Iba Toshiaki, Sasaki Tetsuya, Ohshima Kazutoshi, Sato Koichi, Nagaoka Isao, Thachil Jecko
Department of Emergency and Disaster Medicine, Juntendo University Graduate School of Medicine, Tokyo, Japan.
Nihon Pharmaceutical Co. Ltd., Research Laboratory, Narita, Japan.
TH Open. 2017 Jun 28;1(1):e3-e10. doi: 10.1055/s-0037-1603926. eCollection 2017 Jun.
Antithrombin is a promising option for the treatment of sepsis, and vascular endothelium is an important target for this fatal condition. Here, we aimed to evaluate the protective effects of different glycoforms of antithrombin on histone-induced endothelial cell damage and explore the responsible mechanisms in an experimental model in vitro. Endothelial cells were treated in vitro using histone H4 to induce cellular damage. Various doses of either α- or β-antithrombin were used as treatment interventions, and both cell viability and the levels of lactate dehydrogenase (LDH) in the medium were assessed. Endothelial cell damage was also assessed using microscopic examination and immunofluorescent staining with anti-syndecan-4 and anti-antithrombin antibodies. As a result, both glycoforms of antithrombin significantly improved cell viability when administered at a physiological dose (150 μg/mL). Cellular injury as evaluated using the LDH level was significantly suppressed by β-antithrombin at a supranormal dose (600 μg/mL). Microscopic observation suggested that β-antithrombin suppressed the endothelial cell damage more efficiently than α-antithrombin. β-Antithrombin suppressed the intensity of syndecan-4 staining which became evident after treatment with histone H4, more prominently than α-antithrombin. The distribution of antithrombin was identical to that of syndecan-4. In conclusion, both α- and β-antithrombin can protect vascular endothelial cells from histone H4-induced damage, although the effect was stronger for β-antithrombin. The responsible mechanisms might involve the binding of antithrombin to the glycocalyx on the endothelial surface. These results provide a theoretical basis for the application of antithrombin to the prevention and treatment of sepsis-related endothelial damage.
抗凝血酶是治疗脓毒症的一个有前景的选择，而血管内皮是这种致命病症的一个重要靶点。在此，我们旨在评估不同糖型的抗凝血酶对组蛋白诱导的内皮细胞损伤的保护作用，并在体外实验模型中探索其相关机制。体外使用组蛋白H4处理内皮细胞以诱导细胞损伤。使用不同剂量的α-抗凝血酶或β-抗凝血酶作为治疗干预措施，并评估细胞活力和培养基中乳酸脱氢酶（LDH）的水平。还通过显微镜检查以及用抗syndecan-4和抗抗凝血酶抗体进行免疫荧光染色来评估内皮细胞损伤。结果，当以生理剂量（150μg/mL）给药时，两种抗凝血酶糖型均显著提高了细胞活力。β-抗凝血酶在超正常剂量（600μg/mL）时，能显著抑制用LDH水平评估的细胞损伤。显微镜观察表明，β-抗凝血酶比α-抗凝血酶更有效地抑制内皮细胞损伤。β-抗凝血酶比α-抗凝血酶更显著地抑制了在用组蛋白H4处理后变得明显的syndecan-4染色强度。抗凝血酶的分布与syndecan-4相同。总之，α-抗凝血酶和β-抗凝血酶均可保护血管内皮细胞免受组蛋白H4诱导的损伤，尽管β-抗凝血酶的效果更强。相关机制可能涉及抗凝血酶与内皮表面糖萼的结合。这些结果为抗凝血酶应用于脓毒症相关内皮损伤的预防和治疗提供了理论依据。
