Trichloroethylene (TCE) and perfluorooctanoic acid (PFOA) commonly co-occur at industrially contaminated sites. However, how PFOA affects the microbial reductive dechlorination of TCE remains unclear. This study aims to investigate the effects of PFOA on the anaerobic microbial reductive dechlorination of TCE by an enriched microbial consortium, and to further elucidate the response in the composition and function of extracellular polymeric substances (EPS). The results revealed that the TCE-degrading microbial consortium was capable of completely degrading 76 μmol of TCE within 3 days under both TCE-only and PFOA co-exposure conditions. PFOA exerted a stage-dependent dual effect on the TCE dechlorination process: during the early reaction phase (7 days), PFOA promoted the transformation of TCE to cis-1,2-dichloroethylene and vinyl chloride (VC), accompanied by an increase in electron transport system activity compared to the TCE-only group. However, during the later stage of incubation (21 days), PFOA inhibited the further transformation of VC. Particularly at the high PFOA concentration (10 mg/L), the concentration of VC reached the highest level, while the electron transport system activity decreased by 65.0% compared with the initial stage. Furthermore, analysis of the chemical composition, structure, and electrochemical properties of EPS indicated that PFOA exposure, particularly at 1 mg/L, significantly enhanced the total secretion of EPS and increased the protein/polysaccharide ratio. It also induced nitrogen enrichment in the protein component of EPS and promoted the transformation of oxygen-containing functional groups from carbonyl to alcohol/acetal structures. Concurrently, PFOA strengthened the redox activity and electron transfer capacity of EPS, thereby influencing the transformation of TCE to cis-1,2-DCE and VC. This study demonstrates that PFOA exerts concentration-dependent and stage-specific effects on the anaerobic reductive dechlorination of TCE. The results highlight that EPS play a crucial role in microbial adaptation and maintaining dechlorination performance under PFOA exposure. This study provides a theoretical basis for the bioremediation of chlorinated hydrocarbon contaminated sites in the presence of PFOA.