Ionophore-mediated swelling of erythrocytes as a therapeutic mechanism in sickle cell disease

Abstract

Sickle cell disease (SCD) is characterized by sickle hemoglobin (HbS) which polymerizes under deoxygenated conditions to form a stiff, sickled erythrocyte. The dehydration of sickle erythrocytes increases intracellular HbS concentration and the propensity of erythrocyte sickling. Prevention of this mechanism may provide a target for potential SCD therapy investigation. Ionophores such as monensin can increase erythrocyte sodium permeability by facilitating its transmembrane transport, leading to osmotic swelling of the erythrocyte and decreased hemoglobin concentration. In this study, we treated thirteen blood samples from patients with SCD with 10 nM of monensin ex vivo. We measured changes in cell volume and hemoglobin concentration in response to monensin treatment, and we perfused treated blood samples through a microfluidic device that permits quantification of blood flow under controlled hypoxia. Monensin treatment led to increases in cell volume and reductions in hemoglobin concentration in most blood samples, though the degree of response varied across samples. Monensin treated samples also demonstrated reduced blood flow impairment under hypoxic conditions relative to untreated controls. Moreover, here was a significant correlation between the improvement in blood flow and the decrease in hemoglobin concentration. Thus, our results demonstrate that a reduction in intracellular HbS concentration by osmotic swelling improves blood flow under hypoxic conditions. Although the toxicity of monensin prevents it from being a viable clinical treatment, these results suggest that osmotic swelling should be investigated further as a potential mechanism for SCD therapy.

Magnesium homeostasis in deoxygenated sickle erythrocytes is modulated by endothelin‐1 via Na + /Mg 2+ exchange

“…Erythrocytes dehydrate during periods of deoxygenation through activation of K + efflux through K + transporters such as KCC K‐Cl cotransporters or the KCNN4/KCa3.1 Gardos channel 51–53 . We tested the effect of ET‐1‐induced dehydration in the absence of the Gardos channel activity.…”

“…Erythrocytes dehydrate during periods of deoxygenation through activation of K + efflux through K + transporters such as KCC K‐Cl cotransporters or the KCNN4/KCa3.1 Gardos channel. 51 , 52 , 53 We tested the effect of ET‐1‐induced dehydration in the absence of the Gardos channel activity. Cycling deoxygenation for 3 h produced a significantly greater rightward shift in the density profile of SCD (SS) erythrocytes (Figure 4B ) than in normal (AA) red cells (Figure 4A ).…”

Phenotypic screening of the ReFrame Drug Repurposing Library to discover new drugs for treating sickle cell disease

“…The concept of decreasing the intracellular HbS concentration followed immediately on the decades-old observation that the delay time prior to detectable fiber formation depends on a very high power of the HbS concentration – the 30 th power in the initial experiments, which remains the highest concentration dependence ever observed for any molecular process. It also motivated approaches to treat the disease by decreasing the intracellular HbS concentration by means other than by increasing red cell volume(14, 48), such as introducing an iron deficiency anemia. There are multiple way of making cells iron deficient, including iron restriction, the use of chelating agents, inhibiting the enzymes that catalyze heme synthesis, and even what might be regarded as the heretical method of phlebotomy(7, 27, 49) - the method employed by physicians to treat illnesses 2000 years ago, most notably the Greek Hippocrates and Roman Galen, who subscribed to the theory that disease was caused by too much blood, one of the four humors.…”

“…The concept of decreasing the intracellular HbS concentration followed immediately on the decades-old observation that the delay time prior to detectable fiber formation depends on a very high power of the HbS concentration -the 30 th power in the initial experiments, which remains the highest concentration dependence ever observed for any molecular process. It also motivated approaches to treat the disease by decreasing the intracellular HbS concentration by means other than by increasing red cell volume (14,48), such as introducing an iron deficiency anemia.…”

Fluorescence Lifetime Measurement of Prefibrillar Sickle Hemoglobin Oligomers as a Platform for Drug Discovery in Sickle Cell Disease

“…A previously described − polydimethylsiloxane (PDMS) microfluidic device replicating the geometry of the postcapillary venules was used to study blood rheological behavior. Figure S5 shows the device layout with the inputs and outputs for each of the three layers.…”