Granulocyte-colony stimulating factor (G-CSF) has been reported to improve the function of infarcted heart, but its effects on atherosclerosis are unclear. Here we examined the effects and the potential mechanisms in the high-fat diet rabbit model. Six-month-old male New Zealand white rabbits, fed a high-cholesterol diet or a normal diet for 10 weeks, were treated with vehicle or G-CSF. G-CSF increased lesion area in the thoracic aorta and the plasma levels of total cholesterol (TC) and low-density lipoprotein-cholesterol (LDL-C) at the early phase in the high-fat diet group. High-fat diet-induced arterial endothelium damage and apoptosis were greatly aggravated by G-CSF treatment. In vivo, G-CSF impaired apoptosis induced by oxidized low density lipoprotein (OX-LDL) but it had little effect on cultured endothelial cells (ECs) with vehicle treatment. Further research revealed that G-CSF promoted the upregulation of endothelin-1 (ET-1) and the downregulation of endothelial nitric oxide synthase (eNOS) of thoracic aortae induced by a high-fat diet. In vitro, the effects of G-CSF on expression of ET-1 and eNOS in cultured ECs were consistent with those in vivo. Our results suggested that G-CSF exacerbates lipid abnormity and endothelium damage in hyperlipidemia rabbits, thereby resulting in the deterioration of atherosclerosis and that the ET-1/eNOS system may regulate the progression.