BACKGROUND:Sulfasalazine(SSZ) is an anti-inflammatory and immunomodulatory drug of which mechanism is due to inhibition of NF-κB. In recent years, the transcription factor Nrf2 has been reported to regulate the expression of heme oxygenase-1 (HO-1). We hypothesized that SSZ might reduce neointimal hyperplasia in rat carotid artery balloon injury model and if so, investigated what the underlyng mechanism is. METHODS: Rat vascular smooth muscle cells (rVSMC) were cultured in the presence of SSZ(100 μM). Cell cycle changes and degree of apoptosis was evaluated by FACS analyses. NF-κB activation was evaluated by EMSA and expression of HO-1 and Nrf2 was evaluated by Western blot. For in vivo experiments, rat carotid artery balloon injury model was used to test the effects of SSZ and vehicle on neointimal growth. Rats were divided into 2 groups (n=5/group): SSZ(86 mg/kg/d) and vehicle(normal saline). Drugs were administered via intraperitoneal injection from 3 days before to 2 weeks after balloon injury. RESULTS:1) In vitro, SSZ significantly increased TNFα–mediated apoptosis and induced cell cycle arrest. Apoptosis induction by SSZ was reversed by antioxidant,NAC, suggesting effect was mediated by ROS. Although SSZ significantly inhibited TNFα-mediated activation of NF-κB,the concentration is relatively higher cell cycle arrest and apoptosis (200 uM-1 mM), which suggest that another mechanisms work other than NF-kB inhibition. We found that SSZ significantly increased expression of HO-1 via JNK-Nrf2 complex translocation. The HO-1 inihibitor, ZnPP (100 nM) totally reversed the cell cycle arrest by SSZ, which means that cell cycle arrest is by HO-1. Transfection of Adv-DN Nrf2 reversed expression of HO-1 2) In vivo, SSZ redued neointimal hyperplasia after balloon-injury in rat carotid artery (p<0.05). In SSZ group, expression of HO-1 and JNK was increased in rat carotid artery. Transfection of Adv-DN-Nrf trasfection reversed supression of neointimal hyperplasia by SSZ, which suggests that activation of Nrf2 by SSZ plays important role. CONCLUSIONS: SSZ was found to have a potent inhibiting effect of neointimal hyperplasia mediated by new antioxidant mechanism via HO-1 activation by Nrf2 and JNK pathway.
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