Background: Closed soft-tissue trauma leads to activation of the coagulation cascade and is often complicated by systemic inflammation and infection. Previous investigations have shown potent anti-inflammatory properties of antithrombin. We herein report on the action of antithrombin on skeletal muscle injury in experimental endotoxemia. Materials and Methods: By using a pneumatically driven computer-controlled impact device, closed soft-tissue trauma was applied on the left hind limb of pentobarbital-anesthetized rats. Six hours later, endotoxemia was induced by intraperitoneal injection of Escherichia coli]ipopolysaccharide. An equivalent volume of physiological saline was given in controls. At the same time point, treatment of animals was started by intravenous injection of antithrombin (250 IU/kg body weight) or vehicle solution. Twenty-four hours after trauma, the extensor digitorum longus muscle was microsurgically exposed and analyzed by means of high-resolution multifluorescence microscopy. Results: Traumatic soft-tissue injury with additional endotoxemia was characterized by nutritive perfusion failure (functional capillary density: 379 +/- 20 cm/cm(2)), tissue hypoxia (nicotinamide adenine dinucleotide autofluorescence: 77 +/- 4 aU), and enhanced leukocyte-endothelial cell interaction (773 +/- 35 cells/mm(2)). Therapeutic intervention with antithrombin 6 hrs after trauma restored nutritive perfusion and tissue oxygenation (functional capillary density: 469 +/- 22 cm/cm(2); nicotinamide adenine dinucleotide autofluorescence: 61 +/- 5 aU p < 0.05]) and reduced inflammatory leukocyte adherence (237 +/- 20 cells/mm(2) p < 0.05]) toward values found in nontraumatized controls (functional capillary density: 573 +/- 13 cm/cm(2); nicotinamide adenine dinucleotide autofluorescence: 56 +/- 2 aU; leukocyte adherence: 204 +/- 20 cells/mm(2)). Conclusion: Antithrombin ameliorates microcirculatory dysfunction and tissue injury in traumatized animals during endotoxemia. Furthermore, a reduced inflammatory cell response helps to prevent leukocyte-dependent secondary tissue injury. (Crit Care Med 2013; 41:867-873)