Heparanase (Hpse) is an endo–β-D-glucuronidase, the sole mammalian enzyme responsible for degradation of heparan sulphate (HS) which is a primary structural component of the extracellular matrix (ECM) and the vascular basement membrane. Upregulation of Hpse has been observed under different pathological conditions such as cancer and inflammation. Increased expression of Hpse has been also found in the serum and plaques of patients with atherosclerosis, a chronic inflammatory disease, but the precise roles of this enzyme in atherosclerosis are largely unknown. We investigated the role of Hpse in the advanced stage of atherosclerosis by using ApoE-/-/Hpse-/- or AExH-/- mice and compared atherosclerosis with ApoE-/- mice fed with a high-fat diet for 14 weeks. Compared to ApoE-/- mice, high-fat fed AExH-/- mice showed a significant reduction in atherosclerotic plaque burden at the aorta and aortic sinus confirmed by en face analysis of oil-red O lipid staining and morphometric analysis, respectively. Hpse also modified the immune response by modulating the recruitment of monocytes and dendritic cells to the lesion sites. These data indicated Hpse as a novel inflammatory mediator of atherosclerosis in the advanced stage of this disease. This finding provides a foundation for further investigation into the role of hpse in atherosclerosis and for the development of Hpse inhibitory drugs as novel therapeutics for the treatment of atherosclerotic disease.