Inhibitory effects of spermidine on lipopolysaccharide‑induced inflammation in RAW 264.7 cells

Spermidine (SPD) is a naturally occurring polyamine with anti-inflammatory and antioxidant properties. Given the pivotal role of macrophages in inflammatory pathogenesis (such as rheumatoid arthritis and autoimmune encephalomyelitis), the present study hypothesized that SPD exerts its anti-inflammatory effects by suppressing macrophage polarization. Briefly, RAW 264.7 cells were activated using lipopolysaccharide (LPS) and were treated with a vehicle control (complete DMEM) or varying SPD concentrations. The ratio of M1 and M2 macrophages in each group was determined using flow cytometry. Inflammatory gene expression and cytokine levels were determined using reverse transcription quantitative PCR and ELISA. The levels of NF-κB pathway-associated proteins were measured through western blotting. The findings revealed that the proportion of M1 cells gradually decreased with increasing SPD concentrations. In LPS-stimulated RAW 264.7 cells, SPD markedly decreased the expression of M1 marker genes and notably increased that of M2 marker genes. Furthermore, SPD decreased IL-6 levels and increased IL-10 levels. In addition, the levels of phosphorylated (p)-p65 and p-IκBα, which are NF-κB pathway-associated proteins, were markedly decreased after 24 h of SPD treatment. Overall, SPD treatment inhibited LPS-induced M1 polarization, reduced pro-inflammatory gene/cytokine expression and enhanced anti-inflammatory markers, potentially through NF-κB pathway modulation.