MSCs display P-selectin–dependent rolling and adhesion behavior on ECs. (A-B) P-selectin–dependent interaction of MSCs under flow. (A) MSCs (106) were preincubated or not with anti–P-selectin antibody and analyzed in the parallel plate flow chamber at a calculated shear stress of 0.1 dynes/cm2 (▪) or 2.0 dynes/cm2 (□) on precoated HUVECs stimulated with TNF-α (10 ng/mL for 4-6 hours). Numbers of adhered cells were determined in 3 representative fields. (B) MSCs (106) were flushed over glass slides precoated with P-selectin or control slides coated with BSA-containing buffer only in the parallel plate flow chamber at a calculated shear stress of 0.2 dynes/cm2. Numbers of rolling cells were determined in 3 representative microscopic fields. (C) MSC binding to HUVECs is not blocked by anti–PSGL-1. MSCs or CD34+ progenitors (106) were flushed over HUVEC layers pretreated with 10 ng/mL TNF-α for 4 to 6 hours. Numbers of cells were determined after application of a calculated shear stress of 0.1 dynes/cm2 (▪) or 2.0 dynes/cm2 (□). Pretreatment of MSCs or CD34+ cells with anti–PSGL-1 antibody was performed as indicated. (D) Influence of VCAM-1/VLA-4 on adhesion of MSCs to HUVECs. MSCs were pretreated or not with anti–VLA-4 antibody, HUVECs with anti–P-selectin or anti–VCAM-1 (or both) antibody for 30 minutes prior to analysis as indicated. Subsequently, 106 MSCs were flushed through a parallel plate flow chamber and numbers of adherent cells were recorded after application of a calculated shear stress of 0.1 dynes/cm2 (▪) or 2.0 dynes/cm2 (□). HUVEC monolayers were pretreated with 10 ng/mL TNF-α for 4 to 6 hours prior to the experiments. MSCs used for the experiments were derived from passages 6 to 9. Values are means ± SD; n = 3.
