Ischaemic postconditioning (IPostC) is a powerful protective phenomenon that activates prosurvival intrinsic signalling cascades to limit reperfusion injury. We propose that IPostC confers its infarct-sparing effect via activation of the newly described prosurvival Survivor Activating Factor Enhancement (SAFE) pathway, which involves the activation of the cytokine tumour necrosis factor alpha (TNFalpha) and signal transducer and activator of transcription-3 (STAT-3).Isolated ischaemic/reperfused hearts from TNF knockout, TNF receptor-1 knockout, TNF receptor-2 knockout, cardiomyocyte-specific STAT-3-deficient mice or their respective wild-type, (TNF-WT) or (STAT-3-WT), were postconditioned by ischaemic episodes (IPostC) or with exogenous TNFalpha (0.5 microg/L) (TNF-PostC) at the onset of reperfusion. IPostC reduced infarct size (IS) in TNF-WT and TNFR1(-/-) hearts (by 33 and 27%, respectively, P < 0.05), whereas hearts from TNF(-/-) or TNFR2(-/-) failed to be postconditioned. TNF-PostC reduced IS by 37% (P < 0.05) in STAT-3-WT hearts but failed to protect cardiac-specific STAT-3(-/-) hearts. Administration of wortmannin, an inhibitor of PI-3 kinase/Akt, or PD98059, an inhibitor of extracellular regulated kinase 1/2 (Erk1/2), during the postconditioning stimulus did not abolish the infarct-sparing effect of TNF-PostC. AG490, an inhibitor of STAT-3, abrogated the protective effect of TNFalpha. Western blot analysis did not demonstrate the involvement of Akt or Erk1/2 in TNF-PostC, whereas STAT-3 phosphorylation was increased in both IPostC and TNF-PostC.The protective effect of the SAFE pathway is shown in IPostC, with the activation of TNFalpha, its receptor type 2, and STAT-3. This signalling cascade is activated independently of the well-known Reperfusion Injury Salvage Kinases (RISK) pathway, which involves the kinases Akt and Erk1/2.