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Prevention of erythrocyte dehydration by specific blockade of the transport pathways promoting loss of potassium (K) is a potential therapeutic strategy for sickle cell (SS) disease. Dietary magnesium (Mg) pidolate supplementation over a 4-week period has been shown to inhibit K-Cl co-transport and reduce dehydration. We report here the results in 17 of 20 patients with SS disease treated in an open-label unblinded study of the effects of long-term (6 months) oral Mg pidolate administration (540 mg Mg/d). A significant decrease (P < 0.0025) was observed with Mg therapy in the distribution widths for red cell mean cell haemoglobin concentration (MCHC) (haemoglobin distribution width; HDW), reticulocyte mean cell volume (red cell distribution width of reticulocytes; RDWr) and MCHC (reticulocyte HDW; HDWr), activity of red cell K-Cl co-transport, Na/Mg exchanger and Ca2+-activated (Gardos) K+ channel, whereas red cell K and Mg contents were significantly increased. Hb levels and absolute reticulocyte counts did not change with Mg therapy. Two patients did not complete the trial because of diarrhoea and one did not complete the trial for unrelated reasons. Although the median number of painful days in a 6-month period decreased from 15 (range 0-60) in the year before the trial to 1 (range 0-18; P < 0.0005) during the period of Mg therapy, no firm conclusion on therapeutic efficacy could be drawn from this unblinded open-label trial.

Nd-HPFH are haematological conditions which are natural models to aid understanding of the haemoglobin (Hb) switch. In this paper we describe a new non-deletional hereditary persistence of fetal haemoglobin (nd-HPFH) associated with the highest Hb F level observed to date (up to 49% without haemopoietic stress). Sequence of the G gamma promoter revealed a cytidine insertion within a stretch of four cytidines, located between -200 and -203 bp with respect to the cap site. This insertion is situated within a polypyrimidine-polypurine region which can adopt a triple helix structure, and is therefore of particular interest with respect to the Hb switch mechanism.

The present report provides evidence that hydroxyurea (HU) improves the beta thalassaemic phenotype in mice receiving 200 mg/kg/d for 30 d. The haematocrit rose from 29 +/- 3% at day 0 to 37 +/- 4% at day 30 (P < 0.05), despite myelosuppression and decreased reticulocyte counts. The beta minor/alpha ratio of globin chain synthesis increased from 0.78 at day 0 to 0.97 at day 30 (P < 0.001). Membrane defects improved: the proportion of bound alpha chains decreased, the proportion of spectrin and ankyrin increased and red cell deformability also increased.

The alpha- and beta-thalassaemias are characterized by decreased erythrocyte deformability. To determine what effects excess alpha- and beta-haemoglobin (globin) chains have on cellular and membrane deformability, purified haem-containing alpha- and beta-chains were entrapped within normal erythrocytes. Entrapment of purified alpha-chains in normal erythrocytes resulted in a significant decrease in cellular and membrane deformability similar to that observed in beta-thalassaemia intermedia. The decreased deformability was correlated with alpha-chain membrane deposition, an alteration in membrane proteins and a decrease in membrane reactive thiol groups. These changes in membrane and cellular deformability were time dependent and closely correlated with membrane alpha-chain deposition. The membrane changes and the loss of membrane deformability appeared to account for the loss of cellular deformability in the alpha-chain loaded cells. While both beta-chain loaded and Hb H erythrocytes demonstrated a significant loss of cellular deformability, this loss was less pronounced than in the alpha-chain loaded and beta-thalassaemic cells and may arise from either the increased intracellular viscosity of the beta-chain loaded cells or to the smaller amount of membrane bound globin. In summary, these studies demonstrate that alteration of cellular and membrane deformability occurs very rapidly and as a direct consequence of the autoxidation and membrane binding of the unpaired globin chains.

The ouabain bumetanide resistant (OBR) K+ efflux was investigated in deoxygenated sickle cells in comparison to oxygenated ones, by using a specific inhibitor of the [K+, Cl-] co-transport system, [(DihydroIndenyl)Oxy] Alkanoic acid (DIOA). A DIOA sensitive and a DIOA resistant K+ efflux were measured in deoxygenated sickle cells. The DIOA sensitive K+ efflux shared the properties of the [K+, Cl-] co-transport system, being stimulated by decreased pH and hypoosmolarity. This DIOA sensitive K+ efflux represented 70% of the total K+ efflux at pH 7.0 and at low pO2 (10-15 mmHg). Thus, a small reduction in Ph effectively stimulated the [K+, Cl-] co-transport system in deoxygenated condition, and this may contribute significantly to the sickle cell dehydration. We conclude that at pH lower than 7.4, the [K+, Cl-] co-transport system is permanently activated in sickle cells and leads to sickle cell dehydration in both oxygenated and deoxygenated conditions.

A familial erythrocytosis is described. The propositus is a 52-year-old man who was found, on routine testing, to have an erythrocytosis with a Hb level of 19.8 g/dl. Physical examination and laboratory findings revealed an associated hemolysis. The P50 of the stripped hemolysis was normal. The 2,3-DPG level was reduced to 15% of normal but that of ATP was increased more than twofold. Red cell diphosphoglycerate mutase activity was normal as were the levels of all red cell glycolytic and hexose monophosphate shunt enzymes with the exception of pyruvate kinase (PK). The level of the latter was elevated to 24.5 U/g (normal: 6.2 +/- 1.1). The electrophoretic pattern of hemolysate PK was abnormal, showing three additional bands, one of these migrated as the M2 isozyme. The findings were similar in the propositus's daughter and her daughter and in one of the two brothers of the propositus. The second brother and the son of the first brother was exhibited erythrocytosis but their red cell 2,3-DPG levels were normal. In addition, the level of their RBC PK was reduced to between 50% and 60% of normal and the abnormal electrophoretic bands were absent. The erythrocytosis appears to be inherited as an autosomal-dominant trait. The relationship between the PK abnormalities appears complex.

Erythroid burst colonies derived from the cord blood of nine neonates and from the blood and liver of three fetuses aborted after 20 weeks of gestation were grown in plasma clot culture. Their quantitative study revealed a higher proportion of burst-forming units (BFU-Es) in cord blood than in cord blood of normal adults. In addition, colony-forming units (CFU-Es) were present in cord blood but absent from adult blood. Study of haemoglobin synthesis in 14-day cultures of cord blood BFU-Es showed a significantly higher degree of Hb A synthesis than was found in reticulocytes from fresh cord blood; this proportion was, however, similar to that expected in vivo about three weeks after birth. These data suggest that the hemoglobin switch is already programmed in most of the early erythroid precursors present in cord blood or full-term neonates and indicate that the differentiation time is probably of the same order of magnitude in vivo and in vitro. The proportion of Hb A and F synthesis in erythroid bursts was not influenced by the concentration of erythropoietin in the range studied--ie, from 0.5 to 12 international units. Low but identical proportions of Hb A synthesis were found both in erythroid cells from liver after two hours of incubation with [3H]-leucine, and in 14-day liver bursts from fetuses aborted at 20 weeks of gestation.