Quantitative RT-PCR Analysis of CAR Targets

RT-qPCR was used to examine CAR target gene
expression in PHH, HepaRG cells, or in mouse liver samples. Total
RNA, reverse transcription, and qPCR were performed, and mRNA expression
data was analyzed as we have described before.^{51 (link)}All RT-qPCR experiments were performed in triplicate
samples, and data are presented as fold induction to vehicle-treated
cells with the same reagents we have described before.^{48 (link),52 (link)} PCR TaqMan probes for murine genes have been listed in our previous
reports.^{52 (link)} TaqMan probes for CYP3A4, CYP2C9, and CYP2B6 human genes
as well as for reference genes B2M and GADPH genes were obtained from Thermo Fisher: CYP3A4 (Hs00604506_m1), CYP2C9 (Hs02383631_s1), CYP2B6 (Hs04183483_g1), GAPDH (Hs02758991_g1), and B2M (Hs00984230_m1).

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Mejdrová I., Dušek J., Škach K., Stefela A., Skoda J., Chalupský K., Dohnalová K., Pavkova I., Kronenberger T., Rashidian A., Smutná L., Duchoslav V., Smutny T., Pávek P, & Nencka R. (2023). Discovery of Novel Human Constitutive Androstane Receptor Agonists with the Imidazo[1,2-a]pyridine Structure. Journal of Medicinal Chemistry, 66(4), 2422-2456.

Publication 2023

CYP2C9 CYP2B6

Cells Cyp3a4 Gapdh Genes Liver Mrna Murine Reverse transcription

Corresponding Organization : Charles University

Other organizations : Czech Academy of Sciences, Institute of Molecular Genetics, University of Defence, University of Eastern Finland, University of Tübingen, University Children's Hospital Tübingen

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Variable analysis

independent variables

Treatment conditions (PHH, HepaRG cells, or mouse liver samples)

dependent variables

Expression of CAR target genes
MRNA expression data

control variables

Total RNA extraction, reverse transcription, and qPCR procedures
Reagents used in experiments, as described in previous reports
TaqMan probes for murine genes, CYP3A4, CYP2C9, CYP2B6, B2M, and GAPDH genes

controls

Vehicle-treated cells as a control condition

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