Transmission electron microscopy of the benzbromarone-induced change in mitochondrial morphology in HepG2 cells

Abstract

Drug-induced mitochondrial dysfunction can lead to severe adverse effects. Accordingly, new in vitro assay systems for assessing mitochondrial-related toxicity are required. Current systems evaluate drug-induced mitochondrial dysfunction based on cell death. However, if mitochondria are damaged without cell death, these methods run the risk of overlooking toxic or dangerous compounds. To solve this problem, we attempted to measure morphological changes semi-quantitatively by transmission electron microscopy and to detect subtle changes in mitochondrial function. To this end, we exposed HepG2 cells cultured in galactose-containing medium to benzbromarone (BBR), which impairs mitochondrial function. After 24 hr of BBR exposure, we compared the rate of cell death between galactose and glucose cultures. Before the onset of cell death, BBR increased the ratio of damaged mitochondria to a greater extent in galactose-cultured than glucose-cultured HepG2 cells. Our results suggested that this new in vitro assay system could detect mitochondrial-related toxicity before the onset of cell death.

Hepatocellular organellar abnormalities following elimination of hepatitis C virus

“…Because quantitative analyses are challenging, EM is typically employed to examine organellar abnormalities qualitatively. We recently succeeded in a semi‐quantitative toxicity assessment of mitochondrial abnormalities following treatment at increasing concentrations with drugs toxic to mitochondria 17 . The presence of organellar changes was determined by averaging the scores of ten randomly selected cells using a 5‐point scoring system (−, −/+, +, ++, +++).…”

“…Liver biopsies were prepared as previously described 14 . The ultrastructural status of the liver was observed using TEM (H‐7100, Hitachi Ltd.) and evaluated semi‐quantitatively using the previously described scoring system 15–17 . In this study, hepatocytes near the central vein and portal vein were not used for observation to avoid the influence of the vascular system.…”

“…We recently succeeded in a semi-quantitative toxicity assessment of mitochondrial abnormalities following treatment at increasing concentrations with drugs toxic to mitochondria. 17 The presence of organellar changes was determined by averaging the scores of ten randomly selected cells using a 5-point scoring system (−, −/+, +, ++, +++). In patients with CHC, the hyperfibrosis group had more DMV-like structures (p = .009) than the hypofibrosis group (Figure S1.…”