Several lines of evidence support the view that rapamycin inhibits nuclear factor (NF)-B. Tumor necrosis factor (TNF)- is a potent NF-B inducer. This cytokine is released after artery injury, as occurs for example after balloon angioplasty, and plays an important role in inflammation and restenosis. We investigated the effect of rapamycin on NF-B activation and apoptosis in vascular smooth muscle cells (VSMCs) stimulated with TNF-. Using electrophoretic mobility shift assay, we found that TNF- caused NF-B nuclear translocation in VSMCs after 1 h of incubation. Rapamycin inhibited IB degradation thereby preventing nuclear translocation. Activation of NF-B was accompanied by an increase of Bcl-xL and Bfl-1/A1 proteins, detected by Western blot assay, whereas rapamycin prevented the TNF--induced enhancement of these anti-apoptotic molecules. The extent of death of VSMCs exposed to TNF- was significantly enhanced by rapamycin. The effect of rapamycin appeared to be independent of the phosphatidyl-inositol 3 kinase (PI3k)/Akt-protein kinase B survival pathway because the PI3k inhibitor wortmannin neither prevented IB degradation nor increased apoptosis of cells incubated with TNF-. Finally, we demonstrate that the large immunophilin FKBP51 is essential for TNF--induced NF-B-activation in VSMCs. Our findings show that rapamycin inhibits NF-B activation and act in concert with TNF- in inducing VSMC apoptosis.