Sex Hormone‐Dependent Modulation of Nrf2 and Its Association With Apoptosis, Metastasis, and Drug Resistance in Hepatocellular Carcinoma

Hepatocellular carcinoma (HCC) exhibits a gender disparity, with a 2‐ to fourfold higher incidence in men, as attributed by previous studies to estrogen‐mediated hepatoprotection versus the tumor‐promoting role of androgens. This study investigated the effects of these sex hormones—testosterone and β‐estradiol—on Nrf2 expression and its relation to key factors in metastasis, apoptosis, and drug resistance in HepG2 HCC cell line. HepG2 cells were exposed to testosterone or β‐estradiol at viability‐equivalent doses. Oxidative stress was quantified by total oxidant status (TOS), malondialdehyde (MDA), and total antioxidant capacity (TAC) assays. Apoptosis was evaluated by Annexin V/PI flow cytometry. mRNA levels of Nrf2, MRP‐1 (multidrug resistance protein 1), MMP‐9 (matrix metalloproteinase‐9), BCL2, and SIRT1 were assessed via qRT‐PCR. At viability‐equivalent doses, β‐estradiol elicited dose‐dependent apoptosis in HepG2 cells (total: 47.4% at V₅₀ vs. 32.7% for testosterone; p < 0.001), predominantly late‐stage (34.5% vs. 21.8%; p < 0.0001). Concomitantly, β‐estradiol decreased TOS (maximal at V₄₀, p < 0.05) and MDA (all doses, p < 0.05) while increasing TAC (medium/high doses, p < 0.05)—effects not observed with testosterone ( p > 0.05). β‐estradiol induced downregulation of Nrf2, BCL2, MMP‐9, and MRP‐1 mRNA (moderate/high doses; p < 0.05 vs. control). Conversely, testosterone upregulated SIRT1 across doses ( p < 0.05), unaffected by β‐estradiol ( p > 0.05). β‐Estradiol mitigates oxidative stress, induces apoptosis, and suppresses pro‐survival, metastatic, and chemoresistant pathways—contrasting testosterone's minimal effects. These findings align with HCC's male predominance and highlight the hormone‐modulated strategies' potential for future therapeutic exploration.