Aim: 

Efavirenz (EFV)-containing therapies are related to several side effects including hepatotoxic events and chronic metabolic disorders; however, the possible cellular mechanisms underlying these effects have been the subject of little study. We set out to evaluate the in vitro effects of clinical (10-25 mM) and supraclinical (50 mM) concentrations of EFV in several human cellular models.

Methods: 

Cellular viability was assessed using the MTT assay. Apoptotic features, superoxide production and mitochondrial membrane potential (DY_m) were studied by fluorescence microscopy. Western blot was employed for protein analysis.

Results: 

Cellular viability assays performed after 24h culture in the presence of EFV revealed reduced viability in the human hepatoma cell line Hep3B, human cervix carcinoma cell line HeLa and primary Human Umbilical Vein Endothelial cells (HUVEC). This result was corroborated with 3-day-proliferation experiments in Hep3B cells- significant reduction was detected with 25 mM EFV, whereas cytotoxicity was observed with 50 mM. To evaluate the mechanisms responsible for this diminished cellular viability, we performed bivariate Annexin V/Propidium Iodide analysis and found that EFV-treated cells presented apoptotic features, accompanied by a dose-dependent translocation of the mitochondrial proapoptotic proteins cytochrome c, AIF and Smac/DIABLO. We also detected a dose-dependent activation of caspase-3 and –9 without activation of caspase-8, pointing to induction of the intrinsic (mitochondrial) apoptotic pathway. In addition, short-term exposure of both Hep3B and HeLa cells with EFV compromised mitochondrial function by reducing DY_m and affecting the mitochondrial redox status.

Conclusion: 

Clinical concentrations of EFV are mytotoxic and can activate apoptotic programs in common cellular models.