HMGB1 Triggers Astrocyte Inflammation, Leading to Neurotoxicity through the TLR2 and 4 Pathways

Abstract

Background: Astrocytes are the most abundant cell type in the central nervous system (CNS) and play a crucial role in neuroinflammatory responses to damage and disease in the CNS. High mobility group box 1 (HMGB1) is an immuno-adjuvant factor that acts as a ligand for toll-like receptor (TLR)2/4 and is the receptor for advanced glycation end products (RAGE). It is associated with chronic neurodegeneration and neuroinflammation. We conducted a study to investigate the impact of HMGB1 on astrocytic cytotoxicity and inflammatory evolution. Method: We cultured rat astrocytes and treated them with lipopolysaccharide (LPS) (TLR4 ligand), zymosan (TLR2 ligand), and HMGB1 or left them untreated as a control. Then, we blocked the activity of TLR2 and TLR4 with monoclonal antibodies. Using Luminex, we conducted measurements of cytokines interleukin-6 (IL-6), interleukin-1β (IL-1β), and interleukin-10 (IL-10) at various time intervals. Also, we performed a Flow Cytometry analysis to check for any signs of apoptosis or necrosis in the collected cells. Finally, we tested the effect of astrocytes conditioned medium on neurons after TLRS agonists and antagonists treated them. Results: Our findings showed that the rate of apoptosis was highest after stimulation by HMGB1 (38.8%). However, the antagonists of TLR4 (25.8%) and TLR2 (31.2%) slightly reversed the rate. HMGB1 activated astrocytes to express higher levels of IL-6 and IL-1β, like bacterial ligands. Conversely, the anti-inflammatory cytokine IL-10 was reduced after prolonged interaction with HMGB1. Our data also showed that TLRs antagonists reacted differently according to time intervals for blocking cytokine release after astrocytes were stimulated with HMGB1. Conclusions: HMGB1 exhibits diversity in astrocyte activation. It can trigger TLR pathways, resulting in inflammation and subsequent neurotoxicity, or it can be involved in tissue regeneration at some stage.