MD‐2 as the Target of a Novel Small Molecule, L6H21, in the Attenuation of LPS‐induced Inflammatory Response and Sepsis

Abstract

BackgroundMyeloid differentiation 2 (MD‐2) recognizes LPS, which is required for TLR4 activation, and represents an attractive therapeutic target for severe inflammatory disorders. We previously found that a chalcone derivative, L6H21, could inhibit LPS‐induced overexpression of TNF‐α and IL‐6 in macrophages.MethodsThe binding affinity of L6H21 to MD‐2 protein was analyzed using computer docking, surface plasmon resonance analysis, ELISA, fluorescence measurements, and flow cytometric analysis. The effects of L6H21 on MAPK and NF‐κB signaling were determined using EMSA, fluorescence staining, western blotting, and immunoprecipitation. The anti‐inflammatory effects of L6H21 were confirmed using ELISA and RT‐qPCR in vitro. Finally, the anti‐inflammatory effects of L6H21 in septic C57BL/6 mice were evaluated.ResultsCompound L6H21 was inserted into the hydrophobic region of MD‐2 pocket, forming hydrogen bonds with Arg90 and Tyr102 in the MD‐2 pocket. In vitro, L6H21 subsequently suppressed MAPK phosphorylation, NF‐κB activation, and cytokine expression in macrophages stimulated by LPS. In vivo, L6H21 pretreatment improved survival, protected lung injury, decreased serum and hepatic cytokine level in mice subjected to LPS. In addition, the MD‐2 gene knockout led to universal protection from LPS‐induced septic shock in mice.ConclusionsThis work identified a new chalcone derivative, L6H21, as a potential candidate in the treatment of sepsis; more importantly, the data confirmed that MD‐2 is an important therapeutic target against inflammatory disorders.