Figure 3.

Leptin-induced migration can be abolished by CYT-354 treatment both in vitro and in vivo. A and C, B and D, Chemotaxis of Sca-1⁺ progenitor cells in response to 100 ng/mL of leptin with CYT-354 (A; n=5) or WP1066 (B; n=5) in an 8.0 μm transwell system was identified using 1% crystal violet staining after 16-h incubation (scale bars, 20 μm). Serum-free medium and dimethyl sulfoxide (DMSO) were used as controls. Migration index of transwell assays was defined as the mean ratio of treatment to control of cell numbers counted per 5 random fields at ×20 magnification. E, Sca-1⁺ progenitor cells were successfully transfected by RFP (red fluorescent protein) lentivirus (scale bars, 40 μm). F, 1×10⁶ RFP Sca-1⁺ with (c and d) or without CTY-354 (a and b) were seeded in the adventitial side of injured femoral arteries (n=6). En face staining showed that the RFP cells migrated from the adventitia to the intima on 1 or 3 d post-surgery (scale bar, 100 μm). G, Quantification of migratory cells from the adventitia to intima with or without CYT-354 inhibitor on 1 d post-surgery. H, Quantification of migratory cells from the adventitia to intima with or without CYT-354 inhibitor on 3 d post-surgery. I, Quantification of migratory cells from the adventitia to intima without CYT-354 inhibitor on 1 or 3 d post-surgery. J, Quantification of migratory cells from the adventitia to intima with CYT-354 inhibitor on 1 or 3 d post-surgery. Cell migration assay in vivo was performed in wild-type mice. All graphs are shown as mean±SEM. *P<0.05, **P<0.01, ***P<0.001.