Pathways of anti-inflammatory signaling are potential targets in developing therapeutic options for diseases of inflammation, such as sepsis. Interleukin-10 (IL-10) is an important anti-inflammatory cytokine that can be regulated by epinephrine. It has been shown that epinephrine acts through a mechanism involving cyclic AMP (cAMP), an important intracellular signaling molecule that is critical in regulating the immune response. cAMP has commonly been thought to act only through protein kinase A (PKA), however recent studies suggest that cAMP may also act through PKA-independent pathways. In this study we investigate the mechanism by which epinephrine regulates IL-10 cytokine production. Human promonocytic THP-1 cells were pretreated for 48 h with 100 nM vitamin D prior to stimulation with lipopolysaccharide (LPS, 100 ng/mL) and epinephrine (10 μM) for 24 h. Some cells were pretreated for 15 min with propranolol (10 μM, a β2 adrenergic receptor (β2AR) blocker), H89 (1000 nM, an inhibitor of the catalytic subunit of PKA), Rp-cAMP (300 nM, a competitive inhibitor of PKA), or SQ22536 (1000 μM, an inhibitor of adenylate cyclase) prior to stimulation with LPS and epinephrine. IL-10 cytokine production and mRNA expression levels were measured by enzyme-linked immunosorbent assay (ELISA) and reverse transcriptase-polymerase chain reaction (RT-PCR), respectively. Epinephrine significantly increased LPS-induced IL-10 cytokine production at 24 h (LPS = 24.0 ± 2.1 pg/mL vs. epinephrine + LPS = 93.7 ± 5.8 pg/mL, n=3, p≤0.001) by increasing IL-10 transcription. β2AR blockade with propranolol completely inhibited epinephrine-induced IL-10 production (propranolol + LPS = 27.6 ± 0.8 pg/mL vs. epinephrine + propranolol + LPS = 26.1 ± 1.0 pg/mL, n=3). PKA inhibition with H89 and Rp-cAMP did not reduce epinephrine-induced IL-10 production. In contrast, inhibition of adenylate cyclase with SQ22536 significantly decreased epinephrine-induced IL-10 production by 52% (p≤0.01). These results suggest that epinephrine increases IL-10 cytokine production by a cAMP-dependent, PKA-independent mechanism.
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