Extracellular adenosine production by the GPI-anchored Ecto-5′-Nucleotidase (Ecto-5′-Nu) plays an important role in the cardiovascular system, notably in defense against hypoxia. It has been previously suggested that HMG-CoA reductase inhibitors (HRIs) could potentiate the hypoxic stimulation of Ecto-5′Nu in myocardial ischemia. In order to elucidate the mechanism of Ecto-5′-Nu stimulation by HRIs, Ecto-5′-Nu activity and expression were determined in an aortic endothelial cell line (SVAREC) incubated with lovastatin. Lovastatin enhanced Ecto-5′-Nu activity in a dose-dependent manner. This increase was not supported by de novo synthesis of the enzyme because neither the mRNA content nor the total amount of the protein were modified by lovastatin. By contrast, lovastatin enhanced cell surface expression of Ecto-5′-Nu and decreased endocytosis of Ecto-5′-Nu, as evidenced by immunostaining. This effect appeared unrelated to modifications of cholesterol content or Ecto-5′-Nu association with detergent-resistant membranes. The effect of lovastatin was reversed by mevalonate, the substrate of HMG-CoA reductase, by its isoprenoid derivative, geranyl-geranyl pyrophosphate, and by cytotoxic necrotizing factor, an activator of Rho-GTPases. Stimulation of Ecto-5′-Nu by lovastatin enhanced the inhibition of platelet aggregation induced by endothelial cells. In conclusion, lovastatin enhances Ecto-5′-Nu activity and membrane expression in endothelial cells. This effect seems independent of lowering cholesterol content but could be supported by an inhibition of Ecto-5′-Nu endocytosis through a decrease of Rho-GTPases isoprenylation.