Pioglitazone inhibits homocysteine-induced migration of vascular smooth muscle cells through a peroxisome proliferator-activated receptor gamma-independent mechanism.

1. Peroxisome proliferator-activated receptor (PPAR)-gamma agonists have been demonstrated to exert protective effects against homocysteine (Hcy)-induced pathogenesis. However, the effects of PPAR-gamma agonists on Hcy-induced migration are unknown. In the present study, we examined the effect of pioglitazone on the migration of vascular smooth muscle cells (VSMC) induced by Hcy and the possible mechanism involved. 2. Vascular smooth muscle cells were isolated from the thoracic aortas of male Sprague-Dawley rats. The migration of VSMC was examined using a transwell technique. The generation of intracellular reactive oxygen species (ROS) was measured using the ROS-sensitive fluoroprobe 2',7'-dichlorodihydrofluorescein diacetate. The activity of NAD(P)H oxidase was assessed by lucigenin enhanced chemiluminescence. Activation of p38 mitogen-activated protein kinase (MAPK) was determined by western blotting. 3. The results showed that pioglitazone dose-dependently inhibited the migration of VSMC induced by Hcy. This was not reversed by the PPAR-gamma antagonist GW9662. In addition, pretreatment with the NAD(P)H oxidase inhibitor diphenylene iodonium (DPI), the free radical scavenger N-acetylcysteine and the p38 MAPK inhibitor SB202190 blocked Hcy-induced VSMC migration. Furthermore, we observed that pioglitazone suppressed Hcy-induced intracellular ROS production; similar effects were observed with DPI and NAC. Pioglitazone attenuated Hcy-induced activation of NAD(P)H oxidase. Moreover, pioglitazone blocked Hcy-induced p38 MAPK phosphorylation; similar effects were observed for DPI, NAC and SB202190. 4. The data demonstrate that pioglitazone inhibits Hcy-induced VSMC migration that is independent of PPAR-gamma. Furthermore, part of the biological effect of pioglitazone involves a decrease in the levels of NAD(P)H oxidase derived-ROS and p38 MAPK activation.