Kuross S A, Hebbel R P
Department of Medicine, University of Minnesota Medical School, Minneapolis.
Blood. 1988 Oct;72(4):1278-85.
Previous studies documented the abnormal association of heme and heme proteins with the sickle RBC membrane. We have now examined RBC ghosts and inside-out membranes (IOM) for the presence of nonheme iron as detected by its formation of a colored complex with ferrozine. Sickle ghosts have 33.8 +/- 18.2 nmol nonheme iron/mg membrane protein, and sickle IOM have 4.3 +/- 3.0 nmol/mg. In contrast, normal RBC ghosts and IOM have no detectable nonheme iron. The combination of heme and nonheme iron in sickle IOM averages nine times the amount of membrane-associated iron in normal IOM. Kinetics of the ferrozine reaction show that some of this nonheme iron on IOM reacts slowly and is probably in the form of ferritin, but most (72% +/- 18%) reacts rapidly and is in the form of some other biologic chelate. The latter iron compartment is removed by deferoxamine and by treatment of IOM with phospholipase D, which suggests that it represents an abnormal association of iron with polar head groups of aminophospholipids. The biologic feasibility of such a chelate was demonstrated by using an admixture of iron with model liposomes. Even in the presence of tenfold excess adenosine diphosphate, iron partitions readily into phosphatidylserine liposomes; there is no detectable association with phosphatidylcholine liposomes. To examine the bioavailability of membrane iron, we admixed membranes and t-butylhydroperoxide and found that sickle membranes show a tenfold greater peroxidation response than do normal membranes. This is not due simply to a deficiency of vitamin E, and this is profoundly inhibited by deferoxamine. Thus, while thiol oxidation in sickle membranes previously was shown to correlate with heme iron, the present data suggest that lipid peroxidation is related to nonheme iron. In control studies, we did not find this pathologic association of nonferritin, nonheme iron with IOM prepared from sickle trait, high-reticulocyte, postsplenectomy, or iron-overloaded individuals. These data provide additional support for the concept that iron decompartmentalization is a characteristic of sickle RBCs.
以往的研究记录了血红素和血红素蛋白与镰状红细胞膜的异常关联。我们现在检测了红细胞血影和外翻膜（IOM）中是否存在非血红素铁，检测方法是利用其与亚铁嗪形成有色复合物。镰状血影含有33.8±18.2 nmol非血红素铁/毫克膜蛋白，镰状IOM含有4.3±3.0 nmol/毫克。相比之下，正常红细胞血影和IOM未检测到非血红素铁。镰状IOM中血红素和非血红素铁的总量平均是正常IOM中膜结合铁含量的九倍。亚铁嗪反应动力学表明，IOM上的一些非血红素铁反应缓慢，可能以铁蛋白的形式存在，但大多数（72%±18%）反应迅速，以其他某种生物螯合物的形式存在。后一种铁组分可被去铁胺以及用磷脂酶D处理IOM去除，这表明它代表铁与氨基磷脂极性头部基团的异常结合。通过将铁与模型脂质体混合，证明了这种螯合物在生物学上的可行性。即使存在十倍过量的二磷酸腺苷，铁也很容易分配到磷脂酰丝氨酸脂质体中；未检测到与磷脂酰胆碱脂质体的结合。为了检测膜铁的生物利用度，我们将膜与叔丁基过氧化氢混合，发现镰状膜的过氧化反应比正常膜大十倍。这不仅仅是由于维生素E缺乏所致，且被去铁胺显著抑制。因此，虽然之前已表明镰状膜中的硫醇氧化与血红素铁相关，但目前的数据表明脂质过氧化与非血红素铁有关。在对照研究中，我们未发现非铁蛋白、非血红素铁与镰状性状、高网织红细胞、脾切除术后或铁过载个体制备的IOM存在这种病理关联。这些数据为铁去分隔化是镰状红细胞的一个特征这一概念提供了额外支持。
