Chen Li, Wang Wen-yi, Wang Yi-ping
State Key Laboratory of New Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China.
Acta Pharmacol Sin. 2009 Sep;30(9):1245-52. doi: 10.1038/aps.2009.122. Epub 2009 Aug 24.
To understand the effects of lithospermic acid (LA), a potent antioxidant from the water-soluble extract of Salvia miltiorrhiza, on the migration and proliferation of rat thoracic aorta vascular smooth muscle cells (VSMCs).
VSMC migration, proliferation, DNA synthesis and cell cycle progression were investigated by transwell migration analysis, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, bromodeoxyuridine (BrdU) incorporation assay, and flow cytometric detection, respectively. Intracellular reactive oxygen species (ROS) generation was detected using 2',7'-dichlorofluorescin diacetate (DCFH-DA). The expression of cyclin D1 protein and matrix metalloproteinase-9 (MMP-9) protein, as well as the phosphorylation state of ERK1/2, were determined using Western blots. The activity of MMP-9 and the expression of MMP-9 mRNA were assessed by gelatin zymography analysis and RT-PCR, respectively.
LA (25-100 micromol/L) inhibited both lipopolysaccharide (LPS)- and fetal bovine serum (FBS)-induced ROS generation and ERK1/2 phosphorylation. By down-regulating the expression of cyclin D(1) and arresting cell cycle progression at the G(1) phase, LA inhibited both VSMC proliferation and DNA synthesis as induced by 5% FBS. Furthermore, LA attenuated LPS-induced VSMC migration by inhibiting MMP-9 expression and its enzymatic activity.
LA is able to inhibit FBS-induced VSMC proliferation and LPS-induced VSMC migration, which suggests that LA may have therapeutic effects in the prevention of atherosclerosis, restenosis and neointimal hyperplasia.
了解丹参水溶性提取物中的一种强效抗氧化剂——紫草酸（LA）对大鼠胸主动脉血管平滑肌细胞（VSMC）迁移和增殖的影响。
分别采用Transwell迁移分析、3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐（MTT）法、溴脱氧尿苷（BrdU）掺入法和流式细胞术检测VSMC迁移、增殖、DNA合成及细胞周期进程。使用2',7'-二氯荧光素二乙酸酯（DCFH-DA）检测细胞内活性氧（ROS）的产生。采用蛋白质印迹法测定细胞周期蛋白D1蛋白和基质金属蛋白酶-9（MMP-9）蛋白的表达以及ERK1/2的磷酸化状态。分别通过明胶酶谱分析和RT-PCR评估MMP-9的活性和MMP-9 mRNA的表达。
LA（25 - 100 μmol/L）抑制脂多糖（LPS）和胎牛血清（FBS）诱导的ROS产生以及ERK1/2磷酸化。通过下调细胞周期蛋白D1的表达并使细胞周期进程停滞在G1期，LA抑制5% FBS诱导的VSMC增殖和DNA合成。此外，LA通过抑制MMP-9表达及其酶活性减弱LPS诱导的VSMC迁移。
LA能够抑制FBS诱导的VSMC增殖和LPS诱导的VSMC迁移，这表明LA在预防动脉粥样硬化、再狭窄和新生内膜增生方面可能具有治疗作用。
