METHODS: The attenuating effects of syringin on the productions of prostaglandin E2 (PGE2), cyclooxygenase-2 (COX-2), interleukin-1β (IL-1β), and tumor necrosis factor-α (TNF-α), and the expressions of signaling molecules of the signaling pathways were investigated by using ELISA, Western blot, and qRT-PCR.

RESULTS: Syringin downregulated the NF-κB, MAPKs, and PI3K-Akt signal networks by significantly reducing PGE2 production in the macrophages via suppression of COX-2 gene and protein expression levels. It also reduced TNF-α and IL-1β secretion and their mRNA expression, suppressed phosphorylation of NF-κB (p65), IKKα/β, and IκBα, and restored ability of IκBα to degrade. Syringin dose-dependently attenuated Akt, p38 MAPKs, JNK, and ERK phosphorylation. Also, the expression of corresponding upstream signaling molecules toll-like receptor 4 (TLR4) and myeloid differentiation primary response gene 88 (MyD88) were down-regulated in response to syringin treatment.

METHODS: Molecular docking and molecular dynamics simulations were used for EGFR, p38, ERK1/2, and AKT. The effects of berberine and lapatinib on MAPK and PI3K pathways in MDA-MB231 and MCF-7 cells were evaluated using immunoflorescence assays, and the amounts of phosphorylated kinases were compared to total kinases after treating with different concentrations of berberine.

RESULTS: Simulations showed berberine accurately interacted with EGFR, AKT, P38, and ERK1/2 active sites in silico (scores = -7.57 to -7.92 Kcal/mol) and decreased the levels of active forms of corresponding enzymes in both cell lines; however, berberine binding to p38 showed less stability. Cytotoxicity analysis indicated that MDA-MB231 cells were resistant to berberine compared to MCF-7 cells [72 h IC50 = 50 versus 15 μM, respectively). Also, lapatinib strongly activated AKT but suppressed EGFR in MDA-MB231 cells. The activity of EGFR, AKT, P38, and ERK1/2 were affected by berberine; however, berberine dramatically reduced EGFR and AKT phosphorylation.