Adherent neutrophils on vascular endothelium positively contribute to cell-cell aggregation and vaso-occlusion in sickle cell disease (SCD). In the present study, we demonstrated that pyridoxamine, a derivative of vitamin B6, might be a therapeutic agent to alleviate intravascular cell-cell aggregation in SCD. Using real-time intravital microscopy, we found that one oral administration of pyridoxamine, in a dose-dependent manner, increased the rolling influx of neutrophils and reduced neutrophil adhesion to endothelial cells in cremaster microvessels of SCD mice challenged with hypoxia-reoxygenation. Short-term treatment also mitigated neutrophil-endothelial cell and neutrophil-platelet interactions in the microvessels and improved the survival of SCD mice challenged with tumor necrosis factor-α. The inhibitory effects of pyridoxamine on intravascular cell-cell interactions were enhanced by co-treatment with hydroxyurea. We observed that long-term (5.5 months) oral treatment with pyridoxamine significantly reduced the adhesive function of neutrophils and platelets, and down-regulated the expression of E-selectin and intercellular adhesion molecule-1 on the vascular endothelium in tumor necrosis factor-α-challenged SCD mice. Ex vivo studies revealed that the surface amount of αMβ2 integrin was significantly decreased in stimulated neutrophils isolated from SCD mice treated with pyridoxamine-containing water. Studies using platelets and neutrophils from SCD mice and patients suggested that treatment with pyridoxamine reduced the activation state of platelets and neutrophils. These results suggest that pyridoxamine may be a novel therapeutic and a supplement to hydroxyurea to prevent and treat vaco-occlusion events in SCD.