Effects of α-Linolenic Acid and Arachidonic Acid on the Cytotoxicity and Steatosis of HepG2 Cells

Abstract

Abstract Hepatic steatosis, characterized by excessive triglyceride (TG) accumulation in hepatocytes, serves as the initiating event in the early phase of metabolic dysfunction-associated fatty liver disease (MAFLD). Polyunsaturated fatty acids (PUFAs) exert differential modulatory effects on hepatic steatosis, but the underlying lipidomic remodeling remains incompletely understood. In this study, we systematically investigated the impacts of α-linolenic acid (ALA) and arachidonic acid (AA) on lipid accumulation, inflammation, and cytotoxicity in fattyacid-induced HepG2 cells. Our data indicated that ALA at concentrations >300 μM and AA at >150 μM significantly inhibited cell viability. In fatty acid-induced steatotic HepG2 cells, 100 μM ALA significantly reduced both triglyceride (TG) and total cholesterol (TC) levels, whereas 100 μM AA significantly increased TG levels without affecting TC content. Furthermore, quantitative real-time PCR revealed that 100 μM ALA significantly downregulated the transcriptional expression of NF-κB, TNF-α, and IL-6, while 100 μM AA upregulated the expression of these three factors. The untargeted metabolomic profiling and KEGG pathway enrichment analysis showed that ALA significantly modulated glycerophospholipid metabolism, amino acid metabolism, nucleotide metabolism, and central carbon metabolism in steatotic HepG2 cells. Lipidomics analysis further demonstrated that ALA, through conversion to DHA and EPA, remodeled TG molecular species and was incorporated into multiple membrane phospholipids. Furthermore, Drug Affinity Responsive Target Stability (DARTS) and molecular docking results indicated that choline kinase α (CHKA) might be a potential target of ALA in HepG2 cells. Conclusively, ALA might contribute to the progression of hepatocyte steatosis by remodeling TG molecular species, incorporating into membrane phospholipids, modulating glycerophospholipid metabolism, and targeting CHKA.

The authors' abstract, as published at the source. ACS Omega, 2026 · DOI ↗