Sirtuin 1 (SIRT1), a NAD(+)-dependent class III histone deacetylase, participates in regulating cellular apoptosis, senescence and metabolism by deacetylating histones and multiple transcription factors. In this study, we aimed to determine the effect of SIRT1 on the apoptosis of vascular adventitial fibroblasts (VAFs) and related signaling pathways. SIRT1 was found in the nucleus of VAFs and translocated into the cytoplasm in response to tumor necrosis factor-α (TNF-α). Moreover, SIRT1 protein expression was reduced in VAFs stimulated with TNF-α. In addition, TNF-α increased the apoptosis of VAFs. Activation of SIRT1 by resveratrol (RSV) or overexpression of SIRT1 attenuated TNF-α-induced VAF apoptosis by decreasing the percentage of apoptotic cells and cleaved caspase-3 protein expression and increasing the Bcl-2/Bax ratio. In contrast, inhibition of SIRT1 by sirtinol/nicotinamide or knockdown of SIRT1 enhanced apoptosis of VAFs. On the other hand, knockdown of FoxO1 reduced TNF-α-induced VAF apoptosis. SIRT1 interacted with FoxO1 in VAFs by the co-immunoprecipitation assay. Further study showed that RSV or SIRT1 overexpression decreased acetylated-FoxO1 (Ac-FoxO1) protein expression in VAFs stimulated with TNF-α. Knockdown of SIRT1 resulted in an increase in Ac-FoxO1 protein expression. Taken together, these findings indicate that SIRT1 inhibits the apoptosis of VAFs, whereas FoxO1 promotes VAF apoptosis. Furthermore, the inhibitory effect of SIRT1 on VAF apoptosis is partly mediated by the deacetylation of FoxO1.