β‐Adrenergic Receptor Signaling Reduces Pro‐Inflammatory Signaling and Promotes the M2 Phenotype in the RAW264.7 Macrophage Cell Line

Catecholamines alter macrophage function by binding to adrenergic receptors on the cell membrane. Stimulation through β2-adrenergic receptors is known to down regulate the activity of classically activated M1 macrophages while stimulation through α-adrenergic receptors typically enhances their function. M1 macrophages initiate the pro-inflammatory response to eliminate infection while alternatively activated M2 macrophages down regulate the immune response and promote tissue repair. Our studies with the RAW264.7 murine macrophage cell line as a model for macrophage function support these findings. Previously, we exposed macrophages to the selective β2-adrenergic receptor agonist formoterol or the non-selective α-adrenergic receptor agonist clonidine prior to lipopolysaccharide (LPS) activation and analyzed them for production of the proinflammatory cytokines tumor necrosis factorα (TNFα) and interleukin-6 (IL-6) by ELISA. The results showed that formoterol decreased and clonidine increased macrophage cytokine production. The MAP-Kinase (MAPK) signaling pathway plays an important role in the proinflammatory response, and our recent results obtained by western blotting show that norepinephrine (NE) reduces the activation of extracellular signal-regulated kinase (ERK 1/2) by decreasing the time that the MAPK is phosphorylated. Further studies using quantitative PCR (qPCR) show that expression of inducible nitric oxide synthase (iNOS) mRNA was reduced while expression of arginase-1 (Arg-1) mRNA increased in macrophages pre-treated with NE prior to LPS activation. The iNOS enzyme catalyzes the production of nitric oxide, which has antimicrobial properties, and is a marker for M1 macrophages. The Arg-1 enzyme catalyzes the reaction of arginine to ornithine, a precursor to molecules participating in wound healing, and is indicative of M2 macrophages. Therefore, these findings suggest that stimulation of the adrenergic receptors by NE may also alter the phenotype of the macrophages. Taken together these findings suggest that exposure to NE may not only reduce the proinflammatory activity of macrophages but may also induce development of the alternatively activated M2 phenotype in RAW264.7 macrophages.