Figure 4

MAPK pathways are required for SDF-1-induced ICAM-1 expression and cell adhesion. DLD-1 cells were kept as CL or stimulated with 10 ng/mL SDF-1 for 4 h. Before being kept as CL or stimulated with SDF-1, DLD-1 cells were pretreated with PD98059 (PD), SP600125 (SP) or SB203580 (SB) individually for 1 h, or transfected with control shRNA (sh-CL), or a specific shRNA of sh-ERK, sh-JNK or sh-p38. (A) The gene silencing efficiency of 48-h transfection of shRNA on ERK, JNK, and p38 protein levels of DLD-1 cells. After 48 h of transfection, protein was isolated and the ERK, JNK, and p38 expression was analyzed by Western blotting. (B) All bar graphs represent multiple increases of CL DLD-1 cells normalized to 18S rRNA. (C) DLD-1 cells were stimulated with or without SDF-1 for 4 h, and were added to HUVECs for 1 h. The results are shown as mean ± SEM. *P < 0.05 versus CL. ^#P < 0.05 versus vehicle control (DMSO) or control shRNA (sh-CL) with SDF-1 stimulation. (D) DLD-1 cells were kept as CL, pretreated with DMSO, PD98059 (PD), SP600125 (SP), or SB203580 (SB), and stimulated with SDF-1 for 4 h. The total ICAM-1 protein expressions were determined by Western blot analysis. (E) DLD-1 cells were kept as CL, pretreated with AMD3100, and stimulated with SDF-1; the phosphorylation of ERK, JNK and p38 was determined by Western blotting.