L-selectin-dependent leukocyte rolling on endothelial monolayers under flow

Lcukocyte-endothelial (L/E) interactions during inflammation are regulated by the synchronized binding of endothelial adhesion molecules with corresponding leukocyte counter-receptors, including L-selectin. The dynamic interactions of rat basophilic leukemic cells (RBL), RBL cells expressing human L-selectin (RBL-LS), or human neutrophils (PMN) with human umbilical vein endothelial cells (HUVEC) were analyzed under physiologic shearing flow. HUVEC were cultured on glass slides, activated with tumor necrosis factor α (TNFα), and incorporated into a parallel-plate flow chamber. Leukocytes (105 cells/ml) were infused across the HUVEC monolayer (shear stress=0.8 dyn/cm2), and L/E interactions were monitored within a 0.64 mm2 area via videomicroscopy. Adherent cells were partitioned on the basis of the nature of adhesion, i.e., tethering (initial capture), transient (rolling), or firm (>30 sec). RBL, RBL-LS, and PMN did not interact with unstimulated HUVEC. The numbers of RBL, RBL-LS, and PMN firmly adherent to TNFα-activated HUVEC were similar. The flux of rolling RBL and PMN on activated HUVEC did not differ. RBL-LS demonstrated a greater number of initial captures (4.00±0.82 vs. 0.75±0.48/30 sec) and a greater rolling flux (2.00±0.44 vs. 0.57±0.30 cells/30 sec) than RBL. These data are consistent with a role for L-selectin in the tethering and rolling of leukocytes to activated endothelium, and suggest that this experimental system may be useful for studying the molecular mechanisms regulating L/E interactions.

Duke Scholars

Author Ralph Snyderman Medicine

Author Mark Wesley Dewhirst Radiation Oncology