摘要
目的:探讨黄芪-当归配伍对兔血管内膜增生模型血管外膜成纤维细胞(VAF)活化的影响及其作用机制。方法:清洁级家兔,雌雄各半,随机分为假手术组、模型组、单用黄芪组(1 g·kg^(-1))、单用当归组(1 g·kg^(-1))、黄芪-当归1∶1组(黄芪1 g·kg^(-1)+当归1 g·kg^(-1))、黄芪-当归1∶5组(黄芪1 g·kg^(-1)+当归5 g·kg^(-1))、黄芪-当归5∶1组(黄芪5 g·kg^(-1)+当归1 g·kg^(-1))、阿托伐他汀钙组(5 mg·kg^(-1)),采用颈总动脉套管复合高胆固醇饮食制备家兔血管内膜增生模型,假手术组只进行分离操作,不进行套管。术后第1天开始假手术组喂饲普通饲料,其余各组喂饲高胆固醇饲料(2%胆固醇+3%橄榄油+95%普通饲料),同时各用药组给予相应药物灌胃,假手术组、模型组给予等体积蒸馏水灌胃,持续给药28 d。实验结束后腹主动脉采血,马松(Masson)染色观察血管外膜形态学变化,免疫组化法检测外膜局部促炎因子肿瘤坏死因子-α(TNF-α)及白细胞介素-1β(IL-1β)的表达,免疫荧光双标法测定外膜成纤维细胞标记物波形蛋白(Vimentin)及肌成纤维细胞标记物α-平滑肌肌动蛋白(α-SMA)表达反映VAF活化;蛋白免疫印迹法(Western blot)检测相关通路蛋白转化生长因子-β_(1)(TGF-β_(1))、Smad2、磷酸化(p)-Smad2表达。结果:与假手术组比较,模型组血管外膜增生明显,损伤血管局部TNF-α、IL-1β表达显著升高,受损段血管α-SMA、Vimentin、TGF-β_(1)、Smad2、p-Smad2表达均明显升高(P<0.05,P<0.01);与模型组比较,黄芪-当归1∶1、5∶1组可使血管外膜增生减轻,血管局部TNF-α、IL-1β表达降低,α-SMA、Vimentin表达下调,并且可降低TGF-β_(1)、Smad2、p-Smad2表达(P<0.05,P<0.01)。结论:黄芪当归配伍可抑制血管内膜增生模型的外膜增生,抑制外膜成纤维细胞向肌成纤维细胞转化,减轻血管局部炎症反应,以黄芪-当归1∶1、5∶1组的作用明显。其作用机制可能与黄芪当归配伍抑制TGF-β_(1)/Smad2信号通路激活有关。
Objective:To observe the effect of Astragali Radix combined with Angelicae Sinensis Radix on the activation of adventitial fibroblasts in the rabbit model of intimal hyperplasia and decipher the underlying mechanism.Method:Clean white rabbits of the same number of males and females were selected.They were randomly assigned into a sham operation group,a model group,an Astragali Radix group(1 g·kg^(-1)),an Angelicae Sinensis Radix group(1 g·kg^(-1)),and Astragali Radix-Angelicae Sinensis Radix groups at the ratios of 1∶1(Astragali Radix 1 g·kg^(-1)+Angelicae Sinensis Radix 1 g·kg^(-1)),1∶5(Astragali Radix1 g·kg^(-1)+Angelicae Sinensis Radix 5 g·kg^(-1)),and 5∶1(Astragali Radix 5 g·kg^(-1)+Angelicae Sinensis Radix1 g·kg^(-1)),and an atorvastatin calcium group(5 mg·kg^(-1)).The rabbit model of intimal hyperplasia was established by common carotid artery cannula combined with high-cholesterol diet.The sham operation group only underwent the separation operation and not the cannula.The sham operation group had been fed with normal diet and the other groups with high-cholesterol diet(2%cholesterol+3%olive oil+95%common diet)since the first day after operation.At the same time,drugs were administrated in corresponding groups,and the sham operation group and model group were administrated with distilled water(15 mL·kg^(-1))for 28 days.At the end of the experiment,blood was collected from abdominal aorta,and the morphological changes of adventitia were observed by Masson staining.The expression of tumor necrosis factor-α(TNF-α)and interleukin-1β(IL-1β)in adventitia was detected by immunohistochemical method.The expression of adventitial fibroblast marker vimentin and myofibroblast markerα-smooth muscle actin(α-SMA),which can reflect the activation of adventitial fibroblasts,was detected by immunofluorescence double staining.Western blot was employed to determine the protein levels of transforming growth factor-β_(1)(TGF-β_(1)),Smad2,and p-Smad2.Result:Compared with the normal group,the modeling caused adventitial hyperplasia and up-regulated the expression of TNF-α,IL-1β,α-SMA,Vimentin,TGF-β_(1),Smad2,and phosphorylation(p)-Smad2 in the injured vessels(P<0.05,P<0.01).Compared with the model group,Astragali Radix combined with Angelicae Sinensis Radix at the ratios of 1∶1 and 5∶1 alleviated the adventitial hyperplasia and down-regulated the expression of TNF-α,IL-1β,α-SMA,Vimentin,TGF-β_(1),Smad2,and p-Smad2(P<0.05,P<0.01).Conclusion:Astragali Radix combined with Angelicae Sinensis Radix,especially at the ratios of 1∶1 and 5∶1,can ameliorate the adventitial hyperplasia in the rabbit model of intimal hyperplasia by inhibiting the transformation of adventitial fibroblasts to myofibroblasts and reducing the local inflammation of blood vessels.The mechanism may be associated with the inhibition of TGF-β_(1)/Smad2 signaling pathway.
作者
李霞
曹旺
邓常清
LI Xia;CAO Wang;DENG Changqing(Key Laboratory of Hunan Province for Prevention and Treatment of Integrated Traditional Chinese and Western Medicine on Cardiocerebral Diseases,Hunan University of Chinese Medicine,Changsha 410028,China;Hunan Academy of Chinese Medicine,Changsha 410028,China)
出处
《中国实验方剂学杂志》
CAS
CSCD
北大核心
2023年第3期61-69,共9页
Chinese Journal of Experimental Traditional Medical Formulae
基金
国家自然科学基金项目(81874406)。
