Gestational exposure to hexafluoropropylene oxide trimer acid impairs the reproductive function of male offspring through the LCN2-ferroptosis axis in spermatocytes.

Hexafluoropropylene oxide trimer acid (HFPO-TA), introduced as an industrial replacement for perfluorooctanoic acid (PFOA), has raised concern regarding its potential bioaccumulation and toxicity. However, the reproductive toxicity and mechanistic basis of HFPO-TA prenatal exposure in male offspring remain unclear. A mouse model of gestational HFPO-TA exposure (0.4 and 1 mg/kg/day) was established, and integrated histopathological assessment, single-cell transcriptomics, and molecular simulation approaches were performed to characterize its effects on male offspring. HFPO-TA exposure significantly impaired the reproductive function in F1 male mice, as indicated by reduced testicular weight and sex hormone levels, compromised sperm quality, spermatocyte meiotic arrest, and increased germ cell apoptosis. Single-cell sequencing revealed that prenatal HFPO-TA exposure triggered ferroptosis in spermatocytes of male offspring mice and identified lipocalin-2 (Lcn2) as a key regulator of this response. Molecular docking and dynamics simulations confirmed stable binding between HFPO-TA and LCN2. In vitro experiments further demonstrated that HFPO-TA induced spermatocyte ferroptosis through lipid peroxidation and iron overload, while these effects were reversed by treatment with the ferroptosis inhibitor ferrostatin-1 or by Lcn2 overexpression. Collectively, this study delineated a mechanistic framework in which prenatal HFPO-TA exposure induces spermatocyte-specific injury of male offspring through the LCN2-ferroptosis pathway, providing a scientific basis for health risk assessment and regulation in the use of next-generation per- and polyfluoroalkyl substances.