Drug-Induced Alterations of Mitochondrial DNA Homeostasis in Steatotic and Nonsteatotic HepaRG Cells

Area of Science:

• Hepatotoxicity and Mitochondrial Biology
• Drug-induced Liver Injury
• Cellular Metabolism and Adaptation

Background:

• Mitochondrial toxicity is a key factor in drug-induced liver injury, but alterations in mitochondrial DNA (mtDNA) homeostasis are rarely studied due to the need for prolonged drug exposure.
• Standard toxicity tests using isolated mitochondria or short-term cell cultures may miss delayed toxicity mechanisms related to mtDNA homeostasis.
• Nonalcoholic fatty liver disease (NAFLD) involves lipid overload, which may influence drug-induced mitochondrial dysfunction.

Purpose of the Study:

• To investigate the effects of a 2-week treatment with nine hepatotoxic drugs on mtDNA homeostasis in HepaRG cells.
• To determine if drug-induced mitochondriotoxicity is modulated by lipid overload, mimicking nonalcoholic fatty liver conditions.
• To explore the potential for delayed toxicity secondary to mtDNA homeostasis impairment.

Main Methods:

• HepaRG cells were treated for 2 weeks with nine hepatotoxic drugs at concentrations causing ~10% cytotoxicity.
• Zalcitabine and linezolid were used as positive controls for mtDNA replication and translation effects, respectively.
• Mitochondrial effects, mtDNA levels, and expression of key biogenesis factors (PGC1α, PGC1β, AMPKα) were assessed, with and without lipid overload.

Main Results:

• Imipramine and ritonavir showed mitochondrial effects suggesting altered mtDNA translation; ritonavir toxicity was greater in non-steatotic cells.
• None of the nine drugs decreased mtDNA levels, but five drugs increased mtDNA, particularly in non-steatotic cells.
• Increased mtDNA levels did not correlate with key mitochondrial biogenesis factors, suggesting an adaptive response rather than increased biogenesis.

Conclusions:

• Drug-induced impairment of mtDNA translation may be common, and increased mtDNA levels can be a frequent adaptive response to mild energy deficits.
• This adaptive response to mtDNA translation impairment might be compromised by lipid overload, relevant to NAFLD pathogenesis.
• Prolonged cell culture models are necessary to detect delayed mitochondriotoxicity related to mtDNA homeostasis.