Application of HC-AFW1 hepatocarcinoma cells for mechanistic studies: Regulation of cytochrome P450 2B6 expression by dimethyl sulfoxide and EGR1

Various exogenous compounds, for example the drugs bupropione and propofol, but also various cytostatics, are metabolized in the liver by the enzyme cytochrome P450 (CYP) 2B6. Transcription from the CYP2B6 gene is mainly regulated via the transcription factors CAR (constitutive androstane receptor) and PXR (pregnane-X-receptor). Most hepatic cell lines express no or only low levels of CYP2B6 due to a loss of these two regulators. Dimethyl sulfoxide (DMSO) is frequently used in liver cell cultivation and is thought to affect the expression of various CYP isoforms by inducing or preserving cellular differentiation. Here, we have studied the effects of up to 1.5% of DMSO as cell culture medium supplement on CYP expression in hepatocarcinoma cells from line HC-AFW1. DMSO did not induce differentiation of the HC-AFW1 cell line, as demonstrated by unaltered levels of selected mRNA markers important for hepatocyte differentiation, and also by the lack of a DMSO effect on a broader spectrum of CYPs. By contrast, CYP2B6 mRNA was strongly induced by DMSO. This process was independent of CAR or PXR activation. Interestingly, elevated transcription of CYP2B6 was accompanied by a simultaneous induction of EGR1 (early growth response 1), a transcription factor which is known to influence the expression of CYP2B6. Expression of wild-type EGR1 or of a truncated, dominant-negative EGR1 mutant was able to mimic or attenuate the DMSO effect, respectively. These findings demonstrate that EGR1 is involved in the regulation of CYP2B6 by DMSO in HC-AFW1 cells.

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