RESUMO
BACKGROUND AND PURPOSE: Endothelial cell proliferation, migration and adhesion are necessary for the formation of new blood vessels. We reported previously that baicalein strongly inhibited proliferation of rat heart endothelial cells and here we assess effects on migration and adhesion of these cells. EXPERIMENTAL APPROACH: Effects of baicalein on endothelial migration and adhesion were determined by in vitro wound assays and in modified Boyden chambers. Protein expression and subcellular distribution in rat heart endothelial cells were analysed by immunoblots and immunofluorescence staining. RESULTS: Pretreatment with baicalein for 48 h resulted in a concentration-dependent inhibition of endothelial migration, with an IC(50) of approximately 20 microM. Adhesion assays revealed that baicalein stimulated endothelial cell adhesion to fibronectin and vitronectin, effects blocked by the synthetic peptide Arg-Gly-Asp (RGD). Moreover, treatment with a blocking antibody against integrin alpha5beta1 drastically attenuated baicalein-mediated endothelial adhesion to fibronectin, but not to vitronectin. Furthermore, baicalein-mediated anti-migration effect and adhesion promotion could be partially reversed by the addition of 12(S)-hydroxyeicosatetraenoic acid (12(S)-HETE). Western blot analysis indicated that baicalein increased expression levels of integrin-alpha5beta1, -alphavbeta3 and vinculin proteins. Immunofluorescence staining showed that baicalein induced a marked reorganization of actin stress fibres and the recruitment of vinculin and integrins to focal adhesion plaques, with consequently increased formation of focal adhesion contacts. CONCLUSIONS AND IMPLICATIONS: Baicalein markedly inhibited the migration and enhanced the adhesion of rat heart endothelial cells, possibly by up-regulation of the integrins (alpha5beta1 and alphavbeta3) and vinculin and by promotion of actin reorganization and focal adhesion contact formation.
