Department of Internal Medicine, University of Debrecen, Debrecen, 4032, Hungary.
Department of Cardiology and Cardiac Surgery, University of Debrecen, Debrecen, 4032, Hungary.
Lab Invest. 2020 Jul;100(7):986-1002. doi: 10.1038/s41374-020-0403-x. Epub 2020 Feb 13.
The lysis of red blood cells was shown to occur in human ruptured atherosclerotic lesions and intraventricular hemorrhage (IVH) of the brain. Liberated cell-free hemoglobin was found to undergo oxidation in both pathologies. We hypothesize that hemoglobin-derived peptides are generated during hemoglobin oxidation both in complicated atherosclerotic lesions and IVH of the brain, triggering endothelial cell dysfunction. Oxidized hemoglobin and its products were followed with spectrophotometry, LC-MS/MS analysis and detection of the cross-linking of globin chains in complicated atherosclerotic lesions of the human carotid artery and the hemorrhaged cerebrospinal liquid of preterm infants. The vascular pathophysiologic role of oxidized hemoglobin and the resultant peptides was assessed by measuring endothelial integrity, the activation of endothelial cells and the induction of proinflammatory genes. Peptide fragments of hemoglobin (VNVDEVGGEALGRLLVVYPWTQR, LLVVYPWTQR, MFLSFPTTK, VGAHAGEYGAELERMFLSFPTTK, and FLASVSTVLTSKYR) were identified in ruptured atherosclerotic lesions and in IVH of the human brain. Fragments resulting from the oxidation of hemoglobin were accompanied by the accumulation of ferryl hemoglobin. Similar to complicated atherosclerotic lesions of the human carotid artery, a high level of oxidized and cross-linked hemoglobin was observed in the cerebrospinal fluid after IVH. Haptoglobin inhibited hemoglobin fragmentation provoked by peroxide. The resultant peptides failed to bind haptoglobin or albumin. Peptides derived from hemoglobin oxidation and ferryl hemoglobin induced intercellular gap formation, decreased junctional resistance in the endothelium, and enhanced monocyte adhesion to endothelial cells. Enhanced expression of TNF and the activation of NLRP3 and CASP1 followed by the increased generation of IL-1β and nuclear translocation of the NF-κβ transcription factor occurred in response to hemoglobin-derived peptides, and ferryl hemoglobin in endothelium was upregulated in both pathologies. We conclude that the oxidation of hemoglobin in complicated atherosclerotic lesions and intraventricular hemorrhage of the brain generates peptide fragments and ferryl hemoglobin with the potential to trigger endothelial cell dysfunction.
红细胞裂解被证明发生在人类破裂的动脉粥样硬化病变和脑室内出血 (IVH) 中。在这两种病理情况下，游离的无细胞血红蛋白被发现发生氧化。我们假设在复杂的动脉粥样硬化病变和脑室内出血中，血红蛋白氧化过程中会产生血红蛋白衍生肽，从而触发内皮细胞功能障碍。使用分光光度法、LC-MS/MS 分析和检测人颈动脉复杂动脉粥样硬化病变和早产儿出血性脑脊液中球蛋白链的交联，跟踪氧化血红蛋白及其产物。通过测量内皮完整性、内皮细胞激活和促炎基因的诱导，评估氧化血红蛋白和由此产生的肽的血管病理生理作用。在破裂的动脉粥样硬化病变和人脑 IVH 中鉴定出血红蛋白的肽片段 (VNVDEVGGEALGRLLVVYPWTQR、LLVVYPWTQR、MFLSFPTTK、VGAHAGEYGAELERMFLSFPTTK 和 FLASVSTVLTSKYR)。血红蛋白氧化产生的片段伴随着高铁血红蛋白的积累。与人颈动脉复杂动脉粥样硬化病变相似，在 IVH 后观察到脑脊液中氧化和交联血红蛋白水平升高。触珠蛋白抑制过氧化物引起的血红蛋白片段化。所得肽未能与触珠蛋白或白蛋白结合。来自血红蛋白氧化和高铁血红蛋白的肽诱导细胞间间隙形成，降低内皮细胞的连接电阻，增强单核细胞黏附内皮细胞。针对血红蛋白衍生肽和内皮细胞中铁氧血红蛋白，观察到 TNF 表达增强、NLRP3 和 CASP1 激活以及随后 IL-1β 的核转位和 NF-κβ 转录因子的增加，在这两种病理情况下，内皮细胞中铁氧血红蛋白的表达均上调。我们得出结论，血红蛋白在复杂的动脉粥样硬化病变和脑室内出血中的氧化会产生具有潜在触发内皮细胞功能障碍的肽片段和高铁血红蛋白。
