Hyperhomocysteinemia is a significant risk factor in atherosclerosis and thrombosis. However, its role in the development of intimal hyperplasia after arterial reconstructive procedures remains uncertain. We therefore studied the effect of homocysteine on intimal hyperplasia in a rat model of carotid artery balloon injury. Twenty-four Sprague-Dawley rats were divided into three groups: control (saline infusion), and low dose (0.14 mg/day) and high dose (0.71 mg/day) homocysteine delivered continuously via osmotic pumps implanted intraperitoneally. All animals underwent left common carotid artery balloon denudation with sacrifice after 14 days. Plasma homocysteine levels, intimal hyperplasia, and cell proliferation of rat carotid arteries were determined. In vitro rat smooth muscle cell (SMC) proliferation with homocysteine treatment was also performed. Plasma homocysteine levels at sacrifice were 1.80+/-0.35, 2.65+/-0.05 and 3.50+/-0.22 microM in three groups, respectively. Intimal hyperplasia developed in all balloon-injured arteries in both control and homocysteine-treated animals. The intimal area and intima/media area ratio were increased by 92% (P<0.05) and 105% (P<0.05), respectively, in the high dose-homocysteine-treated animals as compared to the control animals. Homocysteine (high dose) also significantly promoted the intimal cell proliferation (bromodeoxyuridine incorporation) by 2.2-fold as compared to controls. Furthermore, homocysteine treatment in the cell culture study showed a concentration-dependent increase of rat SMC proliferation. These data demonstrate that the continuous intraperitoneal administration of homocysteine significantly increases intimal hyperplasia and SMC proliferation after carotid artery balloon injury in the rat as well as in vitro SMC proliferation. This study suggests that, following arterial reconstructive procedures, elevated plasma homocysteine may increase the complications of clinical restenoses that are associated with intimal hyperplasia.