The Oxidantscan: Evaluating Oxidant-induced Loss of Red Blood Cell Deformability in Sickle Cell Disease and Thalassemia
Sickle cell disease (SCD) and thalassemia are hereditary hemoglobinopathies characterized by chronic hemolytic anemia. Oxidative stress is a key feature in the pathophysiology of both conditions. To better understand the effect of oxidative stress on red blood cell (RBC) deformability in these complex diseases, we have optimized the oxidantscan, a laboratory assay that measures the susceptibility of RBC to oxidant-induced loss of deformability under shear stress.
For the oxidantscan RBCs are exposed to cumene hydroperoxide (CHP, 150 μM)[1]. Whole blood samples were collected from patients with SCD [HbSS (n=11), HbSC (n=11)], thalassemia [β-thalassemia major (transfusion-dependent (TDT) n=4, non-transfusion-dependent (NTDT) n=1), β-thalassemia minor with α-globin triplication (NTDT, n=1), HbH disease (NTDT, n=2)] and healthy controls (HCs; n=11). Three HbSS and four HbSC patients had concomitant α-thalassemia. RBC deformability was measured during 30 minutes at constant shear stress (30 Pa), using the Lorrca MaxSis (RR Mechatronics). Conventional and novel oxidantscan parameters, potentially reflecting distinct aspects of oxidative stress-induced deformability loss, were compared across subgroups.
[1] Larkin, S.K., Hernández, C., van Beers, E.J. et al. The RoxyScan is a novel measurement of red blood cell deformability under oxidative and shear stress. Sci Rep 14, 6344 (2024). https://doi.org/10.1038/s41598-024-56814-8
Exposure to CHP induced a time-dependent reduction of RBC deformability in all groups. Compared to HCs (median T-POD 934.0 s), RBCs from patients with HbSS exhibited significantly earlier onset of deformability loss (median T-POD 600.5 s), defined as T-POD [time (s) to 10% decrease from maximum elongation index (EImax)]. RBCs from HbSC patients showed a greater overall reduction in deformability (median ΔEI 0.62 vs. 0.48 in HCs) and a steeper decline over time (median slope −8.60E4) compared with both HCs (−5.70E4) and HbSS patients (−5.53E4). Among thalassemia subgroups, NTDT patients tended to have a lower T-POD and a higher ΔEI than TDT patients and HCs (significance not assessed due to low sample size).
Within HbSS, hydroxyurea (HU)-treated patients demonstrated a lower T-POD (median T-POD 500.8 s) than HbSS patients not treated with HU (median T-POD 760.7 s; p=0.045). In contrast, HU-treatment in HbSC patients was associated with a lower total loss of deformability (median ΔEI 0.63 vs. 0.56; p=0.038). However, all findings should be interpreted cautiously due to limited sample size and possible confounding by coinherited α-thalassemia.
The optimized oxidantscan is a robust laboratory assay, enabling quantitative assessment of the effect of oxidative stress on RBC deformability. The observed genotype-dependent differences in SCD and thalassemia subgroups suggest differences in sensitivity to oxidative challenge. The oxidantscan may serve as a valuable tool for stratifying disease severity and monitoring therapeutic responses in patients with SCD and thalassemia.
