High-Mobility Group Box-1 Induces Proinflammatory Cytokines Production of Kupffer Cells through TLRs-Dependent Signaling Pathway after Burn Injury

Kupffer cells (KCs) were a significant source of cytokine release during the early stage of severe burns. High mobility group box protein 1 (HMGB1) was recently identified as a new type of proinflammatory cytokine. The ability of HMGB1 to generate inflammatory responses after burn trauma has not been well characterized. KCs were isolated from sham animals and rats with a 30% full-thickness burn, and then were stimulated with increasing concentrations of HMGB1. The levels of Tumor necrosis factor (TNF)-alpha and interleukin (IL)-1 beta in culture supernatant were measured by enzyme-linked immunosorbent assay. Northern blot analysis was performed to detect the expressions of TNF-alpha and IL-1 beta mRNAs. The activities of p38 MAPK and JNK (by Western blot analysis) as well as NF-kappa B (by EMSA) in KCs were also examined. As a result, HMGB1 in vitro upregulated expressions of TNF-alpha and IL-1 beta of KCs in a dose-dependent manner, and HMGB1 promoted KCs from burn rats to produce significantly more TNF-alpha and IL-1 beta proteins than those from sham animals. After harvested from burn rats, KCs were pre-incubated with anti-TLR2 or anti-TLR4 antibody prior to HMGB1 administration. HMGB1 exposure not only significantly increased expressions of TNF-alpha and IL-1 beta mRNAs in KCs from burn rats, but also enhanced activities of p38 MAPK, JNK and NF-kappa B. However, these upregulation events were all reduced by pre-incubation with anti-TLR2 or anti-TLR4 antibody. These results indicate that HMGB1 induces proinflammatory cytokines production of KCs after sever burn injury, and this process might be largely dependent on TLRs-dependent MAPKs/NF-kappa B signal pathway.