Inhibition of cytochrome P450s enhances (+)-usnic acid cytotoxicity in primary cultured rat hepatocytes

Abstract

(+)-Usnic acid (UA) is consumed as a dietary supplement to promote weight loss; however, dietary supplements containing UA have been associated with clinical cases of severe liver injury. UA has been shown to be hepatotoxic in rats and is extensively metabolized by hepatic cytochrome P450s (CYPs); therefore, we examined if UA metabolism results in the formation of cytotoxic metabolites or if metabolism is a detoxification process in primary rat hepatocytes. When CYP activity was suppressed by the non-isoenzyme-selective inhibitor SKF-525A (20μM), or the CYP1A inhibitor alpha-naphthoflavone (10μM), or the CYP3A inhibitor ketoconazole (25μM), the cytotoxicity of UA at 3~6μM after 3~20h of exposure was significantly increased as measured by lactate dehydrogenase (LDH) leakage. At 2h after UA exposure, an earlier time point prior to LDH release, these CYP inhibitors potentiated UA-induced inhibition of cellular respiration as determined by the Clark type oxygen electrode. Cellular adenosine triphosphate (ATP) depletion by UA was also exacerbated by these CYP inhibitors. The CYP2B/2C inhibitor, ticlopidine at 20μM, showed no effects in parallel experiments. These data demonstrate that UA is bio-transformed to less toxic metabolites in rat primary hepatocytes, probably mainly by CYP1A and 3A, but not 2B/2C. Published 2013. This article is a U.S. Government work and is in the public domain in the USA. (+)-Usnic acid (UA) may cause hepatotoxicity. We examined if its metabolism plays a role in toxicity in primary rat hepatocytes. When cytochrome P450 (CYP) activity was suppressed by pan-CYP inhibitor SKF-525A, or CYP1A inhibitor alpha-naphthoflavone, or CYP3A inhibitor ketoconazole, UA induced inhibition of cellular respiration, adenosine triphosphate depletion and cytotoxicity was significantly increased. The CYP2B/2C inhibitor ticlopidine showed no effects. We suggest UA is bio-transformed to less toxic metabolites in rat hepatocytes, mainly by CYP1A and 3A, but not 2B/2C. © 2014 John Wiley & Sons, Ltd.