AbstractGiven its broad effects in endothelium, vascular endothelial growth factor (VEGF) represents the primary rate‐limiting step of angiogenesis. Therefore, VEGF targeting therapies were soon developed. Bevacizumab and ranibizumab are two of these therapeutic agents already in clinical use. Bevacizumab was first used for cancer treatment, whereas ranibizumab was designed to target choroidal neovascularization, the main cause of blindness in age‐related macular degeneration. The present study aims to compare the multiple effects of bevacizumab and ranibizumab in human microvascular endothelial cells (HMECs). HMEC cultures were established and treated during 24 h with the anti‐VEGF agents within the intravitreal‐established concentration range or excipients. Analyses of VEGF content in cell media and VEGF receptor‐2 (VEGFR‐2) expression in cell lysates were performed. No cell cytotoxicity (MTS assay) was found in anti‐VEGF‐treated cultures at any concentration. Apoptosis (TUNEL assay) was significantly increased and cell proliferation (BrdU assay), migration (transwell assay) and assembly into vascular structures were significantly reduced by incubation with both agents at the two doses used. These findings were accompanied by a strong decrease in VEGF release, and in phosphorylated VEGFR‐2 and Akt expression for both agents at the clinical concentration. Interestingly, phosphorylated Erk was only significantly reduced upon bevacizumab treatment. In addition, proliferation was more affected by ranibizumab, whereas migration, capillary formation, and phosphorylated VEGFR2 expression were significantly reduced by bevacizumab as compared to ranibizumab. Therefore, although both agents presented anti‐angiogenic actions, distinct effects were exerted by the two molecules in HMEC. These findings suggest that a careful confirmation of these effects in clinical settings is mandatory. J. Cell. Biochem. 108: 1410–1417, 2009. © 2009 Wiley‐Liss, Inc.