Simoni J, Simoni G, Wesson D E, Griswold J A, Feola M
Department of Surgery, Texas Tech University Health Sciences Center, Lubbock 79430, USA.
ASAIO J. 2000 Nov-Dec;46(6):679-92. doi: 10.1097/00002480-200011000-00007.
Chemically modified hemoglobin (Hb) solutions are under current investigation as potential red cell substitutes. Researchers at Texas Tech University have developed a novel free Hb based blood substitute product. This blood substitute is composed of purified bovine Hb cross-linked intramolecularly with o-adenosine-5'-triphosphate and intermolecularly with o-adenosine, and conjugated with reduced glutathione (GSH). In this study, we compared the effects of our novel blood substitute and unmodified (U) Hb, by using allogenic plasma as the control, on human blood components: red blood cells (RBCs), platelets, monocytes (Mo), and low-density lipoproteins (LDLs). The pro-oxidant potential of both Hb solutions on RBCs was examined by the measurement of osmotic and mechanical fragility, conjugated dienes (CD), lipid hydroperoxides (LOOH), thiobarbituric acid reactants (TBAR-S), isoprostanes (8-iso PGF2alpha) and intracellular GSH. The oxidative modification of LDLs was assessed by CD, LOOH, and TBAR-S, and the degree of apolipoprotein (apo) B cross-linking. The effects of Hb on platelets have been studied by monitoring their responses to the aggregation agonists: collagen, ADP, epinephrine, and arachidonic acid. Monocytes were cultured with Hb solutions or plasma and tested for TNF-alpha and IL-1beta release, then examined by electron microscopy. Results indicate that native UHb initiates oxidative stress of many blood components and aggravates inflammatory responses of Mo. It also caused an increase in RBC osmotic and mechanical fragility (p < 0.001). While the level of GSH was slightly changed, the lipid peroxidation of RBC increased (p < 0.001). UHb was found to be a stimulator of 8-iso PGF2alpha synthesis, a potent modulator of LDLs, and an effective potentiator of agonist induced platelet aggregation. Contrarily, our novel blood substitute did not seem to induce oxidative stress nor to increase Mo inflammatory reactions. The osmotic and mechanical fragility of RBCs was similar to that of the control. Such modified Hb failed to alter LDLs, increase the production of 8-iso PGF2alpha, but markedly inhibited platelet aggregation. The effect of this novel blood substitute can be linked with the cytoprotective and anti-inflammatory properties of adenosine, which is used as a cross-linker and surface modifier, and a modification procedure that lowers the hemoglobin pro-oxidant potential.
化学修饰血红蛋白（Hb）溶液目前正作为潜在的红细胞替代物进行研究。德克萨斯理工大学的研究人员开发了一种新型的基于游离Hb的血液替代产品。这种血液替代物由纯化的牛血红蛋白组成，其分子内与o-腺苷-5'-三磷酸交联，分子间与o-腺苷交联，并与还原型谷胱甘肽（GSH）结合。在本研究中，我们以同种异体血浆为对照，比较了我们的新型血液替代物和未修饰的（U）Hb对人体血液成分的影响：红细胞（RBC）、血小板、单核细胞（Mo）和低密度脂蛋白（LDL）。通过测量渗透脆性和机械脆性、共轭二烯（CD）、脂质氢过氧化物（LOOH）、硫代巴比妥酸反应物（TBAR-S）、异前列腺素（8-异前列腺素F2α）和细胞内GSH，研究了两种Hb溶液对RBC的促氧化潜力。通过CD、LOOH和TBAR-S以及载脂蛋白（apo）B交联程度评估LDL的氧化修饰。通过监测血小板对聚集激动剂（胶原、ADP、肾上腺素和花生四烯酸）的反应，研究了Hb对血小板的影响。将单核细胞与Hb溶液或血浆一起培养，检测TNF-α和IL-1β的释放，然后通过电子显微镜检查。结果表明，天然的U Hb引发了许多血液成分的氧化应激，并加剧了Mo的炎症反应。它还导致RBC渗透脆性和机械脆性增加（p < 0.001）。虽然GSH水平略有变化，但RBC的脂质过氧化增加（p < 0.001）。发现U Hb是8-异前列腺素F2α合成的刺激剂、LDL的有效调节剂以及激动剂诱导的血小板聚集的有效增强剂。相反，我们的新型血液替代物似乎不会诱导氧化应激，也不会增加Mo的炎症反应。RBC的渗透脆性和机械脆性与对照组相似。这种修饰的Hb未能改变LDL，增加8-异前列腺素F2α的产生，但显著抑制血小板聚集。这种新型血液替代物的作用可能与腺苷的细胞保护和抗炎特性有关，腺苷用作交联剂和表面改性剂，以及一种降低血红蛋白促氧化潜力的改性方法。
