Recent evidence suggests that Toll-like receptor 4 (TLR4) is not only involved in innate immunity but is also an important mediator of adverse left ventricular remodeling and heart failure following acute myocardial infarction (MI). TLR4 is activated by lipopolysaccharide (LPS) but also by products of matrix degradation such as hyaluronic acid and heparan sulfate. Although cardioprotective properties of adenosine (Ado) have been extensively studied, its potential to interfere with TLR4 activation is unknown. We observed that TLR4 pathway is activated in white blood cells from MI patients. TLR4 mRNA expression correlated with troponin T levels (R (2) = 0.75; P = 0.01) but not with levels of white blood cells and C-reactive protein. Ado downregulated TLR4 expression at the surface of human macrophages (-50%, P < 0.05). Tumor necrosis factor-α production induced by the TLR4 ligands LPS, hyaluronic acid, and heparan sulfate was potently inhibited by Ado (-75% for LPS, P < 0.005). This effect was reproduced by the A2A Ado receptor agonist CGS21680 and the non-selective agonist NECA and was inhibited by the A2A antagonist SCH58261 and the A2A/A2B antagonist ZM241,385. In contrast, Ado induced a 3-fold increase of TLR4 mRNA expression (P = 0.008), revealing the existence of a feedback mechanism to compensate for the loss of TLR4 expression at the cell surface. In conclusion, the TLR4 pathway is activated after MI and correlates with infarct severity but not with the extent of inflammation. Reduction of TLR4 expression by Ado may therefore represent an important strategy to limit remodeling post-MI.