Synergistic Activation of TLR Adaptors MyD88 and TRIF Induces Inflammatory Cell Death in Infection 2260795

Abstract Introduction Severe infections often cause life-threatening inflammation, multi-organ failure, and death. During infection, pathogens carry multiple pathogen-associated molecular patterns (PAMPs) such as LPS, flagellin, nucleic acids, and lipoproteins which activate more than one pattern-recognition receptor (PRR). These PRRs trigger cytokine secretion and inflammatory cell death to control pathogen load; however, a dysregulated immune response can lead to excessive inflammation and lethality. While earlier studies have primarily focused on single PAMP responses, cells in vivo are exposed to multiple microbial ligands simultaneously. Methods Primary bone marrow-derived macrophages (BMDMs) were stimulated with PAMPs, and real-time imaging of cell death was performed using the IncuCyte system. Microarray analysis was conducted to examine differential gene expression. Results We found that specific PAMP combinations induce lytic cell death, while other combinations do not. Toll-like receptors (TLRs) and their adaptor molecules, MyD88 and TRIF, functioned as upstream regulators that activated inflammatory cell death. In addition, signaling through the interferon-α/β receptor (IFNAR) and interferon regulatory factor 1 (IRF1) was essential for the induction of inflammatory cell death. We also performed microarray analysis to understand the upregulated and downregulated genes in PAMP combinations compared with the untreated control. Through this analysis, we identified significant differences in specific molecules, and we performed genetic, functional validation to show that these molecules directly modulated the induction of inflammatory cell death. Conclusion These findings identify key mediators of lytic cell death and suggest that targeting these pathways can be used to mitigate inflammation and lethality during infections. Funding Source This research was supported by NIH grants AI101935, AI124346, AI160179, AR056296, and CA253095 and the American Lebanese Syrian Associated Charities to Dr. Kanneganti. Topic Categories Innate Immune Responses and Host Defense: Molecular Mechanisms (INM)