Hypoxia results in sulfatide exposure on the erythrocyte membrane and SMase incubation adds to the hypoxic adhesion of sickle red blood cells and anti-sulfatide antibody can inhibit SMase enhanced adhesion. (A) RBCs isolated from peripheral blood of 3 patients with SCD were treated with sodium metabisulfite for 1 hour. Legend denotes the HbS percentage of each sample and percentage of sulfatide-positive cells based on the gating strategy. Erythrocyte size and sulfatide availability increase when oxygen is scavenged by sodium metabisulfite, suggesting that sulfatide is exposed to the membranes of elongated sickle erythrocytes. (B) Sickle red blood cell adhesion to laminin following either 1 hour PBS or SMase (2.5 mU/mL) incubation at 37°C. Antibodies (1:200 v/v) or PBS were added for incubation after 30 minutes for the remaining 30 minutes. Hypoxia enhances the adhesion of sickle red blood cells, and SMase further increases this adhesion. Enzymatic activity of SMase could lead to the outer surface of the sickle red cell membrane with greater sulfatide availability, which is known to promote adhesion to activated endothelial cells. NBS, sodium metabisulfite, Na2S2O5.
