Lycopene attenuates DEHP-induced oxidative stress, mitochondrial dysfunction, and apoptosis in human granulosa cells via SIRT1-dependent mechanisms.

Abstract

Di(2-ethylhexyl) phthalate (DEHP) is a widely used plasticizer and endocrine disruptor that can impair ovarian function by provoking oxidative stress, mitochondrial injury, and apoptosis in granulosa cells. Lycopene is a diet-derived carotenoid with antioxidant and mitochondria-protective activities. This study investigated whether lycopene protects human granulosa-like KGN and COV434 cells against DEHP-induced damage and whether SIRT1 mediates these effects. Cells were exposed to DEHP (50–800 µM) for 24–72 h to define concentration- and time-dependent cytotoxicity, followed by co-treatment with lycopene (1–3 µM). Cell viability was assessed using CCK-8, and apoptosis was evaluated by caspase-3/-7 activity. Intracellular reactive oxygen species (ROS) were quantified using DCFH-DA fluorescence. Mitochondrial function was assessed by JC-1 staining for mitochondrial membrane potential and by qPCR determination of mitochondrial DNA (mtDNA) copy number. Antioxidant defense and lipid peroxidation were evaluated by measuring superoxide dismutase (SOD) and catalase (CAT) activities and malondialdehyde (MDA) levels. SIRT1 expression was examined at the transcript and protein levels, and siRNA-mediated SIRT1 knockdown was used to test mechanistic dependence. DEHP significantly reduced cell viability and increased caspase-3/-7 activity in both cell lines, accompanied by ROS accumulation, loss of mitochondrial membrane potential, reduced mtDNA copy number, decreased SOD/CAT activities, and elevated MDA. Lycopene markedly mitigated these effects, restoring redox balance, antioxidant capacity, and mitochondrial integrity while suppressing apoptotic signaling. DEHP downregulated SIRT1, whereas lycopene restored SIRT1 expression, and SIRT1 silencing partially weakened lycopene-mediated cytoprotection. Collectively, these findings identify a SIRT1-dependent protective action of lycopene against DEHP-induced granulosa cell oxidative and mitochondrial injury, supporting lycopene as a potential strategy to mitigate phthalate-associated ovarian toxicity.