Bertles J F, Milner P F
J Clin Invest. 1968 Aug;47(8):1731-41. doi: 10.1172/JCI105863.
THE AMOUNT OF FETAL HEMOGLOBIN (HB F) IN ERYTHROCYTES OF PATIENTS WITH SICKLE CELL ANEMIA (HB SS DISEASE) WAS MEASURED BY TWO METHODS: (a) photometry of individual cells stained for Hb F by the Kleihauer-Betke technique; and (b) chemical assay of alkali-resistant hemoglobin in cells distributed according to specific gravity by ultracentrifugation. Irreversibly sickled cells (ISC), which could be identified directly during photometry and which were found to gather in high concentration at the bottom of ultracentrifuged cell columns, contained significantly less Hb F than non-ISC. Cell content of total Hb was constant regardless of cell size, shape, or ultracentrifugal behavior: thus absolute amounts of Hb F and S varied reciprocally from cell to cell. In experiments designed to estimate age, at formation, and rate of destruction of ISC, Hb SS blood was incubated with selenomethionine-(75)Se (which labels reticulocytes) or (51)Cr (which labels erythrocytes at random) and reinfused. Sequential blood samples were separated by ultracentrifugation into fractions rich in reticulocytes, non-ISC, and ISC; and chronological changes in the specific activity of each fraction were determined. Analogous information was obtained from radioautography of sequential blood samples after reinfusion of whole blood labeled with amino acids-(3)H: this technique permitted direct visual characterization of labeled erythrocytes as ISC or non-ISC, all of which had been reticulocytes at the time of reinfusion. The transformation of non-ISC into ISC, presumably a manifestation of membrane damage, proved to begin soon after cell release from the marrow; and ISC subsequently underwent rapid removal from the circulating blood. IT IS THEREFORE APPARENT FROM THESE STUDIES THAT, IN HB SS DISEASE, RELATIVELY SMALL RECIPROCAL CHANGES IN THE AMOUNTS OF THE TWO MAJOR HEMOGLOBINS CARRY PREDICTIVE IMPORTANCE: (a) net synthesis of Hb F is least in erythroid cells destined to become ISC; and (b) these irreversibly deformed erythrocytes suffer preferential destruction.
采用两种方法测定镰状细胞贫血（血红蛋白SS病）患者红细胞中的胎儿血红蛋白（Hb F）含量：（a）用Kleihauer - Betke技术对单个细胞进行Hb F染色后的光度测定法；（b）通过超速离心按比重分布的细胞中耐碱血红蛋白的化学测定法。不可逆镰状细胞（ISC）在光度测定过程中可直接识别，且发现在超速离心后的细胞柱底部高度聚集，其所含Hb F明显少于非ISC。无论细胞大小、形状或超速离心行为如何，细胞中总血红蛋白的含量是恒定的：因此，每个细胞中Hb F和S的绝对量呈反比变化。在旨在估计ISC形成时的年龄及其破坏速率的实验中，将血红蛋白SS血液与硒代蛋氨酸 -（75）Se（标记网织红细胞）或（51）Cr（随机标记红细胞）一起孵育，然后再输注。通过超速离心将连续的血样分离成富含网织红细胞、非ISC和ISC的部分；并测定每个部分的比活性随时间的变化。在用氨基酸 -（3）H标记的全血再输注后，通过对连续血样进行放射自显影获得了类似信息：该技术允许将标记的红细胞直接直观地鉴定为ISC或非ISC，所有这些红细胞在再输注时均为网织红细胞。非ISC向ISC的转变，大概是膜损伤的一种表现，事实证明在细胞从骨髓释放后不久就开始了；随后ISC迅速从循环血液中清除。因此，从这些研究中可以明显看出，在血红蛋白SS病中，两种主要血红蛋白含量的相对小的反比变化具有预测意义：（a）在注定要成为ISC的红细胞中，Hb F的净合成最少；（b）这些不可逆变形的红细胞遭受优先破坏。
