Astaxanthin Inhibits Lipopolysaccharide-Induced Immunosuppression in Macrophages

Background: Sepsis is a life-threatening condition characterized by immune dysregulation, with significant mortality occurring during the late immunosuppressive phase. While astaxanthin (AST), a marine-derived tetraterpene, has shown anti-inflammatory potential in sepsis, its role in modulating immunosuppression remains unexplored. Objectives: This study aimed to investigate the immunomodulatory effects of AST on immunosuppressed macrophages in vitro, focusing on its ability to restore inflammatory responses and immune function. Methods: Using lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages, we investigated AST’s immunomodulatory effects during immunosuppression. Cells were pretreated with AST followed by LPS stimulation (LPS1st) and restimulation. Cell viability was assessed using the MTT assay. Inflammatory cytokine tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) production were measured by ELISA. NF-κB activity was determined via ELISA and immunofluorescence (IF). Bactericidal activity was evaluated using an assay with Escherichia coli. Phagocytic capacity was quantified using neutral red uptake. Reactive oxygen species (ROS) and nitric oxide (NO) levels were measured using respective detection kits. Transcriptomic analysis was performed using RNA-Seq, followed by gene set enrichment analysis (GSEA), gene ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. Results: The AST showed no cytotoxicity under both normal and LPS-challenged conditions. In immunosuppressed macrophages, AST pretreatment restored inflammatory responses, specifically TNF-α and IL-6 production, NF-κB activity, bactericidal function, and phagocytic capacity, accompanied by increased ROS and NO production. Transcriptomic analysis revealed AST’s regulation of multiple immune-related pathways, with significant enrichment of both innate and adaptive immune response pathways. Conclusions: Our findings demonstrate AST’s novel dual immunomodulatory properties in addressing both hyperinflammation and immunosuppression in sepsis, suggesting its potential as a therapeutic candidate, particularly for late-stage sepsis treatment.