HCELL on KG1a cells is the primary mediator of shear-resistant E-selectin binding. (A) KG1a cells were perfused over CHO-E cell monolayers for 2 minutes at wall shear stress = 1 dyn/cm²; then detachment assays were performed by increasing shear stress stepwise every 30 seconds. The number of rolling cells at the end of each interval was counted and calculated as the percentage relative to the number of cells rolling at the start of the detachment assay (ie, end of the attachment period at 1 dyn/cm²). All binding was E-selectin–dependent, confirmed by inhibition of attachment in the presence of function-blocking anti–E-selectin mAb 68-5H11 and by treatment with EDTA (data not shown), and lack of binding of KG1a cells to CHO-mock cells. HCELL-mediated E-selectin binding was most evident at shear stress levels > 4 dyn/cm². Data are mean ± SEM; n = 5. *P < .05, relative to untreated and vector controls. Importantly, on CD44 silencing no differences were observed on HECA-452 reactivity of PSGL-1/CLA (Figure 5C), which typically correlates with selectin binding capacity. (B) Velocity of KG1a cell rolling on CHO-E cells at each shear stress was calculated as the distance traveled by the centroid of the cell divided by the time period of observation (5 seconds). Cells lacking HCELL (CD44 siRNA-transduced) rolled significantly faster than those expressing HCELL (untreated, vector-transduced). Data are mean ± SEM; n > 20 cells. *P < .05, relative to untreated and vector control. **P < .01 relative to untreated and vector control.