A mechanistic insight into MDMA-mediated hepatotoxicity

Abstract

methylenedioxymethamphetamine (MDMA, Ecstasy) is a popular drug of abuse among young people that can induce adverse effects. However, these effects lack a specific pattern, as consumption quantities are not correlated with the initiation and severity of the injury. MDMA can cause drug-induced liver injury (DILI). Two suggested pathways that play a role in the onset of DILI are direct hepatotoxicity due to toxic metabolites and adverse immune responses. Therefore, we studied MDMA interactions with phase I and II enzymes and the possible alteration of immune events in several in vitro systems (human liver (THLE) cells transfected with individual CYP450, rat liver microsomes, human PXR-mediated CYP3A4-reporter gene assay, rat primary hepatocytes, HepG2, THP-1 and PBMC). Our data suggests that not only CYP2D6 but also CYP3A4 plays an important role in MDMA bioactivation. Furthermore,MDMA alone or in combination with other therapeutic drugs inhibited CYP3A catalytic activity. This appeared to be due to decreased activation of its main regulator pregnane X receptor (PXR) which subsequently decreases CYP3A gene expression. Therefore, MDMA use in combination with other (therapeutic) drugs could induce adverse drug-drug interactions through interactions with PXR and/or CYP3A. Following metabolism by phase II enzymes, interactions between MDMA/HHMA (a toxic MDMA metabolite) and the glutathione system were observed. HHMA significantly decreased cell viability and depleted GSH levels, resulting in an increased expression of glutamate cysteine ligase catalytic subunit (GCLC) and glutathione-S-transferase (GST). Upon MDMA exposure GSH levels and GCLC expression were not significantly affected, although GST expression was increased. Moreover, we evaluated the potential protective effects of two antioxidants, N-acetyl-cysteine (NAC) and sulforaphane (SFN). NAC counteracted MDMA-induced cytotoxicity and restored GSH levels. Phase II enzyme expression was also reverted. Conversely, SFN increased MDMA-induced cytotoxicity, GSH depletion, GCLC and GST expression. The differential behavior of both antioxidants indicates that care should be taken when using them during treatment of MDMA intoxications, since NAC could very rapidly restore GSH levels, while SFN lacks this effect.Finally, we investigated the role of the immune system on MDMA-mediated liver injury using liver and immune cells in a co-culture model. A protection (HepG2/THP-1) versus impairment (HepG2/PBMC) of MDMA-induced HepG2 cell viability was observed. In THP-1 cells, TNF‑α expression was moderately decreased and IL-8 expression was increased. In the HepG2/PBMC model, MDMA exposure reduced TNF-α and IL-8 but increased IL-10 gene expression, comparable with effects observed in MDMA users. These data suggests that the HepG2/PBMC model is useful to study drug-induced liver toxicity. Using this model, we have shown that immunosuppression appeared to be partially mediated by antagonistic effects of MDMA on peroxisome proliferator activated receptor alpha (PPARα) in HepG2 cells. Taken together, MDMA interactions with metabolizing enzymes and the nuclear receptors that regulate them may alter the metabolic pattern of individuals. This should be taken into account when searching for a clinical treatment to counteract MDMA-mediated side effects. Furthermore, MDMA may cause a greater susceptibility to infectious diseases through a reduction of pro-inflammatory events.