The effect of Diosmin on the blood proteome in a rat model of venousthrombosis
The effect of Diosmin on the blood proteome in a rat model of venousthrombosis
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地奥司明对静脉血栓形成大鼠模型血液蛋白质组的影响
DOI：
10.1016/j.ijbiomac.2017.06.045
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发表时间：
2017
影响因子：
8.2
通讯作者：
Lei Yao
中科院分区：
文献类型：
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作者：
Lei Yao
薯蓣皂苷是薯蓣皂苷（3,5,7三羟基-4甲氧基自由基类黄酮）的苷元部分，是一种天然存在的黄酮类苷，其抗血栓作用已在大鼠体内进行了研究。本研究采用二维凝胶电泳（2-DE）方法，观察地奥司明处理前后Wistar大鼠静脉血栓形成蛋白的变化，并探讨其对人血管内皮细胞生长的影响。先用大孔吸附树脂（AB-8）和聚酰胺进行层析纯化，用溶剂萃取法分离得到白藜芦醇，用高效液相色谱法纯化纯度为98%的白藜芦醇。Wistar大鼠分为对照组、模型组和预防组。并解剖其静脉血栓组织段，进行组织病理学检查和血浆蛋白C （PC）检测。接下来，对样品进行蛋白质组学分析。用二维凝胶电泳（2-DE）对三组低丰度蛋白进行分离。2-DE分析结果显示，三组间存在191个蛋白点的差异表达。在蛋白质鉴定方面，我们选择了6个位点进行基质辅助激光解吸/电离飞行时间质谱（MALDI-TOF-MS）检测，然后在NCBI数据库中进行同源性检索。考虑到这些蛋白的特性，我们认为CEP350与纺锤体组装有关。此外，我们使用Lipofectamine 2000用CEP350 siRNA转染HUVECs，并使用实时聚合酶链反应（RT-PCR）评估沉默程度。用微管蛋白追踪红对细胞进行免疫荧光染色，用激光扫描共聚焦显微镜分析细胞结构变化。我们得出结论，CEP350耗竭降低了微管稳定性。Dosmin可通过调节cep350调节纺锤体的不均匀分布，保护机体免于静脉曲张。
tDiosmin is the aglycone moiety of diosmetin (3,5,7 trihydroxy-4methoxy radicals flavonoids), a naturallyoccurring flavone glycoside, whose antithrombotic effect was studied in rats. This study was designedto find the protein changes of venous thrombosis in Wistar rats comparing conditions with and withoutDiosmin treatment by two-dimensional gel electrophoresis (2-DE), and investigate the effect of a crucialprotein known as CEP350 on human vascular endothelial cell growth.Through prior chromatographic purification with macroporous absorption resin (AB-8) and polyamide,Diosmin was isolated from Galium verum L. by solvent extraction, then purified to 98% purity using HPLC.Wistar rats were divided into control group, model group, and prevention group. And their venous throm-bosis tissue segments were dissected and prepared for histopathological examination and detection ofplasma protein C (PC). Next, proteomic analysis was performed with the samples. Low-abundance pro-teins of the three groups were separated by two-dimensional gel electrophoresis (2-DE). 2-DE analysisrevealed that 191 protein spots were differentially expressed among those three groups. For proteinidentification, we selected six spots to use matrix assisted laser desorption/ionization-time of flight-massspectrometry (MALDI-TOF-MS) detection, and then do the homology search in NCBI database. Consider-ing characteristics of these proteins, we proposed CEP350 is related to spindle assembly. Furthermore, weused Lipofectamine 2000 to transfect HUVECs with CEP350 siRNA and evaluated the extent of silencingusing real time–polymerase chain reaction (RT-PCR). Cells were stained for immunofluorescence withtubulin-tracker red, and structural changes were analyzed using laser scanning confocal microscope.We concluded that CEP350 depletion decreased microtubule stability. Dosmin could modulate theassemble of spindle from unevenly distributing and protect body from varicose veins by regulatingCEP350.