Eicosapentaenoic acid and docosahexaenoic acid synergistically attenuate bile acid-induced hepatocellular apoptosis

Background: Clinical studies have demonstrated improvement of parenteral nutrition (PN)–associated liver disease (PNALD) with ω3 polyunsaturated fatty acid (ω3PUFA) supplementation containing eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). Experiments were designed to test the following hypotheses: (1) therapeutic effects of ω3PUFA are due to attenuation of cellular apoptosis induced by hydrophobic bile acid exposure, which occurs in cholestasis, and (2) attenuation of apoptosis by EPA and DHA is additive or synergistic. Methods: Cultured HepG2 cells were treated with 50–200 µM chenodeoxycholic acid (CDCA) in the presence and absence of EPA, DHA, or EPA + DHA. Apoptosis was evaluated using cell staining with fluorescence microscopy and the Apo‐ONE Homogeneous Caspase‐3/7 assay. Specific apoptotic mediators were evaluated with quantitative RT‐PCR. Results: Treatment with EPA alone and DHA alone resulted in 22% and 9% attenuation of caspase‐3/7 activity, respectively. Caspase‐3/7 activity was attenuated by 52% when cells were treated with a combination of EPA and DHA (P = .0034). Treatment with EPA alone, DHA alone, and the combination of EPA and DHA all resulted in equal attenuation of apoptotic mediator gene expression. Conclusions: The combination of EPA and DHA resulted in a synergistic attenuation of bile acid–induced hepatocellular apoptosis, as assessed by caspase‐3/7 activity, compared to EPA and DHA separately. The combination of EPA and DHA did not result in a synergistic attenuation of the upregulation of Fas or TRAIL‐R2. These data suggest that EPA and DHA may be working via multiple intracellular pathways to attenuate bile acid–induced apoptosis.