ER stress in human hepatic cells treated with Efavirenz: mitochondria again.

BACKGROUND & AIMS: ER stress is associated with a growing number of liver diseases, including drug-induced hepatotoxicity. The non-nucleoside analogue reverse transcriptase inhibitor Efavirenz, a cornerstone of the multidrug strategy employed to treat HIV1 infection, has been related to the development of various adverse events, including metabolic disturbances and hepatic toxicity, the mechanisms of which remain elusive. Recent evidence has pinpointed a specific mitochondrial effect of Efavirenz in human hepatic cells. This study assesses the induction of ER stress by Efavirenz in the same model and the implication of mitochondria in this process. METHODS: Primary human hepatocytes and Hep3B were treated with clinically relevant concentrations of Efavirenz and parameters of ER stress were studied using standard cell biology techniques. RESULTS: ER stress markers, including CHOP and GRP78 expression (both protein and mRNA), phosphorylation of eIF2α, and presence of the spliced form of XBP1 were upregulated. Efavirenz also enhanced cytosolic Ca(2+) content and induced morphological changes in the ER suggestive of ER stress. This response was greatly attenuated in cells with altered mitochondrial function (Rho°). The effects of Efavirenz on the ER, and particularly in regard to the mitochondrial involvement, differed from those elicited by a standard pharmacological ER stressor. CONCLUSIONS: This newly discovered mechanism of cellular insult involving ER stress and UPR response may help comprehend the hepatic toxicity that has been associated with the widespread and life-long use of Efavirenz. In addition, the specificity of the actions of Efavirenz observed expands our knowledge of the mechanisms that trigger ER stress and shed some light on the mitochondria/ER interplay in drug-induced hepatic challenge.

Profile of stress and toxicity gene expression in human hepatic cells treated with Efavirenz.

Hepatic toxicity and metabolic disorders are major adverse effects elicited during the pharmacological treatment of the human immunodeficiency virus (HIV) infection. Efavirenz (EFV), the most widely used non-nucleoside reverse transcriptase inhibitor (NNRTI), has been associated with these events, with recent studies implicating it in stress responses involving mitochondrial dysfunction and oxidative stress in human hepatic cells. To expand these findings, we analyzed the influence of EFV on the expression profile of selected stress and toxicity genes in these cells. Significant up-regulation was observed with Cytochrome P450, family 1, subfamily A, polypeptide 1 (CYP1A1), which indicated metabolic stress. Several genes directly related to oxidative stress and damage exhibited increased expression, including Methalothionein 2A (MT2A), Heat shock 70kDa protein 6 (HSPA6), Growth differentiation factor 15 (GDF15) and DNA-damage-inducible transcript 3 (DDIT3). In addition, Early growth response protein 1 (EGR1) was enhanced, whereas mRNA levels of the inflammatory genes Chemokine (C-X-C motif) ligand 10 (CXCL10) and Serpin peptidase inhibitor (nexin, plasminogen activator inhibitor type 1), member 1 (SERPINE1) decreased and increased, respectively. This profile of gene expression supports previous data demonstrating altered mitochondrial function and presence of oxidative stress/damage in EFV-treated hepatic cells, and may be of relevance in the search for molecular targets with therapeutic potential to be employed in the prevention, diagnosis and treatment of the hepatic toxicity associated with HIV therapy.

Autophagy as a rescue mechanism in efavirenz-induced mitochondrial dysfunction: a lesson from hepatic cells.

Efavirenz (EFV) is the most widely used non-nucleoside reverse transcriptase inhibitor applied in highly active antiretroviral therapy (HAART), the combined pharmacological treatment of the human immunodeficiency virus infection. Its use has been associated with the development of several adverse events including hepatotoxicity. The molecular pathogenesis of this effect is poorly understood but recent reports have highlighted features of mitochondrial dysfunction in hepatic cells exposed to clinically relevant concentrations of EFV. In this study, we investigated the activation of autophagy and, in particular, mitophagy, in human hepatic cells exposed to EFV. We detected the presence of altered mitochondria with abnormal morphology and relative increase in mitochondrial mass. Several autophagic markers reveal specific induction of autophagy. Of special note, while moderate levels of EFV activate autophagy, higher concentrations exceeding the threshold of mitochondrial dysfunction, lead to a blockage in the autophagic flux, thus promoting "autophagic stress". Pharmacological inhibition of autophagy exacerbates the deleterious effect of EFV on cell survival/proliferation thereby promoting apoptosis, a finding which points to the fact that autophagy is triggered as a rescue mechanism enabling cell survival. The effect described in this study could be involved in the EFV-associated hepatotoxicity. It may constitute a new mechanism implicated in the genesis of pharmacological liver damage and in the recovery of hepatic homeostasis upon a drug-induced cellular insult.

Compromising mitochondrial function with the antiretroviral drug efavirenz induces cell survival-promoting autophagy.

UNLABELLED: Hepatotoxicity is a very common side effect associated with the pharmacological treatment of human immunodeficiency virus (HIV) infection and its pathogenesis is poorly understood. Efavirenz (EFV) is the most widely used nonnucleoside reverse transcriptase inhibitor administered for the control of HIV and some of its toxic effects in hepatic cells have been recently shown to display features of mitochondrial dysfunction. Here we studied the activation of autophagy and, in particular, mitophagy, the main mitochondrial turnover mechanism, in human hepatic cells treated with clinically relevant concentrations of this drug. EFV-treated cells had altered mitochondria, characterized by a relative increase in mitochondrial mass and defective morphology. This was followed by induction of autophagy as shown by the presence of autophagic vacuoles and the presence of the specific autophagic marker proteins microtubule-associated protein 1A/1B light chain 3 and Beclin-1. Importantly, whereas moderate levels of EFV activated autophagy, higher concentrations led to blockage in the autophagic flux, a condition that promotes "autophagic stress" and produces severe cellular damage. Finally, pharmacological inhibition of autophagy exacerbated the deleterious effect of EFV on cell survival/proliferation promoting apoptosis, which suggests that autophagy acts as an adaptive mechanism of cell survival. CONCLUSION: Clinical concentrations of EFV induce autophagy and, in particular, mitophagy in hepatic cells. Activation of this process promotes cell survival, but exceeding a certain threshold of mitochondrial dysfunction is associated with an autophagic overload or stress. This effect could be involved in the EFV-associated hepatotoxicity and may constitute a new mechanism implicated in the genesis of drug-induced liver damage.