Legacy and Replacement PFAS Differentially Regulate Intestinal Epithelial Barrier and Inflammatory Gene Expression

Perfluorooctanoic acid (PFOA), a common per- and polyfluoroalkyl substance (PFAS), has been previously associated with inflammatory bowel disease. Hexafluoropropylene oxide dimer acid (HFPO-DA) is a replacement compound for PFOA, yet its impact on primary intestinal epithelial cells (IECs) remains poorly characterized. We hypothesize that exposure to HFPO-DA will alter intestinal epithelial barrier function- and inflammation-related gene expression in primary IECs. Investigating these effects will define mechanistically relevant dose-response relationships and establish a foundational framework for future comparative studies of legacy and replacement PFAS in human intestinal epithelial models. Primary murine IECs were exposed to 100μM PFOA or HFPO-DA for 24 hours, followed by RNA isolation and real-time qPCR for gene expression quantification. A methanol vehicle control was included to account for solvent effects. Expression of inflammatory and junctional genes differed between groups. Downregulation of Tnf and Il-6 along in IECs exposed to HFPO-DA relative to IECs exposed to PFOA suggests HFPO-DA may elicit a comparatively reduced inflammatory response. Upregulation of Tslp in HFPO-DA-exposed cells compared to PFOA-exposed cells may indicate an altered capacity for intestinal epithelial barrier repair. Inconsistent differences in junctional gene expression could be further clarified through protein-level assays and trans-epithelial electrical resistance measurements. These findings highlight the need for continued investigation into HFPO-DA’s effects on the intestinal epithelial barrier. Future studies should incorporate additional biological replicates, a broader dose range, and expanded gene panels, complemented by protein-level and functional integrity analyses alongside transcriptional expression analyses.