Ivabradine inhibits SDF-1-induced activation of Rac1 and reduces phosphorylation of MLC. (a) Human CD4-positive cells were pretreated with ivabradine (0.01, 0.05, and 0.1 μmol/L) for 15 minutes before cells were stimulated with SDF-1 (100 ng/mL). After 3 minutes, GTPase activity assay was performed (affinity precipitation with GST-PAK). Densitometric analysis of 5 independent experiments. Bars represent mean ± SD. * compared with SDF-1-stimulated cells. (b) SDF-1 leads to phosphorylation of MLC. Isolated CD4-positive lymphocytes were pretreated with ivabradine in different concentrations indicated before stimulation with 100 ng/mL SDF-1 for 5 min. Total lysates were analyzed by immunoblotting employing antibodies against phospho-MLC. Equal loading of intact protein was confirmed by staining for GAPDH. Densitometric analyses were performed of 5 independent experiments. Data are expressed as p-MLC normalized to GAPDH. Bars represent mean ± SD. * compared with SDF-1-stimulated cells; .
