AbstractInflammatory and invasive fibroblast‐like synoviocytes (FLS) contribute to the pathology of rheumatoid arthritis (RA). Isoginkgetin (IGKG) has been identified as having anti‐inflammatory properties. This study investigated whether IGKG could be utilized to treat RA. Primary FLS were isolated from synovial tissues derived from six RA patients, which were over‐expressed with matrix metallopeptidase 9 and cultured with or without tumor necrosis factor (TNF)‐α and then further treated with IGKG. IGKG down‐regulated the content of various interleukins (ILs), namely, IL‐1β, IL‐6, and IL‐8, in RA‐FLS supernatant with or without TNF‐α stimulation, with diminished migration and invasion properties as assayed by the transwell system. Furthermore, down‐regulated inflammatory cytokine secretion and down‐regulated migration and invasion properties could be reversed through matrix metallopeptidase 9 overexpression. Dual‐luciferase reporter gene assay indicated that IGKG could inhibit nuclear factor kappa B transcription activity. Western blot analysis also demonstrated that IGKG down‐regulated the expression of p‐IκBα, p‐p65, and MMP9. IGKG displayed the ability to inhibit the inflammatory response of RA‐FLS through the NF‐κB/MMP9 pathway with diminished migration and invasion.