Papassotiriou I, Kister J, Griffon N, Abraham D J, Kanavakis E, Traeger-Synodinos J, Stamoulakatou A, Marden M C, Poyart C
Hematology Laboratory, Aghia Sophia Children's Hospital, Athens, Greece.
Exp Hematol. 1998 Sep;26(10):922-6.
Patients with the nondeletion genotype of hemoglobinopathy H (HbH or beta4) disease have higher proportions of HbH and more severe tissue hypoxia than patients with the deletion genotype. Because these patients' red blood cells (RBCs) contain mainly two Hb species, HbH and HbA, the high proportion of HbA can be exploited by lowering its oxygen affinity; this would probably increase oxygen delivery to the RBCs and improve the patients' clinical phenotype. Allosteric effectors that induce a low-affinity Hb may be useful in this regard. We investigated the effect of a bezafibrate derivative, RSR-4, on the oxygen affinity of RBCs and purified hemolysates containing HbA and HbH. This allosteric effector crosses RBC membranes and binds reversibly to the alpha-chains of deoxy-Hb, decreasing hemoglobin oxygen affinity. The blood used was obtained from a patient with HbH disease (alphaTSaudi homozygote) whose HbH level was 33.5% as measured by high-performance liquid chromatography. Oxygen binding studies were performed in RBCs and purified hemolysates. RBCs incubated in the presence of 500 microM RSR-4 (2-[[[(3,5-dichloroanilino)-carbonyl]methyl]phenoxy]-2-methylpropi onic acid) in standard conditions (pH 7.4, 0.14 M NaCl, 37 degrees C) displayed an increase in their P50 value from 14.5 to 35.2 mm Hg. Oxygen binding studies in purified stripped hemolysates (pH 7.2, 0.1 M NaCl, 25 degrees C) showed that addition of both 500 microM RSR-4 and 1 mM of 2,3 diphosphoglycerate (DPG) led to an 11-fold decrease in oxygen affinity, whereas the addition of the natural effector DPG or RSR-4 alone produced a 2.7- and 5.7-fold decrease, respectively. In both cases, the oxygen equilibrium curves (OECs) were biphasic due to the presence of the noncooperative, high-oxygen-affinity HbH (beta4) component. After addition of RSR-4, the lower part of the OEC (corresponding to HbH) was not shifted compared with the upper part (corresponding to HbA). These results were confirmed by kinetic studies of CO recombination. Both experiments demonstrated that RSR-4 does not affect beta4 Hb. Our findings provide an experimental model for lowering the oxygen affinity of HbA in HbH-containing cells and suggest that the oxygen delivery capability of the latter would be thereby improved.
血红蛋白病H（HbH或β4）疾病非缺失基因型患者比缺失基因型患者有更高比例的HbH和更严重的组织缺氧。因为这些患者的红细胞（RBC）主要含有两种血红蛋白，即HbH和HbA，所以可以通过降低HbA的氧亲和力来利用其高比例；这可能会增加向RBC的氧输送并改善患者的临床表型。诱导低亲和力Hb的变构效应剂在这方面可能有用。我们研究了苯扎贝特衍生物RSR-4对RBC以及含有HbA和HbH的纯化溶血产物氧亲和力的影响。这种变构效应剂可穿过RBC膜并与脱氧Hb的α链可逆结合，降低血红蛋白的氧亲和力。所用血液取自一名HbH病患者（α沙特纯合子），通过高效液相色谱法测得其HbH水平为33.5%。在RBC和纯化溶血产物中进行了氧结合研究。在标准条件（pH 7.4、0.14 M NaCl、37℃）下，在500μM RSR-4（2-[[[(3,5-二氯苯胺基)-羰基]甲基]苯氧基]-2-甲基丙酸）存在下孵育的RBC，其P50值从14.5 mmHg增加到35.2 mmHg。在纯化的脱铁溶血产物（pH 7.2、0.1 M NaCl、25℃）中进行的氧结合研究表明，添加500μM RSR-4和1 mM 2,3-二磷酸甘油酸（DPG）会导致氧亲和力降低11倍，而单独添加天然效应剂DPG或RSR-4分别导致氧亲和力降低2.7倍和5.7倍。在这两种情况下，由于存在非协同、高氧亲和力的HbH（β4）成分，氧平衡曲线（OEC）呈双相。添加RSR-4后，OEC的下部（对应于HbH）与上部（对应于HbA）相比没有移动。这些结果通过CO重组的动力学研究得到证实。两个实验均表明RSR-4不影响β4 Hb。我们的研究结果为降低含HbH细胞中HbA的氧亲和力提供了一个实验模型，并表明由此可改善后者的氧输送能力。
