Antithrombin is expected to modulate both prothrombotic and proinflammatory reactions in sepsis; vascular endothelium is the primary target. In the present study, we sought to evaluate the protective effects of a newly developed fucose-deficient recombinant antithrombin. Endothelial cells were treated in vitro with histone H4 to induce cellular damage. Low to high doses of either plasma-derived antithrombin or recombinant thrombomodulin were used as treatment interventions. Morphological change, apoptotic rate, cell viability, cell injury, and syndecan-4 level in the medium were evaluated. Immunofluorescent staining with anti-syndecan-4 was also performed. Both types of antithrombin reduced cellular damage and apoptotic cell death. Both plasma-derived and recombinant antithrombin improved cell viability and reduced cellular injury when administered at a physiological concentration or higher. Syndecan-4 staining became evident after treatment with histone H4, and both antithrombins suppressed the staining intensity at similar levels. The syndecan-4 level in the medium was significantly decreased by both antithrombins. None of the indicators showed a significant difference between plasma-derived and recombinant antithrombin. In conclusion, both recombinant and plasma-derived antithrombin can protect vascular endothelial cells. Recombinant antithrombin may represent a useful new therapeutic agent for sepsis-associated vascular damage.