title: Evaluation of seven drug metabolisms and clearances by cryopreserved human primary hepatocytes cultivated in microfluidic biochips.
abstract: We present characterization of the metabolic performance of human cryopreserved hepatocytes cultivated in a platform of parallelized microfluidic biochips. The RTqPCR analysis revealed that the mRNA levels of the cytochromes P450 (CYP 1A2, 2B6, 2C8, 2C9, 2C19, 2D6, 2E1, 3A4) were reduced after the adhesion period (when compared to the post-thawing step). The microfluidic perfusion played a part in stabilizing and partially recovering the levels of the HNF4α, PXR, OAPT2, CYP 1A2, 2B6, 2C19 and 3A4 mRNA on contrary to non-perfused cultures. Fluorescein diacetate staining and P-gp mRNA level illustrated the hepatocytes' polarity in the biochips. Drug metabolism was assessed using midazolam, tolbutamide, caffeine, omeprazole, dextromethorphan, acetaminophen and repaglinide as probes. Metabolite detection and quantification revealed that CYP1A2 (via the detection of paraxanthine), CYP3A4 (via 1-OH-midazolam, and omeprazole sulfone detection), CYP2C8 (via hydroxyl-repaglinide detection), CYP2C19 (via hydroxy-omeprazole detection) and CYP2D6 (via dextrorphan detection) were functional in our microfluidic configurations. Furthermore, the RTqPCR analysis showed that the drugs acted as inductors leading to overexpression of mRNA levels when compared to post-thawing values (such as for HNF4α, PXR and CYP3A4 by dextromethorpahn and omeprazole). Finally, intrinsic in vitro biochip clearances were extracted using a PBPK model for predictions. The biochip predictions were compared to literature in vitro data and in vivo situations.
authors: Laboratoire de Biomécanique et Bio ingénierie, CNRS UMR 7338, Université de Technologie de Compiègne, France.
authors: Régis
authors: R
authors: Baudoin
authors: Jean Matthieu
authors: JM
authors: Prot
authors: Grégory
authors: G
authors: Nicolas
authors: Jessy
authors: J
authors: Brocheton
authors: Céline
authors: C
authors: Brochot
authors: Cécile
authors: C
authors: Legallais
authors: Henri
authors: H
authors: Benech
authors: Eric
authors: E
authors: Leclerc
mesh_terms: D002454:Cell Differentiation; D002478:Cells, Cultured; D022781:Hepatocytes / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D008658:Inactivation, Metabolic; D007700:Kinetics; D008657:Metabolic Clearance Rate; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation; D015262:Xenobiotics / Q000378:metabolism*
chemical_list: D015262:Xenobiotics