Inhibitory Effects of Rap1GAP Overexpression on Proliferation and Migration of Endothelial Cells via ERK and Akt Pathways / 华中科技大学学报（医学）（英德文版）

ABSTRACT

Rap 1 is expressed in human umbilical vein endothelial cells (HUVECs).Rap1-GTPase activating protein (Rap 1 GAP),with its specific target,Rap 1,has been shown to be important in the regulation of many physiological and certain pathological processes.In this study,we investigated the effect of RaplGAP expression on endothelial cell function,or,more specifically,proliferation and migration of endothelial cells.HUVECs were transfected with pcDNA3.1 (empty vector),pcDNA3.1 containing Flag-tagged-Rap1GAP or Myc-tagged-Rap1N17.The proliferation,migration and tube formation were examined and compared among the 3 groups.Expression of Rap1,Rap1GAP,extracellular signal-regulated kinase (ERK),phospho-ERK,Akt,phosphor-Akt was detected by Western blotting.The results showed that the proliferation,migration and tube formation were significantly reduced in Rap1GAP- and Rap1N17-transfected HUVECs as compared with empty vector-transfected control.These changes were coincident with increased expression of Rap 1 GAP and decreased expression of activated Rap 1,phospho-ERK and -Akt.After treatment of Rap 1 GAP-transfected HUVECs with a stimulator of Rapl guanine-nucleotide-exchange factor (Rap1GEF) 8CPT-2'OMe-cAMP,it was found that Rap1 activity was decreased as compared with empty vector-transfected control.Pretreatment of HUVECs with an ERK inhibitor PD98059 or a PI3K inhibitor LY294002 prior to stimulation not only blocked 8CPT-2'OMe-cAMP-induced phosphorylation of ERK and Akt,but also significantly reduced cell proliferation and migration.Finally,we examined the effect of vascular endothelial growth factor (VEGF) on HUVECs overexpressing Rap1GAP.VEGF-stimulated Rap1 activity,phosphorylation of ERK and Akt,cyclin D1 expression and cell proliferation were repressed in HUVECs overexpressing Rap 1 GAP as compared to empty vector-transfected control.Taken together,our findings demonstrate that Rap1GAP/Rap1 and their downstream effectors regulate proliferation and migration of HUVECs via ERK and Akt pathways.