TLR4/TNFR1 blockade suppresses STAT1/STAT3 expression and increases SOCS3 expression in modulation of LPS-induced macrophage responses

Due to the urgent need to create appropriate treatment techniques, which are currently unavailable, LPS-induced sepsis has become a serious concern on a global scale. The primary active component in the pathophysiology of inflammatory diseases such as sepsis is the Gram-negative bacterial lipopolysaccharide (LPS). LPS interacts with cell surface TLR4 in macrophages, causing the formation of reactive oxygen species (ROS), TNF-alpha, IL-1(3 and oxidative stress. It also significantly activates the MAPKs and NF-kappa B pathway. Excessive production of proinflammatory cytokines is one of the primary characteristic features in the onset and progression of inflammation. Cytokines mainly signal through the JAK/STAT pathway. We hypothesize that blocking of TLR4 along with TNFR1 might be beneficial in suppressing the effects of STAT1/STAT3 due to the stimulation of SOCS3 proteins. Prior to the LPS challenge, the macrophages were treated with antibodies against TLR4 and TNFR1 either individually or in combination. On analysis of the macrophage populations by flowcytometry, it was seen that receptor blockade facilitated the phenotypic shift of the M1 macrophages towards M2 resulting in lowered oxidative stress. Blocking of TLR4/TNFR1 upregulated the SOCS3 and mTOR expressions that enabled the transition of inflammatory M1 macrophages towards the anti-inflammatory M2 phenotype, which might be crucial in curbing the inflammatory responses. Also the reduction in the production of inflammatory cytokines such as IL-6, IL-1(3 due to the reduction in the activation of the STAT1 and STAT3 molecules was observed in our combination treatment group. All these results indicated that neutralization of both TLR4 and TNFR1 might provide new insights in establishing an alternative therapeutic strategy for LPS-sepsis.