The function of blood-brain barrier is often disrupted during the progression of multiple sclerosis and its animal model, experimental autoimmune encephalomyelitis (EAE). However, the molecular mechanism of blood-brain barrier modulation during neuroinflammation remains unclear. Herein, we show that the expression of interferon-γ (IFNγ) receptor on endothelial cells (ECs) protected mice from the brain inflammation during EAE. IFNγ stabilized the integrity of the cerebral endothelium and prevented the infiltration of leukocytes into the brain. Further analysis revealed that IFNγ increased the expression of tight junction proteins zonula occludens protein 1 and occludin, as well as membranous distribution of claudin-5, in brain ECs. Silencing claudin-5 abolished the IFNγ-mediated improvement of EC integrity. Taken together, our results show that IFNγ, a pleiotropic proinflammatory cytokine, stabilizes blood-brain barrier integrity and, therefore, prevents brain inflammation during EAE.