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

Departamento de Farmacología, Facultad de Medicina, Universitat de València, Valencia, Spain.
Antiviral research (Impact Factor: 3.43). 04/2012; 94(3):232-41. DOI: 10.1016/j.antiviral.2012.04.003
Source: PubMed

ABSTRACT 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.

1 Follower
  • [Show abstract] [Hide abstract]
    ABSTRACT: Epidermal keratinocytes serve as the primary barrier between the body and environmental stressors. They are subjected to numerous stress events and are likely to respond with a repertoire of heat shock proteins (HSPs). HSPA6 (HSP70B') is described in other cell types with characteristically low to undetectable basal expression, but is highly stress induced. Despite this response in other cells, little is known about its control in keratinocytes. We examined endogenous human keratinocyte HSPA6 expression and localized some responsible transcription factor sites in a cloned HSPA6 3 kb promoter. Using promoter 5' truncations and deletions, negative and positive regulatory regions were found throughout the 3 kb promoter. A region between -346 and -217 bp was found to be crucial to HSPA6 basal expression and stress inducibility. Site-specific mutations and DNA-binding studies show that a previously uncharacterized AP1 site contributes to the basal expression and maximal stress induction of HSPA6. Additionally, a new heat shock element (HSE) within this region was defined. While this element mediates increased transcriptional response in thermally stressed HaCaT keratinocytes, it preferentially binds a stress-inducible factor other than heat shock factor (HSF)1 or HSF2. Intriguingly, this newly characterized HSPA6 HSE competes HSF1 binding a consensus HSE and binds both HSF1 and HSF2 from other epithelial cells. Taken together, our results demonstrate that the HSPA6 promoter contains essential negative and positive promoter regions and newly identified transcription factor targets, which are key to the basal and stress-inducible expression of HSPA6. Furthermore, these results suggest that an HSF-like factor may preferentially bind this newly identified HSPA6 HSE in HaCaT cells.
    Cell Stress and Chaperones 07/2014; 20(1). DOI:10.1007/s12192-014-0529-0 · 2.54 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Background and PurposeThe non-nucleoside analog reverse transcriptase inhibitor Efavirenz (EFV) has been associated with hepatic toxicity and metabolic disturbances. Although the mechanisms involved are not clear, recent evidence has pinpointed a specific mitochondrial action of EFV accompanied by ER-stress/UPR induction in human hepatic cells. The aim of this study was to further investigate the implication of this organelle by evaluating EFV action on cells lacking functional mitochondria (rho°), and comparing it with that of the typical mitotoxic agent, rotenone (Rot), a standard Complex I inhibitor, and the ER-stress inductor thapsigargin (TG).Experimental approachHep3B rho+ and rho° cells were treated with clinically relevant concentrations of EFV, then mitochondrial function and cytotoxicity were studied using standard cell biology techniques.Key resultsEFV-treated rho° cells exhibited a substantial reduction in parameters indicative of mitochondrial interference, such as increased superoxide production, mitochondrial mass/morphology alterations and enhanced expression of LONP, a highly conserved mitochondrial protease. In line with these results, the cytotoxic effect (cell number, chromatin condensation, cell cycle alterations and induction of apoptosis) of EFV was less pronounced in Hep3B respiration-depleted cells than in WT cells. The response of EFV reveals both similarities and differences with those of two distinct mitochondrial stressors, TG and Rot.Conclusions and ImplicationsCells lacking normal mitochondria (rho°) are less vulnerable to EFV. Our results provide further evidence that the hepatic damage induced by EFV involves acute interference with mitochondria and extend our knowledge of the mitochondria/ER interplay that takes place upon a stress stimulus.
    British Journal of Pharmacology 11/2014; 172(7). DOI:10.1111/bph.13018 · 4.99 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: The non-nucleoside reverse transcriptase inhibitor efavirenz is a widely prescribed antiretroviral drug used in combined antiretroviral therapy. Despite being an essential and life-saving medication, the required lifelong use of HIV drugs has been associated with a variety of adverse effects, including disturbances in lipid metabolism and increased cardiovascular risk. Efavirenz belongs to those HIV drugs for which cardiovascular and endothelial dysfunctions have been reported. It is here shown that elevated concentrations of efavirenz can inhibit endothelial meshwork formation on extracellular matrix gels by normal and immortalized human umbilical vein cells. This inhibition was associated with an increase in oxidative stress markers, endoplasmic reticulum (ER) stress markers, and autophagy. Induction of ER stress occurred at pharmacologically relevant concentrations of efavirenz and resulted in reduced proliferation and cell viability of endothelial cells, which worsened in the presence of elevated efavirenz concentrations. In combination with the HIV protease inhibitor nelfinavir, both oxidative stress and ER stress became elevated in endothelial cells. These data indicate that pharmacologically relevant concentrations of efavirenz can impair cell viability of endothelial cells and that these effects may be aggravated by either elevated concentrations of efavirenz or by a combined use of efavirenz with other oxidative stress-inducing medications.
    Cardiovascular Toxicology 02/2015; DOI:10.1007/s12012-015-9314-2 · 2.06 Impact Factor