The Properties of Red Blood Cells from Patients Heterozygous for HbS and HbC (HbSC Genotype)

Abstract

Sickle cell disease (SCD) is one of the commonest severe inherited disorders, but specific treatments are lacking and the pathophysiology remains unclear. Affected individuals account for well over 250,000 births yearly, mostly in the Tropics, the USA, and the Caribbean, also in Northern Europe as well. Incidence in the UK amounts to around 12-15,000 individuals and is increasing, with approximately 300 SCD babies born each year as well as with arrival of new immigrants. About two thirds of SCD patients are homozygous HbSS individuals. Patients heterozygous for HbS and HbC (HbSC) constitute about a third of SCD cases, making this the second most common form of SCD, with approximately 80,000 births per year worldwide. Disease in these patients shows differences from that in homozygous HbSS individuals. Their red blood cells (RBCs), containing approximately equal amounts of HbS and HbC, are also likely to show differences in properties which may contribute to disease outcome. Nevertheless, little is known about the behaviour of RBCs from HbSC heterozygotes. This paper reviews what is known about SCD in HbSC individuals and will compare the properties of their RBCs with those from homozygous HbSS patients. Important areas of similarity and potential differences will be emphasised.

Figure 1

Schematic diagram of the main transport pathways activated in red blood cells (RBCs) from sickle cell patients. In RBCs from homozygous HbSS individuals, high cation permeability is accounted for by three main pathways [28, 37]. Under oxygenated conditions, the KCl cotransporter (KCC) is highly active. It is overexpressed in HbSS cells compared to HbAA ones and does not become quiescent as RBCs mature. It is stimulated further by low pH (reduction in extracelluar pH from 7.4 to 7). Under deoxygenated conditions, KCC remains active—again unlike the situation in HbAA RBCs [40]. In addition, two other pathways are observed. The deoxygenation-induced cation conductance (or P _sickle) is activated as HbS polymerises. It mediates entry of Ca²⁺. Elevation in intracellular Ca²⁺ then leads to activation of the third pathway, the Ca²⁺-activated K⁺ channel, or Gardos channel. These three pathways result in solute loss, cell shrinkage and dehydration, and consequent increase in [HbS]. They thereby contribute to pathogenesis of sickle cell disease. They are also likely to be involved in solute loss from RBCs of patients heterozygous for HbS and HbC (HbSC genotype), though details are lacking and differences in their behaviour compared to that in HbSS cells are expected.

Figure 2

Effect of oxygen tension on the activity of KCl cotransport (KCC) or P _sickle in red blood cells (RBCs) from normal individuals or patients with sickle cell disease. The activity of each transport pathway is normalised—to the value in oxygenated cells (150 mmHg O₂) for KCC activity and for that in deoxygenated RBCs (0 mmHg) in the case of P _sickle—and given as a percentage. Solid circles give KCC activity in RBCs from normal HbAA individuals; open symbols give KCC activity (open circles) or P _sickle activity (open triangles) in RBCs from sickle cell patients (HbSS homozygotes). In these experiments, total magnitude of KCC activity was about 10-fold greater in RBCs from HbSS individuals compared with HbAA ones. Note how the deoxygenation-induced KCC activity and activation of P _sickle follow a similar dependence on O₂ tension. Data taken from [67].

Figure 3

Components of K⁺ transport pathways and sickling in red blood cells (RBCs) from sickle cell disease patients heterozygous for HbS and HbC (HbSC genotype). K⁺ influxes are given as flux units [mmol·(l cells·h)⁻¹] measured at 5 mM [K⁺]_o and numbers of sickled cells as a percentage of total RBCs in fully oxygenated (150 mmHg O₂) or deoxygenated (0 mmHg) conditions. Although this technique measures a K⁺ influx, because of the high K⁺ content of RBCs, net solute movement through the transport systems will be outwards. KCl cotransport activity was calculated as the Cl⁻-dependent K⁺ influx, Gardos channel activity as the clotrimazole (5 μM)-sensitive K⁺ influx, and P _sickle as the Cl⁻-independent K⁺ influx (Cl⁻ substituted with NO₃ ⁻). Sickling, P _sickle, and Gardos channel activation occurs in deoxygenated conditions—as for HbSS RBCs—but KCC activity is low when O₂ is removed (as in RBCs from HbAA cells). Data taken from [64].

Figure 4

Conductance of red blood cells (RBCs) from sickle cell patients heterozygous for HbS and HbC (HbSC genotype). (a), (b) Representative whole-cell recordings from (a) oxygenated and (b) deoxygenated RBCs. (c) Mean whole-cell currents + S.E.M., n = 5. Test potentials from −80 to +80 mV were applied for 300 ms in 10 mV increments from a holding potential of −10 mV. Measurements were made using Na⁺-containing bath and pipette solutions. Data taken from [20]. See [66] for experimental details. The conductance of RBCs from HbSC patients is high and increases further on deoxygenation.