黄芪甲苷对大鼠平滑肌细胞及颈动脉内膜增生的影响及机制研究  

Effect of asragaloside IV on vascular smooth muscle cells and neointima hyperplasia in rat carotid artery and their mechanism

作　　者：尉希清[1] 刘帅[1] 牛珩[1] 胡玲爱[1] 张金国[1] 

机构地区：[1]济宁医学院附属医院心内科,山东省心脏疾病诊疗重点实验室,山东济宁272129

出　　处：《中草药》2016年第19期3432-3440,共9页Chinese Traditional and Herbal Drugs

基　　金：山东省自然科学基金项目(ZR2011HL006)

摘　　要：目的通过细胞培养及建立大鼠颈动脉球囊损伤模型,观察黄芪甲苷对血管平滑肌细胞(VSMCs)及血管内皮损伤后内膜增生的影响,并初步探讨黄芪甲苷抑制内膜增生的可能机制。方法采用组织贴块法培养大鼠VSMCs,以重组大鼠肿瘤坏死因子(TNF-α,100 ng/m L)作为刺激因素,建立体外VSMCs增殖模型,应用CCK-8法检测黄芪甲苷(0、0.5、5、25、50μg/m L)对TNF-α诱导的VSMCs增殖能力的影响。采用Fogarty(2F)球囊导管制备大鼠颈总动脉球囊损伤模型,SD大鼠随机分为假手术组、模型组、黄芪甲苷(20、40、60 mg/kg)组,各组于造模前3 d开始ig给药,连续给药17 d,术后第15天,取损伤颈总动脉进行HE染色,光镜下观察血管内膜的形态学变化,并用计算机图像分析系统测定管腔面积、内膜面积、内膜面积/中膜面积;采用免疫组织化学法检测内膜增殖细胞核抗原(PCNA)和血小板源性生长因子(PDGF-BB)的表达。结果在TNF-α刺激下,VSMCs增殖活性与对照组相比均明显提高,差异显著(P<0.01),经黄芪甲苷预处理后,各质量浓度黄芪甲苷组细胞增殖均受到抑制,与TNF-α组相比差异显著(P<0.05),且抑制作用具有时间和浓度依赖性。颈总动脉球囊损伤模型中,模型组大鼠颈动脉内膜增生明显,管腔狭窄,与假手术组比较,内膜面积、内膜面积/中膜面积增大(P<0.01),管腔面积减小(P<0.01);与模型组比较,黄芪甲苷各剂量组内膜面积、内膜面积/中膜面积减小(P<0.05、0.01),管腔面积增大(P<0.05、0.01)。与假手术组比较,模型组大鼠颈总动脉血管内膜PCNA、PDGF-BB的阳性表达明显升高(P<0.01);与模型组比较,黄芪甲苷各组大鼠颈总动脉内膜PCNA、PDGF-BB的阳性表达明显降低(P<0.05),且呈剂量依赖性。结论黄芪甲苷能够以时间和剂量依赖方式抑制TNF-α诱导的VSMCs增殖;黄芪甲苷能够抑制颈总动脉球囊损伤大鼠颈总动脉内膜的增生,黄芪甲苷抑制生长因子PObjective To investigate the effects ofasragaloside IV （AS-IV） on cultured vascular smooth muscle cells （VSMCs） and neointima hyperplasia in the carotid artery of rats after ballon injury and explore the inhibitory mechanisms. Methods For next related researches, primary cultures of VSMCs were prepared from the thoracoabdominal aorta of rats using explant method. Taking the recombinant rat tumor necrosis factor （TNF-α, 100 ng/mL ） as the stimulating factor, the model of VSMCs proliferation was established by TNF-ct inducer in vitro and the effects of AS-IV （0, 0.5, 5, 25, and 50μg/mL） on the VSMCs proliferation induced by TNF-a were determined by CCK-8 method. Rat carotid artery balloon injury model was prepared by Fogarty （2F） balloon catheter. Fifty healthy male Sprague-Dawley rats were randomly divided into five groups： a Sham-operation group （Sham）, a model group （model）, and three AS-IV-treated （20, 40, and 60 mg/kg） groups. Three days before the surgery, 1% CMC, AS-IV 20, 40, and 60 mg/kg were ig administered to each group once daily for continuous 17 d. Fifteen days after the surgery, rats were killed, and the carotid arterys were harvested. Hematoxylin-elsin staining was carried out to observe the pathomorphological change in vascular intima. The measurement of lumen area, intimal area, intimal area/medium film area were measured by computer image analysis system; Immunohistochemistry staining was performed to measure the expression of proliferating cell nuclear antigen （PCNA） and platelet-derived growth factor-BB （PDGF-BB）. Results The proliferative activity of VSMCs was obviously increased in the TNF-a group under the stimulation of TNF-u. There was a significant difference compared with the control group （P 〈 0.01）. However, when pretreated with AS-IV ahead of time, we found that AS-IV significantly inhibited TNF-a-induced VSMCs proliferation in a dose- and time-dependent manner compared to the TNF-u group. Compared with the model group, the area of int

关 键 词：黄芪甲苷 球囊损伤 内膜增生 血管平滑肌细胞 再狭窄 

分 类 号：R285.5[医药卫生—中药学]