Abstract
Honokiol, a lignan found in Magnolia plant species, exerts diverse pharmacological effects, and induces apoptosis in several cancer cell lines, including human hepatocellular carcinoma (HCC). The present study aimed to investigate whether it could induce paraptosis-like cell death, a type of non-canonical programmed cell death characterized by vacuolation and dysfunction of the mitochondria and endoplasmic reticulum (ER), in HCC Hep3B cells. Our results showed that honokiol significantly induced cytotoxicity and autophagy, both of which were associated with cytoplasmic vacuolation. Honokiol also enhanced ER stress, increased cellular calcium ion (Ca(2+)) levels, and caused mitochondrial dysfunction. Honokiol upregulated the expression of mitophagy regulators such as PTEN-induced kinase 1 and Parkin in the mitochondria, whereas the expression of apoptosis-linked gene 2-interacting protein X (Alix), involved in suppressing paraptosis, was downregulated. In addition, honokiol-induced cytotoxicity was accompanied by excessive generation of intracellular reactive oxygen species (ROS) and mitochondrial ROS (mtROS). However, the addition of Mito-TEMPO, a mitochondria-targeting antioxidant, neutralized the honokiol-induced increase in Ca(2+) levels and changes in autophagy, ER stress, and mitophagy regulatory protein expression, thereby counteracting ER stress. Moreover, Mito-TEMPO pretreatment significantly improved honokiol-induced mitochondrial impairment, cytotoxicity, and Alix expression. Collectively, our findings demonstrate that honokiol-induced oxidative stress in HCC Hep3B cells critically contributes to subsequent paraptotic events such as ER stress and mitochondrial damage, highlighting the potential of honokiol as a therapeutic agent for liver cancer treatment. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s43188-025-00291-2.