Ethanol extract of Liriodendron tulipifera leaves displays anti-inflammatory activity by suppressing the Syk/Src/NF-κB pathway.

Liriodendron tulipifera L. (family Magnoliaceae), native to North America, has traditionally been used to treat inflammation-related diseases, such as rheumatism, catarrh, stomachache, dysentery, or headache. Historical records also suggest its strong vermifuge activity against parasites. Although L. tulipifera has been reported to exhibit notable anti-inflammatory activity, its underlying molecular mechanisms remain unclear. This study aimed to comprehensively evaluate the anti-inflammatory effects of an ethanol extract of L. tulipifera (Lt-EE) through in vitro and in vivo experiments and to elucidate its molecular mechanisms and target proteins. RAW264.7 cells and HEK293T cells were used to analyze cellular activities and determine the molecular targets of Lt-EE. Griess reagent assays were used to evaluate the influence of Lt-EE on nitric oxide (NO) generation. In addition, western blotting was conducted to analyze the in vitro effects of Lt-EE at the protein level while real-time polymerase chain reaction (PCR) and reverse transcription PCR were used to assess the in vitro effects of Lt-EE at the transcript level. Luciferase reporter assays and cellular thermal shift assays (CETSAs) were employed to confirm the molecular target(s) of Lt-EE. To evaluate the in vivo effects, Lt-EE was administered at 50 and 150 mg/kg intraperitoneally in an LPS-induced peritonitis model, and at 100 and 150 mg/kg orally in an HCl/EtOH-induced acute gastritis mouse model. Lt-EE was not cytotoxic but effectively suppressed the production of NO. mRNA expression levels of the pro-inflammatory cytokines iNOS, IL-6, and IL-1β were decreased by Lt-EE in a concentration-dependent manner. Moreover, the overall pathway of Lt-EE was revealed by western blotting. Through CETSA, the targets of Lt-EE were confirmed to be Syk and Src. Lt-EE significantly ameliorated inflammation in both LPS-induced peritonitis and HCl/EtOH-induced gastritis models. An ethanol extract of L. tulipifera leaves showed significant anti-inflammatory effects in vitro and in vivo by inhibiting Syk and Src and the NF-κB signaling pathway. These results indicate that Lt-EE is a potent anti-inflammatory agent.