3A28, 3A38, 3A48. Also conjugative DMEs, namely UGT and GST, were constitutively expressed. Among the tested inducers, best results in terms of mRNA up-regulation were observed with PCB126 and βNF (CYP1A), FL81 (CYP2B), and RU486 (CYP3A). This preliminary study confirms BFH12 cell line as a promising in vitro method for studying drug metabolism in cattle. Nevertheless, prospective studies are needed to better characterize BFH12 as a useful in vitro model for regulation, induction and functional studies. Acknowledgements Project supported by University of Padua (DOR173229).

Preliminary Characterisation Of a Novel Cattle Foetal Hepatocyte-Derived Cell Line (BFH12)

Mery Giantin;Roberta Tolosi;Mauro Dacasto
2018

Abstract

Introduction Actually, the obtainment of cattle hepatocyte primary cultures to be used for drug metabolism studies is still troublesome; therefore, a reliable hepatic in vitro model is needed for cattle. To this purpose, we preliminarly characterised a novel bovine foetal hepatocyte-derived (BFH12) cell line, measuring gene expression profiles of principal drug metabolizing enzymes (DMEs) and drug transporters (DTs) either in control conditions than following the incubation with common DMEs inducers. Materials and methods After the definition of the best conditions for cell culturing, BFH12 cells at different passages were seeded and cultivated either at standard conditions or in presence of some prototypical DMEs inducers, i.e., β-naphthoflavone (βNF), PCB126, phenobarbital, CITCO, FL81, dexamethasone, rifampicin, pregnenolone 16α-carbonitrile (PCN), SR12813 and RU486. Total RNA was extracted, and mRNA levels of main cytochromes P450 (CYPs), glucuronosyl- and glutathione-transferase (UGT and GST, respectively) as well as of nuclear receptors, i.e. aryl hydrocarbon receptor (AhR), pregnane X receptor (PXR), constitutive androstan receptor (CAR), retinoid X receptor alpha (RXRα), and hepatocyte nuclear factor (HNF) 1α, 1β and 4α were measured by using quantitative Real time PCR (qPCR) assays. Results and conclusions BFH12 cells expressed detectable amounts of AhR, PXR, CAR, RXRα, HNF-1α, -1β and 4α mRNAs as well as main mixed function oxidases, i.e. CYP1A1, 1A2, 1B1, 2B22, 3A28, 3A38, 3A48. Also conjugative DMEs, namely UGT and GST, were constitutively expressed. Among the tested inducers, best results in terms of mRNA up-regulation were observed with PCB126 and βNF (CYP1A), FL81 (CYP2B), and RU486 (CYP3A). This preliminary study confirms BFH12 cell line as a promising in vitro method for studying drug metabolism in cattle. Nevertheless, prospective studies are needed to better characterize BFH12 as a useful in vitro model for regulation, induction and functional studies. Acknowledgements Project supported by University of Padua (DOR173229).
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11577/3271870
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