Toll-like receptor 4 contributes to microvascular inflammation and barrier dysfunction in thermal injury

Systemic and microvascular inflammation plays a key role in the development of multiple organ failure after infection, sepsis, and traumatic injury. Toll-like receptors (TLRs) regulate host responses to pathogens and sterile, injury-associated inflammatory responses. We investigated whether TLR-4 contributes to microvascular dysfunction during thermal injury in vivo in anesthetized wild-type or TLR-4 (-/-) mice receiving either a 25% total body surface area full-thickness scald burn or sham treatment on the dorsal skin. Using intravital microscopy, we assessed the hemodynamics and leukocyte dynamics in the mesenteric microvasculature as representative of the splanchnic microcirculation at a site remote from the burn wound. The transvascular flux of fluorescein isothiocyanate-albumin across mesenteric venules was measured as an indicator of microvascular permeability. Furthermore, cultured microvascular endothelial cell models were used to evaluate the endothelial-specific mechanisms involved in TLR-4-mediated barrier dysfunction. The results showed significantly elevated microvascular permeability in wild-type mice after burn, whereas this response was markedly attenuated in TLR-4 (-/-) mice. Burn injury also increased leukocyte adhesion in mesenteric venules of wild-type mice, and a blunted leukocyte response was seen in the TLR-4 mice. Treatment of endothelial cell monolayers with burn plasma induced a rapid reduction in the transendothelial electrical resistance measured by electric cell-substrate impedance sensing, indicative of endothelial cell-cell adhesive barrier dysfunction. Reducing expression of TLR-4 with siRNA treatment attenuated this response. Taken together, these data indicate that TLR-4 plays an important role in microvascular leakage and leukocyte adhesion under the inflammatory condition associated with nonseptic thermal injury.