Persistent endotoxin exposure in macrophages elicits an immunometabolic profile susceptible to NF-κB and p53 perturbation

The prolonged exposure of macrophages to endotoxin occurs in various chronic inflammatory conditions, including long-term inflammation triggered by particle-releasing implanted biomaterials, progressive pulmonary obstructive conditions, chronic bronchitis, colitis and hepatitis characterized by sustained exposure to gut-derived lipopolysaccharide (LPS). However, immune cellular metabolism in models of persistent inflammation remains undercharacterized, especially when compared to the well-studied immunometabolic changes following short-term LPS exposure. Here, we demonstrate that persistent LPS stimulation induces increased oxygen consumption alongside reduced levels of ATP and elevated glycolysis, with a mixed inflammatory profile indicative of endotoxin tolerance in primary bone marrow-derived macrophages. Pharmacological blockade of complex I of the mitochondrial electron transport chain lowered oxygen consumption and mitochondrial ROS by reducing NF-κB signaling. Surprisingly, NF-κB inhibition at TAK1 or IκBα phosphorylation, and p53 activation, reduced both oxygen consumption and glycolysis with minimal mitochondrial ROS impact. The absence of elevated mitochondrial membrane potential in prolonged LPS exposure suggests mitochondrial ROS production is likely reverse electron transport-independent, supported by the lack of effect of a mitochondrial depolarizer on oxygen consumption. These findings reveal a distinct immunometabolic phenotype in macrophage endotoxin tolerance, extending our understanding of immunobiology in chronic inflammatory diseases.