Decoding PFAS immunotoxicity: a NAMs-based comparison of short vs. long chains.

Introduction: Per- and polyfluoroalkyl substances (PFAS) are persistent environmental pollutants with potential immunotoxic effects. Most toxicological studies have focused on long-chain PFAS such as perfluorooctanesulfonic acid (PFOS) and perfluorooctanoic acid (PFOA). However, short-chain and ultra-short-chain alternatives, including trifluoroacetic acid (TFA), are increasingly used despite limited toxicological data. Methods: This study evaluated and compared the immunotoxic effects of PFAS with varying chain lengths-long-, short-, ultra-short-chain compounds, and fluoropolymer representatives (polytetrafluoroethylene, PTFE)-using human-relevant new approach methodologies (NAMs). Two complementary in vitro models were employed. Peripheral blood mononuclear cells (PBMCs) from healthy donors to assess antibody (IgG and IgM) production. THP-1-derived dendritic cells (DCs) to evaluate maturation marker expression (CD83, CD86, HLA-DR). Environmentally and occupationally relevant PFAS concentrations were tested. Results: PFOS, PFOA, and perfluorononanoic acid (PFNA) suppressed antibody production and impaired DC maturation in a concentration-dependent manner, consistent with previous in vivo and epidemiological data. Short-chain PFAS (PFHxS, PFBS, PFHxA, PFBA) showed modest to intermediate immunomodulatory activity, with subtle immunosuppressive trends in female donors. Notably, TFA reduced antibody production at levels comparable to PFOS, indicating that chain length alone is not a reliable predictor of immunotoxic potential. PTFE exhibited no suppressive effects; instead, increased antibody release was observed in female donors, suggesting possible sex-dependent immunostimulation. Discussion: Findings support a nuanced, compound-specific approach to PFAS risk assessment rather than a simple long- vs. short-chain distinction. In vitro NAMs provided mechanistic, human-relevant insights and reinforce their integration into regulatory frameworks.