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: 3.94).
04/2012; 94(3):232-41. DOI: 10.1016/j.antiviral.2012.04.003
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.
Available from: Darren Ferrell
- "Brain oligomeric  and Aβ1-40,42
 have been correlated with cognitive impairment. Since the EFV containing regimen may promote mitochondrial dysfunction , , , ,  which could result in increased BACE-1 activity, we investigated the effect of a commonly used EFV containing cART regimen – for its ability to upregulate Aβ production via activation of BACE-1 and amyloidogenic APP processing and also for its ability to reduce microglial phagocytosis of Aβ1-40,42. "
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ABSTRACT: Efavirenz (EFV) is among the most commonly used antiretroviral drugs globally, causes neurological symptoms that interfere with adherence and reduce tolerability, and may have central nervous system (CNS) effects that contribute in part to HIV associated neurocognitive disorders (HAND) in patients on combination antiretroviral therapy (cART). Thus we evaluated a commonly used EFV containing regimen: EFV/zidovudine (AZT)/lamivudine (3TC) in murine N2a cells transfected with the human "Swedish" mutant form of amyloid precursor protein (SweAPP N2a cells) to assess for promotion of amyloid-beta (Aβ) production. Treatment with EFV or the EFV containing regimen generated significantly increased soluble amyloid beta (Aβ), and promoted increased β-secretase-1 (BACE-1) expression while 3TC, AZT, or, vehicle control did not significantly alter these endpoints. Further, EFV or the EFV containing regimen promoted significantly more mitochondrial stress in SweAPP N2a cells as compared to 3TC, AZT, or vehicle control. We next tested the EFV containing regimen in Aβ - producing Tg2576 mice combined or singly using clinically relevant doses. EFV or the EFV containing regimen promoted significantly more BACE-1 expression and soluble Aβ generation while 3TC, AZT, or vehicle control did not. Finally, microglial Aβ phagocytosis was significantly reduced by EFV or the EFV containing regimen but not by AZT, 3TC, or vehicle control alone. These data suggest the majority of Aβ promoting effects of this cART regimen are dependent upon EFV as it promotes both increased production, and decreased clearance of Aβ peptide.
- "This result corresponds to the findings of Apostolova et al. , in which they reported that EFV at similar concentrations as ours failed to show significant cytotoxic effects in human Hep3B cells following a 3-day treatment period. A follow-up study by the same team in 2012 , revealed that EFV at similar concentrations did not alter the mRNA expression of Bax and BCL-2-like 1 genes following a 24 h treatment period in human Hep3B cells. This report is consistent with our findings that EFV did not alter the mRNA expression of both cervical cell lines after a 4-day exposure. "
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ABSTRACT: Protease inhibitors (PIs) and reverse transcriptase drugs are important components of highly active antiretroviral therapy (HAART) for treating human acquired immunodeficiency syndrome (AIDS). Long-term clinical therapeutic efficacy and treatment compliance of these agents have been limited by undesirable adverse effects and their oncogenicity has been queried. This study investigated the effects of selected antiretroviral agents on the expression of key apoptotic regulatory genes; Bax and Bcl-2 in two cervical cell lines HCS-2 and NCE16IIA by real-time qPCR gene expression and immunocytochemistry. The anti-apoptotic effects of the PI-LPV/r were investigated by cell death detection ELISA and acridine orange staining. All the antiretroviral drugs and combinations tested had no effects on Bax and Bcl-2 gene expression and protein localisation in both cell lines. The protease inhibitors-LPV/r exhibited significant (P<0.05) inhibition of camptothecin-induced apoptosis in the cervical cancer HCS-2 cell line but not in the normal immortalised NCE16IIA cell line. This anti-apoptotic property of HIV protease inhibitors, although shown so far not to involve protein and RNA synthesis might promote the development of cancer.
Available from: Fernando Alegre
- "This interference is manifested in reduced respiration with specific inhibition of Complex I, decreased mitochondrial membrane potential (DW m ), bioenergetic changes and elevated ROS generation   , culminating in the adaptive response of autophagy . Parallel to these manifestations, several genes implicated in ER stress/UPR are upregulated, namely DDIT3/CHOP (C/EBPhomologous protein, also known as DNA damage inducible gene 153 (GADD153)), DNAJB4 (DnaJ (Hsp40) homolog, subfamily B, member 4), HSPA6 (Heat shock 70 kDa protein 6 (HSP70B')), and HSPD1 (Heat shock 60 kDa protein 1 (chaperonin 60) . "
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ABSTRACT: 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 EFV in human hepatic cells. This study assesses the induction of ER-stress by EFV in the same model and the implication of mitochondria in this process.
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.
ER-stress markers, including CHOP and GRP78 expression (both protein and mRNA), phosphorylation of eIF2α and presence of the spliced form of XBP1 were up-regulated. 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.
This newly discovered mechanism of cellular insult involving ER-stress and UPR response may help to 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 EFV observed expands our knowledge of the mechanisms that trigger ER-stress and throw light on the mitochondria/ER interplay in drug-induced hepatic challenge.
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