β2-Adrenergic receptor regulates Toll-like receptor-4-induced nuclear factor-κB activation through β-arrestin 2

Toll-like receptors (TLRs) play an important role in innate immunity while, beta(2)-adrenergic receptors (beta(2)AR) provide the key linkages for the sympathetic nervous system to regulate the immune system. However, their role in macrophages remains uncertain. Here, we demonstrate the cross-talk between beta(2)AR and TLR signalling pathways. Expression of beta(2)AR was down-regulated by TLR4 ligand lipopolysaccharide (LPS) stimulation. To investigate the physiological consequence of this down-regulation RAW264 cells, a macrophage cell line, were transfected with a beta(2)AR expression vector (RAWar). Both LPS-stimulated inducible nitric oxide synthase (NOS II) expression and NO production were markedly suppressed in the RAWar cells. The activation of nuclear factor-kappa B (NF-kappa B) and degradation of the inhibitor of NF-kappa B (I kappa B alpha) in response to LPS were markedly decreased in these cells. The level of beta-arrestin 2, which regulates beta(2)AR signalling, was also reduced in RAW264 cells after stimulation with LPS, but not in RAWar cells. Overexpression of beta-arrestin 2 (RAWarr2) also inhibited NO production and NOS II expression. Furthermore, we demonstrated that beta-arrestin 2 interacted with cytosolic I kappa B alpha and that the level of I kappa B alpha coimmunoprecipitated by anti-beta-arrestin 2 antibodies was decreased in the RAW264 cells but not in RAWar or RAWarr2 cells. These findings suggest that LPS-stimulated signals suppress beta(2)AR expression, leading to down-regulation of beta-arrestin 2 expression, which stabilizes cytosolic I kappa B alpha and inhibits the NF-kappa B activation essential for NOS II expression, probably to ensure rapid and sufficient production of NO in response to microbial attack.