Perfluorooctanoic acid (PFOA) induces lipid accumulation, oxidative stress, and reduced neurogenesis in primary human neuronal progenitor cells

Introduction Per- and polyfluoroalkyl substances (PFAS), particularly perfluorooctanoic acid (PFOA), are persistent environmental contaminants known for bioaccumulation and adverse health effects, including neurodevelopmental toxicity. This study investigated the impact of PFOA on primary human neuronal progenitor cells (phNPCs) derived from fetal brain tissue from genetically diverse donors, focusing on lipid metabolism and neuronal differentiation. Methods phNPCs were exposed in vitro to PFOA at high concentrations (10,000–156 μM range) to determine cell viability and cytotoxicity using Alamar blue and lactate dehydrogenase (LDH) assays, respectively. Further experiments were conducted in 300–0.3 μM range where no effects on cell viability or cytotoxicity were observed. phNPCs were treated acutely (2 days) and assessed for changes in lipid droplet accumulation, fatty acid metabolism, lipid peroxidation, mitochondrial damage, and proliferation (EdU, Ki67, pHH3 staining). phNPCs were then exposed to PFOA for 14-days in neuronal differentiation media and assessed for changes in neuronal gene expression using quantitative reverse transcription polymerase chain reaction (RT-qPCR) and MAP2 protein expression and neuronal morphology using high content imaging. To assess differences in cytotoxicity between neuronal progenitors and neurons, fully differentiated neurons and phNPCs were both exposed to high concentrations (10,000–156 μM range) for 14 days and assessed for impacts on cell viability and death using Alamar Blue assays and flow cytometry using Calcein-AM/7-AAD stained cells. Results Acute PFOA exposure induced dose-dependent lipid droplet accumulation, increased fatty acid uptake, reduced lipid turnover, elevated lipid peroxidation, mitochondrial reactive oxygen species, and fragmented mitochondrial morphology. The PFOA-induced lipid droplet accumulation was attenuated by inhibition of autophagy and lipolysis pathways, suggesting PFOA-induced lipotoxicity. PFOA exposure had minimal effects on phNPC proliferation but 14-day exposure during neuronal differentiation reduced MAP2-positive neurons, neuronal branching and gene expression of neuronal markers ( TUBB3, SYN1, MAP2 ), while increasing the gene expression of progenitor-associated FABP7 . Principal component analysis revealed PFOA-exposed cells exhibited intermediate gene expression between progenitors and mature neurons. Treatment of fully differentiated neurons during the same time window resulted in increased death cell and reduced viability compared phNPCs, suggesting neurons are more susceptible to PFOA cytotoxicity. Across donors, greater PFOA-induced lipid accumulation negatively correlated with neuronal differentiation outcomes. Discussion These findings indicate that PFOA disrupts human neurodevelopment primarily by impairing neuronal differentiation, potentially through lipotoxicity and mitochondrial stress, highlighting a mechanistic link between dysregulated lipid metabolism and reduced neurogenesis.