Objective To study inhibition of rosuvastatin (Rosu) on homocysteine (Hcy)-induced reactive oxygen species (ROS) and apoptosis in endothelial progenitor cells (EPCs). Methods EPCs were isolated from peripheral blood and then incubated with Hcy, or pre-incubated with Rosu or with different stress signaling pathway inhibitors including mevalonate (100 μmol/L), acetyl-cysteine (NAC, 10 μmol/L), NADPH oxidase inhibitor (DPI, 10 μmol/L), and endothelial nitric oxide synthase inhibitor (LNMA, 1 mmol/L) before adding Hcy. Apoptosis rate was evaluated by fluorescence activated cell sorting (FACS) analysis. ROS levels were detected by 2’,7’-dichlorodihydrofluorescein diacetate (H2DCFH-DA). NADPH oxidases were evaluated with lucigenin-enhanced chemiluminescence. Expression of Nox4 mRNA was measured by RT-PCR. Results Rosu remarkably inhibited Hcy-induced ROS accumulation and apoptosis of EPCs, and antagonized Hcy-induced activation of NADPH oxidase and Nox4 mRNA expression. Nox4 siRNA transfected EPCs with a similar effect. Conclusion The protective effect of Rosu on EPCs possibly involves inhibition of Hcy-induced activation of Nox, ROS accumulation, and apoptosis of EPCs through Nox4 dependent mechanisms.