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目的观察大黄素对颈动脉狭窄的影响,并对狭窄变化的分子机制进行初步探讨。方法选取72只SD大鼠(200 g)采用随机数字表法分为3组:假手术组(n=24)、损伤组(n=24)、大黄素+损伤组(n=24)。损伤组及大黄素+损伤组利用球囊建立颈动脉狭窄模型;假手术组除无球囊抽拉损伤外其余处理与其他两组一致。术后大黄素+损伤组灌胃大黄素(70 mg/kg);其余两组灌胃等体积溶剂(生理盐水含二甲亚砜)。于第14天取下3组颈总动脉行HE染色;另取第14天3组颈总动脉行二氧乙啶(dihydroethidium,DHE)测量活性氧(reactive oxygen species,ROS)含量,免疫组化染色检测平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)、Ki67及c-myc表达变化,Western blot检测3组胞浆MEK1/2、p-MEK1/2、ERK1/2、p-ERK1/2及核内c-myc含量变化。结果 HE染色显示颈动脉狭窄模型建立成功,14 d新生内膜增殖速度为3个时间点最快(P<0.01)。新生内膜成分主要为平滑肌细胞(vascular smooth muscle cells,VSMCs)。与假手术组比较,损伤组ROS表达量急剧升高(P<0.01),大黄素+损伤组显著下调ROS水平(P<0.01)。与损伤组比较,大黄素+损伤组显著下调增殖蛋白Ki67表达(P<0.01),抑制新生内膜中VSMCs的增殖(P<0.01),降低MAPK-ERK通路活化(P<0.01),减少核内c-myc活化(P<0.01)。结论大黄素能够缓解大鼠颈动脉狭窄,其机制可能是大黄素下调ROS/MAPK/ERK/c-myc通路活化,降低增殖蛋白Ki67表达,从而抑制新生内膜中VSMCs的过度增殖。
Objective To observe the effect of emodin on carotid artery stenosis, and to investigate the molecular mechanism of stenosis. Methods Seventy two SD rats (200 g) were randomly divided into three groups: sham operation group (n = 24), injury group (n = 24) and emodin + injury group (n = 24). In the injury group and the emodin + injury group, the balloon was used to establish the model of carotid artery stenosis. The rats in the sham operation group were the same as those in the other two groups except without balloon injury. After emodin + injury group emodin (70 mg / kg); the other two groups were given an equal volume of solvent (saline containing dimethyl sulfoxide). Three groups of common carotid arteries were removed for HE staining on the 14th day. The other three groups of carotid artery on the 14th day were examined for the content of reactive oxygen species (ROS) by dihydroethidium (DHE) The expression of α-smooth muscle actin (α-SMA), Ki67 and c-myc were detected by Western blotting. The expressions of MEK1 / 2, p-MEK1 / 2, ERK1 / 2 and p-ERK1 / 2 and nuclear c-myc content changes. Results Hematoxylin-eosin staining showed that the model of carotid artery stenosis was successfully established. The rate of intimal hyperplasia was the fastest at 14 days (P <0.01). The neointimal components are mainly vascular smooth muscle cells (VSMCs). Compared with the sham operation group, ROS expression in the injury group was significantly increased (P <0.01), and emodin + injury group significantly reduced the ROS level (P <0.01). Compared with the injury group, emodin + injury group significantly down-regulated the expression of Ki67 (P <0.01), inhibited the proliferation of neointimal VSMCs (P <0.01), decreased the activation of MAPK-ERK pathway (P <0.01) Intra-c-myc activation (P <0.01). Conclusion Emodin can attenuate carotid artery stenosis in rats. The mechanism may be that emodin can down-regulate the activation of ROS / MAPK / ERK / c-myc pathway and decrease the expression of proliferating cell protein Ki67, thereby inhibiting over proliferation of neointimal VSMCs.