Anti‑inflammatory effects of Daehwangmokdantang, a traditional herbal formulation, in lipopolysaccharide‑stimulated RAW 264.7 macrophages

: Daehwangmokdantang (DHMDT) is a traditional polyherbal formulation that has known antidiarrheal and anti-inflammatory activities. However, the underlying mechanisms of these activities are poorly understood. In the present study, the inhibitory effects of DHMDT on the production of proinflammatory mediators and cytokines in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages were investigated. The inhibitory effects of DHMDT on LPS-induced nitric oxide (NO), prostaglandin (PG)E2, tumor necrosis factor (TNF)-α and interleukin (IL)-1β production were examined using Griess reagent and ELISA detection kits. The effects of DHMDT on the expression of inducible NO synthase (iNOS), cyclooxygenase (COX)-2, IL-1β and TNF-α, and their upstream signal proteins, including nuclear factor (NF)-κB, mitogen-activated protein kinases (MAPKs) and RAC-α serine/threonine-protein kinase (Akt), a phosphatidylinositol 3-kinase (PI3K) downstream effector, were investigated using western blotting and immunofluorescence staining. The results revealed the pretreatment with DHMDT significantly inhibited the LPS-induced production of NO, PGE2, TNF-α, and IL-1β, and expression of iNOS, COX-2 TNF-α, and IL-1β, without any significant cytotoxicity. DHMDT also efficiently prevented the translocation of the NF-κB subunit p65 into the nucleus by interrupting the activation of the upstream mediator inhibitor of NF-κB kinase α/β. Furthermore, the anti-inflammatory effects of DHMDT were associated with the suppression of LPS-induced phosphorylation of Akt and MAPKs in RAW 264.7 macrophages. Therefore, the results of the present study indicate that DHMDT exhibited anti-inflammatory activity via the inhibition of proinflammatory mediators and cytokines, in which the inactivation of NF-κB, PI3K/Akt, and MAPKs may be involved. These results suggest that DHMDT may be a potential anti-inflammatory drug candidate.