Okamoto Takayuki, Hattori Mai, Katsube Yukiko, Ota Junichi, Asanuma Kunihiro, Usuda Haruki, Wada Koichiro, Suzuki Koji, Nikai Tetsuro
Department of Pharmacology, Faculty of Medicine, Shimane University, 89-1 Enya-cho, Izumo-city, Shimane 693-8501, Japan.
Department of Anesthesiology, Faculty of Medicine, Shimane University, 89-1 Enya-cho, Izumo city, Shimane 693-8501, Japan.
Biochim Biophys Acta Mol Cell Res. 2025 Feb;1872(2):119891. doi: 10.1016/j.bbamcr.2024.119891. Epub 2024 Dec 15.
Thrombomodulin is predominantly expressed on vascular endothelial cells and modulates endothelial cell functions by interacting with multiple ligands. The specific thrombomodulin receptor or cofactor active on the endothelial cell surface remains elusive. This study aims to identify interacting partners of thrombomodulin on endothelial cells. Here, using a liquid chromatograph-tandem mass spectrometer, hornerin was identified as a candidate protein. We then investigated hornerin protein and mRNA expression in endothelial cells. Hornerin protein was detected in the mouse endothelium of the aorta and lung. Both human- and mouse-cultured endothelial cells expressed hornerin mRNA and protein. Moreover, immunoprecipitation analysis suggested the direct protein interaction between thrombomodulin and hornerin. Lipopolysaccharides administration increased serum hornerin concentrations in mice and reduced hornerin protein levels on the surface of cultured endothelial cells as same as thrombomodulin protein. Thrombomodulin-targeting siRNA decreased not only thrombomodulin protein levels but also hornerin protein levels in cultured endothelial cells. Thrombomodulin- or hornerin-targeting siRNA impaired tube formation and leukocyte adhesion to endothelial cells. Our findings reveal that hornerin is located on vascular endothelial cells in the presence of thrombomodulin and suggest that endothelial thrombomodulin and hornerin may interact, which may play an important role in endothelial cell functions such as vascular inflammation and angiogenesis.
血栓调节蛋白主要表达于血管内皮细胞，并通过与多种配体相互作用来调节内皮细胞功能。在内皮细胞表面具有活性的特定血栓调节蛋白受体或辅助因子仍不明确。本研究旨在鉴定内皮细胞上血栓调节蛋白的相互作用伙伴。在此，使用液相色谱 - 串联质谱仪，角蛋白被鉴定为一种候选蛋白。然后我们研究了角蛋白在内皮细胞中的蛋白和mRNA表达。在小鼠主动脉和肺的内皮中检测到角蛋白。人源和鼠源培养的内皮细胞均表达角蛋白mRNA和蛋白。此外，免疫沉淀分析表明血栓调节蛋白与角蛋白之间存在直接的蛋白相互作用。给予脂多糖可增加小鼠血清角蛋白浓度，并降低培养的内皮细胞表面的角蛋白水平，与血栓调节蛋白水平降低情况相同。靶向血栓调节蛋白的小干扰RNA不仅降低了培养的内皮细胞中血栓调节蛋白的水平，也降低了角蛋白的水平。靶向血栓调节蛋白或角蛋白的小干扰RNA损害了内皮细胞的管腔形成和白细胞黏附。我们的研究结果表明，在有血栓调节蛋白存在的情况下，角蛋白定位于血管内皮细胞，提示内皮血栓调节蛋白和角蛋白可能相互作用，这可能在血管炎症和血管生成等内皮细胞功能中发挥重要作用。
