Lyn protein depletion increases adhesion of CD34⁺ bone marrow cells and cell lines to stromal cells in the presence of SDF-1. (A) CD34⁺ cell attachment was measured at 48 hours after nucleoporation with Lyn siRNA or control siRNA. See Figure 1 for control Western blot. Values in adhesion assays are mean plus or minus SD (n = 3). (B) HL-60 and Nalm-6 cell attachment to stromal cells in the presence of SDF-1 was measured at 48 hours after nucleoporation with Lyn siRNA or control siRNA. Values are mean plus or minus SD (n = 3). (C) FACS analysis of integrin expression in control siRNA–(light lines) and Lyn siRNA–(dark lines) transfected CD34⁺ cells (both curves almost completely match). Cells were stained with or without (dotted lines) anti–human integrin monoclonal antibodies followed by secondary antibodies and FACS analysis. (D) Attachment of SDF-1–stimulated or unstimulated Mo7e cells to surfaces coated with ICAM-1, VCAM-1, or BSA at 48 hours after transfection with Lyn siRNA or control siRNA. Values in adhesion assays are mean plus or minus SD (n = 4). (E) Attachment of SDF-1–stimulated HL-60 and Nalm-6 cells to surfaces coated with ICAM-1 after transfection with Lyn siRNA or control siRNA. Values are mean plus or minus SD (n = 4). (F) Adhesion of SDF-1–stimulated Mo7e cells to stromal marrow cells at 48 hours after transfection with Lyn siRNA or control siRNA. Antihuman ICAM antibody was used for ICAM-1 blockade. Values in adhesion are mean plus or minus SD (n = 4). (G) FACS analysis of β₂ integrin-activation epitope expression in control siRNA–(dotted lines) and Lyn siRNA–(dark lines) transfected HL-60 and MO7e cells. Cells were stained with or without (light lines) anti–human integrin monoclonal antibody 24 (it binds to activated CD11/CD18) or anti–human Mac-1 antibodies (they bind only to activated CD11b), followed by secondary antibodies and FACS analysis.