Noradrenaline Synergistically Enhances <em>Porphyromonas gingivalis </em>LPS and OMV-Induced Interleukin-1β Production in BV-2 Microglia Through Differential Mechanisms

Infection with Porphyromonas gingivalis (Pg), which is a major periodontal pathogen, causes a large number of systemic diseases based on chronic inflammation such as diabetes, rheumatoid arthritis and Alzheimer’s disease. However, it remains unclear how Pg can augment local systemic immune and inflammatory responses, because Pg and its virulence factor lipopolysaccharide (LPS) have low biological activities. There is a strong association between depression and elevated levels of inflammation. Noradrenaline (NA) is a key neurotransmitter that modulates microglial activation during stress conditions. In this study, we have thus investigated the regulatory mechanisms of NA on the production of interleukin-1β (IL-1β) by microglia following stimulation with Pg virulence factors, LPS and outer membrane vesicles (OMVs). NA significantly enhanced the mRNA level, promoter activity, and protein level of IL-1β up to 20-fold in BV-2 microglia following treatment with Pg LPS and OMVs in a dose-dependent manner. Pharmacological studies have suggested that NA synergistically augments the responses induced by Pg LPS and OMVs through different mechanisms. AP-1 is activated by the β2 adrenergic receptor (Aβ2R)-mediated pathway. NF-κB, which is activated by the Pg LPS/Toll-like receptor 2-mediated pathway, is required for the synergistic effect of NA on the Pg LPS-induced IL-1β production by BV-2 microglia. Co-immunoprecipitation combined with western blotting and the structural models generated by AlphaFold2 suggested that cross-coupling of NF-κB p65 and AP-1 c-Fos transcription factors enhances the binding of NF-κB p65 to the IκB site, resulting in the synergistic augmentation of the IL-1 promoter activity. In contrast, OMVs phagocytosed by BV-2 microglia and then activated the TLR9/p52/RelB-mediated pathway. The Aβ2R/Epac-mediated pathway, which promotes phagosome maturation, may be responsible for the synergistic effect of NA on the OMV-induced production of IL-1β in BV-2 microglia.