Establishment of HK-2 Cells as a Relevant Model to Study Tenofovir-Induced Cytotoxicity

Abstract

Tenofovir (TFV) is an antiviral drug approved for treating Human Immunodeficiency Virus (HIV) and Hepatitis B. TFV is administered orally as the prodrug tenofovir disoproxil fumarate (TDF) which then is deesterified to the active drug TFV. TFV induces nephrotoxicity characterized by renal failure and Fanconi Syndrome. The mechanism of this toxicity remains unknown due to limited experimental models. This study investigated the cellular mechanism of cytotoxicity using a human renal proximal tubular epithelial cell line (HK-2). HK-2 cells were grown for 48 h followed by 24 to 72 h exposure to 0–28.8 μM TFV or vehicle, phosphate buffered saline (PBS). MTT (MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide) and Trypan blue indicated that TFV diminished cell viability at 24–72 h. TFV decreased ATP levels at 72 h when compared to vehicle, reflecting mitochondrial dysfunction. TFV increased the oxidative stress biomarkers of protein carbonylation and 4-hydroxynonenol (4-HNE) adduct formation. Tumor necrosis factor alpha (TNFα) was released into the media following exposure to 14.5 and 28.8 μM TFV. Caspase 3 and 9 cleavage was induced by TFV compared to vehicle at 72 h. These studies show that HK-2 cells are a sensitive model for TFV cytotoxicity and suggest that mitochondrial stress and apoptosis occur in HK-2 cells treated with TFV.

Mitochondrial pathway of apoptosis and necrosis contribute to tenofovir disoproxil fumarate–induced renal damage in rats

“…Our findings are supported by a recent report by Murphy et al 30 These authors have investigated the cellular mechanism of TDF cytotoxicity using a human renal proximal tubular epithelial cell line (HK-2).They have demonstrated that incubation of HK-2 cells with different concentrations of TDF resulted in decreased cell viability, increased oxidative stress biomarkers–protein carbonylation and 4-hydroxynonenol, mitochondrial dysfunction and increased expression of cleaved caspases 3 and 9. These authors have suggested that TDF induces apoptosis in HK-2 cells and that mitochondrial dysfunction may be a major contributing factor to loss of cell viability.…”

In vitro evaluation of biomarkers of nephrotoxicity through gene expression using gentamicin

“…We found that gentamicin has a higher toxicity to HK‐2 than HEK‐293, which was expected since the proximal tubule cells are more sensitive to aminoglycosides, such as gentamicin. This effect was also observed in other studies, even for animal strains such as proximal and distal convoluted tubulus LLC‐PK1 and MDCK, respectively …”

Cinnabar protects serum-nutrient starvation induced apoptosis by improving intracellular oxidative stress and inhibiting the expression of CHOP and PERK

“…Therefore, proximal convoluted tubules are the main toxic target cells of inorganic mercury. The HK-2 cells, which are immortalized human renal cortex proximal convoluted tubule epithelial cells, have various characteristics of normal cortical juxta convoluted tubule epithelial cells, and are often used as an effective model for the study of mercury nephrotoxicity in vitro [ 33 , 34 ]. Therefore, the Hk-2 cells were employed in the present study as well.…”