〔Abstract〕 Objective To study the effects and potential mechanisms of the aquaporin 1(AQP1) inhibitor acetazolamide(AZ) on the proliferation, apoptosis, and inflammatory response of rheumatoid arthritis(RA) fibroblast-like synoviocytes(FLS). Methods TNF-α-induced RA-FLS was served as in vitro RA model. MTT assay, IF staining, and EdU incorporation assay were applied to study AZ′s effects on RA-FLS proliferation. Hoechst staining, flow cytometry analysis of Annexin V-FITC/PI-stained cells, and mitochondrial membrane potential detection experiments were used to detect cell apoptosis. ELISA and quantitative real-time PCR methods were used to measure pro-inflammatory cytokine production. Cell autophagy was evaluated using IF staining and mCherry-GFP-LC3B puncta assay. Western blot was performed to detect the levels of autophagy, apoptosis, and proliferation-related proteins. Moreover, the role of autophagy inhibition in AZ′s effects on RA-FLS was examined by co-treating with the autophagy activator rapamycin(RAPA) or the autophagy inhibitor 3-methyladenine(3-MA). Results AZ dose-dependently inhibited cell proliferation, promoted apoptosis, and reduced the production of pro-inflammatory cytokines in RA-FLS. Furthermore, AZ suppressed cytoprotective autophagy in these cells, as evidenced by decreased LC3B levels levels ( P < 0. 05 ) , increased p62 expression ( P < 0. 05 ) , and reduced autophagic flux ( P <0. 01) . Particularly , AZ ′s beneficial effects were reversed by RAPA⁃induced autophagy activation and enhanced by 3 ⁃MA⁃induced autophagy inhibition. Conclusion This study provides the first evidence that AZ hinder tective autophagy , thereby alleviating the hyperproliferation , apoptosis resistance , and aberrant inflammatory re⁃sponse of RA⁃FLS , revealing the core role of autophagy inhibition in AZ ′s anti⁃RA effects.