益气活血通络方调控Drp1抑制血管新生减轻单侧输尿管梗阻大鼠肾纤维化  

Experimental Study of Yiqi Huoxue Tongluo Recipe Inhibits Angiogenesis to Attenuate Renal Fibrosis in UUO Rats

作　　者：张孟娟 王洪双 许畅 刘佳志 何珍 徐贺朋 周文平 钟艳 王筝 王香婷 ZHANG Mengjuan;WANG Hongshuang;XU Chang;LIU Jiazhi;HE Zhen;XU Hepeng;ZHOU Wenping;ZHONG Yan;WANG Zheng;WANG Xiangting(Hebei University of Chinese Medicine,Shijiazhuang 050200 Hebei,China;Hebei Key Laboratory of Integrative Medicine on Liver-Kidney Patterns Research,Shijiazhuang 050091 Hebei,China;Institute of Integrative Medicine,Hebei University of Chinese Medicine,Shijiazhuang 050091 Hebei,China)

机构地区：[1]河北中医药大学,河北石家庄050200 [2]河北省中西医结合肝肾病证研究重点实验室,河北石家庄050091 [3]河北中医药大学中西医结合研究所,河北石家庄050091

出　　处：《中药新药与临床药理》2023年第10期1354-1362,共9页Traditional Chinese Drug Research and Clinical Pharmacology

基　　金：河北省自然科学基金中医药联合基金重点项目(H2022423351);河北中医学院研究生创新资助项目(XCXZZSS2022009)。

摘　　要：目的探讨益气活血通络方(黄芪、丹参、川芎、地龙等)调控动力相关蛋白1(Drp1)抑制血管新生,减轻单侧输尿管梗阻(UUO)大鼠肾纤维化的作用机制。方法将雄性Wistar大鼠随机分为假手术组、模型组、氯沙坦钾组(20 mg·kg^(-1))、益气活血通络方组(1.92 g·kg^(-1)颗粒剂),每组12只。采用单侧输尿管结扎的方式复制肾纤维化大鼠模型。术后第1天开始灌胃给药,每日1次,连续14 d。采用HE、Masson染色法观察肾组织病理变化;采用免疫组化法检测肾组织中Ⅰ型胶原蛋白(ColⅠ)、α-平滑肌肌动蛋白(α-SMA)、波形蛋白(Vimentin)、血管内皮生长因子A(VEGF-A)、CD34的表达;Western Blot法检测肾组织中α-SMA、Vimentin、VEGF-A、血管内皮生长因子受体2(VEGFR2)、Drp1及Ser616位点磷酸化Drp1(p-Drp1S616)蛋白表达;Real-time PCR法检测肾组织中VEGF-A、VEGFR2、CD34 mRNA表达。结果与假手术组比较,模型组大鼠肾组织出现明显的肾小管上皮细胞变性、坏死、脱落,大量炎性细胞浸润,肾小管萎缩,肾间质胶原纤维沉积明显增多(P<0.05);肾间质中的ColⅠ、α-SMA、Vimentin、CD34蛋白表达量及胞质中VEGF-A蛋白表达量明显升高(P<0.05);肾组织中α-SMA、Vimentin、VEGF-A、VEGFR2及p-Drp1/Drp1蛋白表达水平均明显升高(P<0.05),VEGF-A、VEGFR2、CD34 mRNA表达水平明显升高(P<0.05)。与模型组比较,益气活血通络方组大鼠的肾脏炎性细胞浸润有一定程度减轻,胶原纤维沉积明显减少(P<0.05);肾间质中的ColⅠ、α-SMA、Vimentin、CD34蛋白表达量及胞质中VEGF-A蛋白表达量明显降低(P<0.05);肾组织中α-SMA、Vimentin、VEGF-A、VEGFR2及p-Drp1/Drp1蛋白表达水平均明显降低(P<0.05),VEGF-A、VEGFR2、CD34 mRNA表达水平明显降低(P<0.05)。结论益气活血通络方能够减轻UUO大鼠肾纤维化水平,其机制可能与下调Drp1磷酸化水平、抑制血管新生有关。Objective To investigate the mechanism of action of Yiqi Huoxue Tongluo Recipe(Astragali Radix,Salviae Miltiorrhizae Radix et Rhizoma,Chuanxiong Rhizoma,Pheretima,etc.)to regulate the inhibition of angiogenesis by dynamin-related protein 1(Drp1)and to attenuate renal fibrosis in rats with unilateral ureteral obstruction(UUO).Methods Male Wistar rats were randomly divided into a sham-operation group,a model group,a Losartan Potassium group(20 mg·kg^(-1)),and a Yiqi Huoxue Tongluo Recipe group(1.92 g·kg^(-1) in granules),with 12 rats in each group.The drug was administered by gavage starting from the first day after surgery,once a day for 14 days.The pathological changes of renal tissue were observed by HE and Masson staining;and immunohistochemistry was used to detect expressions of collagen typeⅠ(ColⅠ),α-SMA,Vimentin in renal tissue,VEGF-A,CD34 in kidney tissue;the protein expressions ofα-smooth muscle actin(α-SMA),vimentin,vascular endothelial growth factor A(VEGF-A),vascular endothelial growth factor receptor 2(VEGFR2),Drp1 and phosphorylated Drp1(p-Drp1S616)at Ser616 site in renal tissue were detected by Western Blot method;Real-time PCR was used to detect mRNA expressions of VEGF-A,VEGFR2,CD34 in renal tissue.Results Compared with the sham-operation group,the renal tissue of rats in the model group showed obvious degeneration,necrosis and detachment of renal tubular epithelial cells,infiltration of a large number of inflammatory cells,renal tubular atrophy,and obvious increase in collagen fibre deposition in rat renal interstitium(P<0.05);the protein expression levels of ColⅠ,α-SMA,Vimentin,and CD34 in the interstitial tissue of the rat kidneys,and that of VEGF-A in the cytoplasm,were obviously elevated(P<0.05);protein expression levels ofα-SMA,Vimentin,VEGF-A,VEGFR2 and p-Drp1/Drp1 in rat kidney tissue were all significantly elevated(P<0.05),and the mRNA expression levels of VEGF-A,VEGFR2,and CD34 were significantly elevated(P<0.05).Compared with the model group,the infiltration of renal inflamm

关 键 词：益气活血通络方 慢性肾脏病 单侧输尿管梗阻 肾纤维化 血管新生 动力相关蛋白1 大鼠 

分 类 号：R285.5[医药卫生—中药学]