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Recombinant thrombomodulin protects against LPS-induced acute respiratory distress syndrome via preservation of pulmonary endothelial glycocalyx.

ABSTRACT: BACKGROUND AND PURPOSE:Disruption of the endothelial glycocalyx is causally related to microvascular endothelial dysfunction, a characteristic of sepsis-induced acute respiratory distress syndrome (ARDS). Recombinant human thrombomodulin (rhTM) attenuates vascular endothelial injuries, but the underlying mechanism remains elusive. Here, we investigated the structural basis and molecular mechanisms of rhTM effects on vascular endothelial injury in a model of sepsis. EXPERIMENTAL APPROACH:LPS (20 mg·kg-1 ) was intraperitoneally injected into 10-week-old male C57BL6 mice, and saline or rhTM was intraperitoneally injected 3 and 24 h after LPS injection. Using serum and/or lung tissue, histological, ultrastructural, and microarray analyses were performed. KEY RESULTS:Survival rate of rhTM-treated mice was significantly higher than that of control mice 48 h after LPS injection. Serum concentrations of IL-6 and high-mobility group box 1 were lower in the rhTM-treated group than in the control. Injury to the endothelial glycocalyx in pulmonary capillaries was attenuated by rhTM treatment. Gene set enrichment analysis revealed up-regulation of gene sets corresponding to cell proliferation/differentiation and anti-inflammation, such as the TGF-? pathway, and negative regulation of IL-6, upon rhTM treatment. Gene expression of heparan sulfate 6-O-sulfotransferase 1 and endothelial cell-specific molecule 1 (components of the endothelial glycocalyx) was significantly preserved by rhTM treatment, and their protein expression levels were maintained in endothelial cells. CONCLUSION AND IMPLICATIONS:Our findings show that rhTM treatment affected inflammation, cell proliferation/differentiation, and glycocalyx synthesis in serum and lung tissue, subsequently attenuating ARDS caused by endothelial injury.

SUBMITTER: Suzuki K

PROVIDER: S-EPMC7429482 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

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Recombinant thrombomodulin protects against LPS-induced acute respiratory distress syndrome via preservation of pulmonary endothelial glycocalyx.

Suzuki Kodai K Okada Hideshi H Takemura Genzou G Takada Chihiro C Tomita Hiroyuki H Yano Hirohisa H Muraki Isamu I Zaikokuji Ryogen R Kuroda Ayumi A Fukuda Hirotsugu H Nishio Ayane A Takashima Shigeo S Suzuki Akio A Miyazaki Nagisa N Fukuta Tetsuya T Yamada Noriaki N Watanabe Takatomo T Doi Tomoaki T Yoshida Takahiro T Kumada Keisuke K Ushikoshi Hiroaki H Yoshida Shozo S Ogura Shinji S

British journal of pharmacology 20200714 17

<h4>Background and purpose</h4>Disruption of the endothelial glycocalyx is causally related to microvascular endothelial dysfunction, a characteristic of sepsis-induced acute respiratory distress syndrome (ARDS). Recombinant human thrombomodulin (rhTM) attenuates vascular endothelial injuries, but the underlying mechanism remains elusive. Here, we investigated the structural basis and molecular mechanisms of rhTM effects on vascular endothelial injury in a model of sepsis.<h4>Experimental approa ...[more]

PMID: 32497259

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Project description:IntroductionMyocardial dysfunction occurs in patients with sepsis due to vascular endothelial injury. Recombinant human thrombomodulin (rhTM) attenuates vascular endothelial injuries through endothelial glycocalyx (eGC) protection.HypothesisWe hypothesized that rhTM attenuates myocardial dysfunction via the inhibition of vascular endothelial injury during sepsis.MethodsTen-week-old male C57BL6 mice were injected intraperitoneally with 20 mg/kg of lipopolysaccharide (LPS). In rhTM-treated mice, rhTM was injected intraperitoneally at 3 and 24 h after LPS injection. Saline was injected intraperitoneally as control. To assess for eGC injury, intensity score was measured 48 h after the LPS injection. To confirm vascular endothelial injuries, ultrastructural analysis was performed using scanning (SEM) and transmission electron microscopy (TEM).ResultsThe survival rate of the rhTM group at 48 h after LPS injection was significantly higher than that of the control group (68% vs. 17%, p < 0.05). The serum level of troponin I in the rhTM group was lower than that in the control (2.2 ± 0.4 ng/dL vs 9.4 ± 1.1 ng/dL, p < 0.05). The expression of interleukin-6 (IL-6) was attenuated in the rhTM-treated group than in the control (65.3 ± 15.3 ng/mL vs 226.3 ± 19.4 ng/mL, p < 0.05). The serum concentration of syndecan-1, a marker of glycocalyx damage, was significantly decreased 48 h post-administration of LPS in the rhTM-treated group than in the control group. In ultrastructural analysis using SEM and TEM, eGC peeled off from the surface of the capillary lumen in the control. Conversely, the eGC injury was attenuated in the rhTM group. Gene set enrichment analysis revealed that osteomodulin, osteoglycin proline/arginine-rich end leucine-rich repeat protein, and glypican-1, which are proteoglycans, were preserved by rhTM treatment. Their protein expression was retained in endothelial cells.ConclusionrhTM attenuates sepsis-induced myocardial dysfunction via eGC protection.

Project description:IntroductionRecent studies have shown that histones, the chief protein component of chromatin, are released into the extracellular space during sepsis, trauma, and ischemia-reperfusion injury, and act as major mediators of the death of an organism. This study was designed to elucidate the cellular and molecular basis of histone-induced lethality and to assess the protective effects of recombinant thrombomodulin (rTM). rTM has been approved for the treatment of disseminated intravascular coagulation (DIC) in Japan, and is currently undergoing a phase III clinical trial in the United States.MethodsHistone H3 levels in plasma of healthy volunteers and patients with sepsis and DIC were measured using enzyme-linked immunosorbent assay. Male C57BL/6 mice were injected intravenously with purified histones, and pathological examinations were performed. The protective effects of rTM against histone toxicity were analyzed both in vitro and in mice.ResultsHistone H3 was not detectable in plasma of healthy volunteers, but significant levels were observed in patients with sepsis and DIC. These levels were higher in non-survivors than in survivors. Extracellular histones triggered platelet aggregation, leading to thrombotic occlusion of pulmonary capillaries and subsequent right-sided heart failure in mice. These mice displayed symptoms of DIC, including thrombocytopenia, prolonged prothrombin time, decreased fibrinogen, fibrin deposition in capillaries, and bleeding. Platelet depletion protected mice from histone-induced death in the first 30 minutes, suggesting that vessel occlusion by platelet-rich thrombi might be responsible for death during the early phase. Furthermore, rTM bound to extracellular histones, suppressed histone-induced platelet aggregation, thrombotic occlusion of pulmonary capillaries, and dilatation of the right ventricle, and rescued mice from lethal thromboembolism.ConclusionsExtracellular histones cause massive thromboembolism associated with consumptive coagulopathy, which is diagnostically indistinguishable from DIC. rTM binds to histones and neutralizes the prothrombotic action of histones. This may contribute to the effectiveness of rTM against DIC.

Dataset Information

Recombinant thrombomodulin protects against LPS-induced acute respiratory distress syndrome via preservation of pulmonary endothelial glycocalyx.

Publications

Recombinant thrombomodulin protects against LPS-induced acute respiratory distress syndrome via preservation of pulmonary endothelial glycocalyx.

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