Characterization of the molecular basis of labile heme as a prothrombotic modulator under hemolytic conditions

金属卟啉血红素是一种铁离子的络合物，它广泛存在于体内，是血红蛋白，特别是氧转运蛋白的重要组成部分，广泛存在于体内。不平衡的血红素动态平衡导致血红素的释放，其清除通常受包括血凝素在内的几种蛋白质的调节。然而，血管内血红素水平的增加与血栓形成、细胞毒性、促炎和补体激活效应有关，这些效应是患有溶血性疾病的患者观察到的有害并发症的特征，如镰状细胞病和输血相关溶血。考虑到血栓形成的高患病率和相关的死亡率，令人惊讶的是，不稳定的血红素和血栓前病变之间的机制联系尚未在分子水平上完全阐明。在这一背景下，关于最相关的蛋白质，即凝血系统的组成部分，不同的蛋白质，其中包括第八因子，活化蛋白C，和纤维蛋白原，已被鉴定为与血红素结合或甚至是受血红素调节的蛋白质，而其他蛋白质(凝血酶，FXIIIa)不与血红素结合。许多关于血红素与大多数凝血因子的潜在相互作用的问题仍然悬而未决。因此，这一建议旨在通过生物物理(如UV/Vis，SPR)和结构分析(Rmann)方法来阐明血红素与凝血因子FVIII和von Willebrand因子的瞬时相互作用。此外，还将进行生化和血液学功能分析以及具有高度生理相关性的细胞学方法(例如，血小板/内皮细胞功能分析)。所获得的结果将被合并到机制知识图HEMEKG中，以连接已知的影响，并揭开迄今尚不清楚的血红素的凝血调节作用的方面。综上所述，利用多种光谱方法和功能分析，将在分子、生化、生物信息学和细胞水平上针对血红素的促凝血作用进行研究。这将允许更详细地描述血红素的血栓前性质，并为溶血相关血栓的治疗提供新的视角。

The metalloporphyrin heme is a coordination complex of an iron ion incorporated in a porphyrin ring system, which is widely distributed in the body as an essential part of hemoproteins, especially the oxygen-transport protein hemoglobin. An unbalanced heme homeostasis leads to the release of heme, whose clearance is usually regulated by several proteins including hemopexin. Increased intravascular heme levels, however, have been associated with prothrombotic, cytotoxic, proinflammatory, and complement-activating effects, which characterize the harmful complications observed in patients suffering from hemolytic disorders, such as sickle cell disease and transfusion-related hemolysis. Considering the high prevalence of thrombosis and the associated mortality it is surprising that the mechanistic link between labile heme and prothrombotic effects is not yet completely elucidated on the molecular level. Concerning the most relevant proteins in this context, namely the components of the blood coagulation system, distinct proteins, among them factor VIII, activated protein C, and fibrinogen, have been identified as heme-binding or even heme-regulated proteins, while others (thrombin, FXIIIa) do not bind heme. Many questions regarding a potential interaction of heme with most of the coagulation factors are still open. This proposal thus aims at the elucidation of the transient interaction of heme with the coagulation factor FVIII and von Willebrand factor by means of biophysical (e.g., UV/Vis, SPR) and structural analysis (rRaman) methods. In addition, biochemical and hematological functional assays as well as cellular approaches with high physiological relevance (e.g., platelet/endothelial cell function analysis) will be performed. The results obtained will be incorporated into the mechanistic knowledge graph HemeKG to crosslink the already known effects and to unravel so far unknown aspects of heme’s coagulation-modulating effects. In conclusion, the procoagulant effects of heme will be targeted on different levels, i.e. the molecular, biochemical, bioinformatic, and cellular level applying a variety of spectroscopic methods and functional assays. This will allow for a more detailed characterization of heme’s prothrombotic nature and provide new perspectives for the treatment of hemolysis-associated thrombosis.