黄芩苷通过降低活性氧生成抑制血管平滑肌细胞伪足形成和迁移

已知黄芩苷（baicalin）通过削弱肌动蛋白相关蛋白（actin-related protein, Arp）2/3复合物的活性抑制血管平滑肌细胞（vascular smooth muscle cell, VSMC）伪足形成和迁移，然而，其抑制该信号途径的机制尚不明确。本研究证明，黄芩苷通过抑制VSMC活性氧（reactive oxygen species，ROS）生成降低Arp2/3活性，发挥阻止细胞伪足形成和迁移的功能。分别利用TRITC 鬼笔环肽和ROS荧光探针标记VSMCs，结果显示，黄芩苷能显著抑制血小板源性生长因子（platelet derived growth factor, PDGF）-BB诱导的VSMC伪足形成和迁移，伴有ROS生成减少。用超氧物歧化酶（superoxide dismutase, SOD）清除胞内过氧化物后，PDGF-BB引发的VSMC伪足形成被逆转，且该过程与降低皮层肌动蛋白微丝（F-actin）成核蛋白Arp2/3活性有关。免疫沉淀分析结果进一步表明，黄芩苷降低p47phox磷酸化水平，与ROS生成减少相一致。体内的实验也表明，黄芩苷（70 mg/kg/d）能有效抑制球囊损伤诱导的大鼠颈总动脉ROS生成。以上结果表明，黄芩苷通过抑制NADPH氧化酶介导的ROS生成，降低细胞皮质区F-actin成核活性，阻止细胞伪足形成、迁移，进而发挥血管保护作用。

Baicalin Inhibits Lamellipodium Formation and Migration via Repressing ROS Production in Vascular Smooth Muscle Cells

Our recent study has demonstrated that baicalin, a herb derived flavonoid compound, inhibits platelet derived growth factor (PDGF) BB induced lamellipodium formation and migration via reducing the activation of actin related protein (Arp)2/3 complex in vascular smooth muscle cells (VSMCs). However, the mechanisms responsible for baicalin regulating this signal pathway are not fully understood. In the present study, our finding suggests that baicalin blocks Arp2/3 activity via reducing reactive oxygen species (ROS) generation, and subsequently inhibits lamellipodium formation and cell migration in VSMCs.Using TRITC-Phalloidin and ROS specific probe, fluorescent staining assays showed that baicalin pretreatment inhibited PDGF-BB induced lamellipodium formation and cell migration, accompanied by less ROS generation. To determine the role of ROS in lamellipodium formation of migrating cells, superoxide dismutase (SOD), a ROS scavenger, was used. Results showed SOD attenuated lamellipodium formation and actin-nucleating activation of Arp2/3, which is essential for lamellipodium formation in VSMCs, suggesting oxidative stress probably has a strong effect on cell migration through regulating cortex actin remodeling. Immunoprecipitation analysis further showed that baicalin dramatically suppressed phosphorylation of p47phox induced by PDGF-BB, which was associated with the decreased ROS generation. Moreover, balloon injury-induced ROS generation in rat carotid arteries also attenuated by administration with baicalin (70 mg/kg/d). These data demonstrate that NADPH oxidase mediated oxidative stress may play a crucial role in the activation of Arp2/3 and cortex actin polymerization in vitro and in vivo. Therapeutic blockade of ROS generation by baicalin may represent a safe and effective approach to prevent vascular injury. Our findings are important for understanding of baicalin involved events that contribute to the development and progression in vascular diseases.