Burn injury sensitizes rat Kupffer cells via mechanisms dependent on gut-derived endotoxin.

BACKGROUND

Bacterial translocation occurs after thermal injury in association with intestinal barrier loss. Recently, we found that sensitization of Kupffer cells involved gut-derived endotoxin; therefore, the purpose of this work was to study the mechanisms of sensitization of Kupffer cells in burn injury.

METHODS

Rats received a 30% body surface area full-thickness steam burn 24 h before experiments. Serum alanine aminotransferase (ALT) was measured to assess liver damage, and plasma endotoxin in the portal vein were measured. Kupffer cells were isolated 24 h after the burn. Intracellular calcium ([Ca2+]i) in Kupffer cells was measured using a microspectrofluorometer with the fluorescent indicator, fura-2, and tumor necrosis factor (TNF)-alpha was measured by enzyme-linked immunosorbent assay (ELISA).

RESULTS

Lipopolysaccharide (LPS)-induced mortality was increased by burn treatment. This increase was blocked by gadolinium chloride, a Kupffer-cell toxicant. Accordingly, Kupffer cells were involved in this system. The LPS-induced increase of ALT was upregulated by the burn injury. This increase was blocked by pretreatment with antibiotics. Endotoxin levels were increased to almost 300 pg/ml (normal, <20 pg/ml) in the portal veins of rats that received a burn. This increase was blunted by antibiotics. In Kupffer cells isolated from untreated control rats, [Ca2+]i increased to 82+/-7 nM after the addition of LPS (100 ng/ml). Levels were elevated twofold over control levels in the cells from rats with burn (174+/-15 nM). In addition, TNF-alpha production by Kupffer cells isolated from rats with burn was increased fourfold over the based level. Sterilization of the gut with antibiotics completely blocked all effects of the burn on [Ca2+]i and TNF-alpha release.

CONCLUSIONS

Kupffer cells isolated from rats with burn exhibited sensitization to LPS, involving gut-derived endotoxin. It is concluded that burns sensitize Kupffer cells to LPS via mechanisms that are dependent on gut-derived endotoxin.