Locomotion and brain gene expression exhibit sex-specific non-monotonic dose-response to HFPO-DA during Drosophila melanogaster lifespan

Background: Legacy per-and polyfluoroalkyl substances (PFAS), known for their environmental persistence and bio-accumulative properties, have been phased out in the U.S. due to public health concerns. A newer poly-merization aid used in the manufacture of some fluoropolymers, hexafluoropropylene oxide-dimer acid (HFPO-DA), has lower reported bioaccumulation and toxicity, but is a potential neurotoxicant implicated in dopami-nergic neurodegeneration.Objective: We investigated HFPO-DA's bio-accumulative potential and sex-specific effects on lifespan, locomo-tion, and brain gene expression in fruit flies.Methods: We quantified bioaccumulation of HFPO-DA in fruit flies exposed to 8.7 x 104 mu g/L of HFPO-DA in the fly media for 14 days via UHPLC-MS. Long-term effect on lifespan was determined by exposing both sexes to 8.7 x 102 - 8.7 x 105 mu g/L of HFPO-DA in media. Locomotion was measured following 3, 7, and 14 days of exposures at 8.7 x 101 - 8.7 x 105 mu g/L of HFPO-DA in media, and high-throughput 3 '-end RNA-sequencing was used to quantify gene expression in fly brains across the same time points.Results: Bioaccumulation of HFPO-DA in fruit flies was not detected. HFPO-DA-induced effects on lifespan, locomotion, and brain gene expression, and lowest adverse effect level (LOAEL) showed sexually dimorphic patterns. Locomotion scores significantly decreased in at least one dose at all time points for females and only at 3-day exposure for males, while brain gene expression exhibited non-monotonic dose-response. Differentially expressed genes correlated to locomotion scores revealed sex-specific numbers of positively and negatively correlated genes per functional category.Conclusion: Although HFPO-DA effects on locomotion and survival were significant at doses higher than the US EPA reference dose, the brain transcriptomic profiling reveals sex-specific changes and neurological molecular targets; gene enrichments highlight disproportionately affected categories, including immune response: female -specific co-upregulation suggests potential neuroinflammation. Consistent sex-specific exposure effects necessi-tate blocking for sex in experimental design during HFPO-DA risk assessment.