PURPOSE: Prostaglandin E2 (PGE2), a pivotal lipid metabolite, plays a dual role in inflammation, manifesting both pro-inflammatory and anti-inflammatory effects, which are significantly influenced by the cellular microenvironment and receptor subtype. Although recent studies have highlighted the anti-inflammatory potential of PGE2, its role in toll-like receptor (TLR)-associated inflammation and the underlying mechanisms have not fully elucidated. Consequently, the primary aim of this study was to assess the anti-inflammatory efficacy of PGE2 in TLR-related inflammation and to elucidate the associated mechanisms. METHODS: In vitro, the anti-inflammatory effect of PGE2 on TLR-related inflammation were investigated by measuring pro-inflammatory cytokine protein and gene levels using ELISA and RT-qPCR, respectively. Western blot analysis was used to explore the corresponding anti-inflammatory signaling pathways. In vivo, the anti-inflammatory effects of PGE2 were further validated using ALI and sepsis models, employing the PGE2 analog 16,16-dimethyl prostaglandin E2 (dmPGE2). RESULTS: The findings revealed that PGE2 inhibited the LPS-induced inflammatory response and activation of the IKK/NF-κB signaling pathway via the EP4 receptor-mediated downstream cAMP/PKA pathway. Additionally, PGE2 analog, dmPGE2, effectively mitigated pathological injury and the inflammatory response in lung tissue of mice subjected to LPS-induced ALI and sepsis. Notably, dmPGE2 suppressed LPS-induced activation of the IKK/NF-κB signaling pathway in lung tissue. CONCLUSION: This study demonstrated that PGE2 can inhibit the IKK/NF-κB signaling pathway through the EP4/cAMP/PKA pathway, thereby alleviating the LPS-induced inflammatory response and providing a protective effect against LPS-induced ALI and sepsis. Consequently, PGE2 holds promise as a candidate for drug development aimed at preventing ALI and sepsis.