Cardiovascular disease and its thrombotic complications, particularly myocardial infarction and stroke, are a main cause of death in industrialized countries. Currently, little is known about the underlying pathomechanisms of cigarette smoke on thrombosis and subsequent vessel wall remodeling, partly due to a lack of adequate in vivo models. We investigated the biological effects of sub-chronic inhalation exposure to cigarette mainstream smoke (MS) from the University of Kentucky reference cigarette 2R4F at doses of 600, 1200, 1800, and 2400 µg total particulate matter per liter per day on arterial thrombosis and intimal hyperplasia using the ferric chloride model of vascular injury in male apolipoprotein e-deficient (ApoE-/-) mice. Arterial thrombosis was analyzed in the left carotid artery by determining the time to thrombotic occlusion and patency rate after vascular injury. A dose-dependent decrease was seen in median time to occlusion (14.9 min for control, 9.7 min for MS-1800, and 8.0 min for MS-2400; p < 0.001 compared to control) and lower vascular patency rates after 25 minutes (40% for control, 10% for MS-1800, and 0% for MS-2400; p < 0.001 compared to control). Three weeks later, intimal hyperplasia was analyzed using immunohistochemistry and morphometry. Immunohistochemistry of neointimal lesions revealed an MS exposure-associated increase in the amount of alpha-actin-positive smooth muscle cells (range: 16% (control) to 25% (MS-2400); p < 0.05 for MS-2400 compared to control). No differences were seen in the number of apoptotic cells. Morphometry revealed an MS exposure-associated increase intima-media ratio (p < 0.05 for MS-1800 compared to control and p < 0.01 for MS-2400 compared to control). These data indicate that subchronic exposure of ApoE-/- mice to MS promotes arterial thrombosis and modulates the size and composition of vascular lesions.