Excessive and unresolved neuroinflammation plays an important role in the pathophysiology of many neurological disorders, such as ischemic stroke, yet there are no effective treatments. Tripartite motif-containing 67 (TRIM67) plays a crucial role in the control of inflammatory disease and pathogen infection-induced inflammation; however, the role of TRIM67 in cerebral ischemia‒reperfusion injury remains poorly understood.In the present study, we demonstrated that the expression level of TRIM67 was significantly reduced in middle cerebral artery occlusion and reperfusion (MCAO/R) mice and primary cultured microglia subjected to oxygen-glucose deprivation and reperfusion. Furthermore, a significant reduction in infarct size and neurological deficits was observed in mice after TRIM67 upregulation. Interestingly, TRIM67 upregulation alleviated neuroinflammation and cell death after cerebral ischemia‒reperfusion injury in MCAO/R mice. A mechanistic study showed that TRIM67 bound to IκBα, reduced K48-linked ubiquitination and increased K63-linked ubiquitination, thereby inhibiting its degradation and promoting the stability of IκBα, ultimately inhibiting NF-κB activity after cerebral ischemia.Taken together, this study demonstrated a previously unidentified mechanism whereby TRIM67 regulates neuroinflammation and neuronal apoptosis and strongly indicates that upregulation of TRIM67 may provide therapeutic benefits for ischemic stroke.© 2023. The Author(s).