Reentry into the cell cycle of contact-inhibited vascular endothelial cells by a phosphatase inhibitor - Possible involvement of extracellular signal-regulated kinase and phosphatidylinositol 3-kinase

Vascular endothelial cells are unique in that they exit from the cell cycle when they come into contact with each other. Although the phenomenon is ca lled "contact inhibition," little is known about the cellular mechanisms in volved. Here we show that the phosphatase inhibitor sodium orthovanadate (S OV) induced the reentry of contact-inhibited human umbilical vascular endot helial cells (HUVECs) into the cell cycle and that reentry was associated w ith activation of the extracellular signal-regulated kinase (ERK) and phosp hatidylinositol 3-kinase (PI 3-K)/Akt pathways. SOV stimulated [H-3]thymidi ne uptake of contact-inhibited HUVECs in a time- and dose-dependent manner. SOV-induced increase in [SH]thymidine uptake was significantly inhibited b y the mitogen-activated protein kinase kinase inhibitor PD98059 and by the PI 3-K inhibitor LY294002, SOV also stimulated the expression of cyclin D1, cyclin E, and cyclin A, and the activity of CDK2 kinase, whereas it decrea sed the expression of p27(kip1). In marked contrast, growth media alone did not induce these changes. Furthermore, these SOV-induced changes were abol ished by pretreatment with PD98059 and LY294002, SOV stimulated phosphoryla tion of ERK and Akt in contact-inhibited HUVECs, while growth media alone d id not. This phosphorylation was associated with inhibition of phosphatase activity in the cells. Finally, overexpression of high cell density-enhance d protein tyrosine phosphatase 1 inhibited c-fos and cyclin A promoter acti vity. Taken together, our results suggest that in contact-inhibited HUVECs, increased phosphatase activity suppressed the ERK and PI 3-K/Akt pathways, resulting in exit from the cell cycle by down-regulation of cyclin D1, cyc lin E, and cyclin A and by upregulation of p27(kip1).