HMGB1 modulates inflammatory responses in LPS-activated macrophages

Objective: As a late mediator of inflammation, high mobility group box 1 protein (HMGB1) amplifies the inflammatory responses to tissue injury and infection by inducing and extending the production of proinflammaory cytokines. The aim was to investigate whether HMGB1 mediates such effects by affecting the production of anti-inflammatory mediators. Materials and methods: The murine macrophage RAW 264.7 cells were stimulated with 0.5 mu g/ml of LPS and the levels of HMGB1, TNF alpha, IL-1 beta, IL-10 and TGF-beta 1 in the culture supernatants were measured by ELISA. Also, the mRNA expression for IL-10 and TGF-beta 1 was assessed by RT-PCR. Results: LPS induced HMGB I release at 8 h and reached a peak at 48 h. Significant (p < 0.05) production of TNF alpha, IL-1 beta, IL-10, and TGF-beta 1 was seen after 8 h. However, while the levels of TNF alpha and IL-beta remained elevated, IL-10 and TGF-beta 1 release markedly declined by 24 h after stimulation. When cells were stimulated in the presence of conditioned medium derived from a 24 h LPS-stimulated culture, the production of TNF alpha and IL-1 beta was increased while IL-10 and TGF-beta 1 release and mRNA transcripts were decreased. A neutralizing anti-HMGB1 antibody added to the conditioned media reveresed these responses. Conclusions: HMGB1 modulates the inflammatory cascade in activated macrophages by inducing proinflammatory, while suppressing anti-inflammatory responses.