化瘀明目方下调PI3K/Akt/mTOR-HIF-1α/VEGFA信号通路干预DR大鼠视网膜血管新生  

Huayu Mingmu Prescription Downregulates PI3K/Akt/mTOR-HIF-1α/VEGFA Signaling Pathway to Intervene in Retinal Angiogenesis of DR Rats

作　　者：马孝秋 赵磊[2] 周慧敏 郑方卉 杨国庆 左韬[2] 马贤德[3] MA Xiaoqiu;ZHAO Lei;ZHOU Huimin;ZHENG Fanghui;YANG Guoqing;ZUO Tao;MA Xiande(The Second Clinical College of Liaoning University of Traditional Chinese Medicine(TCM),Shenyang 110000,China;The Second Affiliated Hospital of Liaoning University of TCM,Shenyang 110000,China;Teaching and Experimental Center of Liaoning University of TCM,Shenyang 110000,China)

机构地区：[1]辽宁中医药大学第二临床学院,沈阳110000 [2]辽宁中医药大学附属第二医院,沈阳110000 [3]辽宁中医药大学教学实验中心,沈阳110000

出　　处：《中国实验方剂学杂志》2025年第8期78-87,共10页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金项目(82104714);国家中医药管理局“中医药循证能力建设项目”(2019XZZX-YK008)。

摘　　要：目的:观察化瘀明目方对糖尿病视网膜病变(DR)大鼠视网膜新生血管生成及磷脂酰肌醇3-激酶/蛋白激酶B/哺乳动物雷帕霉素靶蛋白(PI3K/Akt/mTOR)-缺氧诱导因子-1α/血管内皮生长因子A(HIF-1α/VEGFA)信号通路的影响。方法:64只SPF级雄性SD大鼠,随机选取11只为正常组,其余53只采用高糖高脂饮食联合小剂量链脲佐菌素(STZ)诱导法复制2型糖尿病(T2DM)大鼠模型,大鼠糖尿病病程12周时进行DR模型评价,将大鼠分为模型组,化瘀明目方低、中、高剂量组(9.29、18.57、37.14 g·kg^(-1)),羟苯磺酸钙组(0.16 g·kg^(-1)),每组10只,并予相应剂量化瘀明目方和羟苯磺酸钙灌胃,正常组和模型组予等容生理盐水灌胃,连续8周。眼底照相观察眼底血管改变,苏木素-伊红(HE)染色观察视网膜组织病理形态改变,视网膜血管网铺片过碘酸雪夫(PAS)染色观察视网膜微血管病理改变,免疫荧光(IF)检测视网膜组织VEGFA、血管生成素-2(Ang-2)表达,蛋白免疫印迹法(Western blot)检测视网膜组织PI3K、Akt、mTOR、HIF-1α、VEGFA、VEGFR2蛋白表达,实时荧光定量聚合酶链式反应(Real-time PCR)检测视网膜组织PI3K、Akt、mTOR、HIF-1α、VEGFA、VEGFR2 mRNA表达。结果:与正常组比较,模型组大鼠视网膜组织病理改变显著,可见无细胞毛细血管出现,内皮细胞(E)增生及周细胞(P)显著丢失(P<0.01),E/P显著上升(P<0.01),视网膜组织PI3K、Akt、mTOR、HIF-1α、VEGFA、VEGFR2蛋白及mRNA的表达量显著升高(P<0.01),Ang-2蛋白表达量显著升高(P<0.01);与模型组比较,各治疗组大鼠视网膜组织病理改变减轻,内皮细胞增生及周细胞丢失明显缓解(P<0.05,P<0.01),其中化瘀明目方高剂量及羟苯磺酸钙组E/P显著降低(P<0.01),各治疗组视网膜组织PI3K、Akt、mTOR、HIF-1α、VEGFA、VEGFR2蛋白及mRNA的表达量明显降低(P<0.05,P<0.01),Ang-2蛋白表达量显著降低(P<0.01)。结论:化瘀明目方能够干预DR大鼠视网膜新生�Objective:To observe the effect of Huayu Mingmu prescription on retinal angiogenesis and phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin(PI3K/Akt/mTOR)-hypoxia inducible factor-1α/vascular endothelial growth factor A(HIF-1α/VEGFA)signaling pathway in diabetic retinopathy(DR)rats.Methods:Sixty-four SPF-grade male SD rats were used in the study.Eleven rats were randomly selected as the normal group,while the remaining 53 rats were fed a high-sugar,high-fat diet combined with low-dose streptozotocin(STZ)intraperitoneal injection to establish a type 2 diabetes mellitus(T2DM)rat model.DR model evaluation was performed after 12 weeks of diabetes.The rats were then divided into model,low-dose,medium-dose,and high-dose groups of Huayu Mingmu prescription(9.29,18.57,37.14 g·kg^(-1)),and a calcium dobesilate group(0.16 g·kg^(-1)),with 10 rats in each group.The rats were orally administered the corresponding doses of Huayu Mingmu prescription and calcium dobesilate.The normal and model groups received equal volumes of physiological saline via gavage for 8 consecutive weeks.Retinal vascular changes were observed through fundus photography,and pathological changes in retinal tissue were evaluated using hematoxylin-eosin(HE)staining.Retinal microvascular pathological changes were examined through retinal vascular network preparation and periodic acid-Schiff(PAS)staining.Immunofluorescence(IF)was used to detect the expression of VEGFA and angiopoietin-2(Ang-2)in retinal tissue.Western blot was employed to detect the protein expression of PI3K,Akt,mTOR,HIF-1α,VEGFA,and VEGFR2 in retinal tissue.Real-time fluorescence quantitative polymerase chain reaction(Real-time PCR)was used to assess the mRNA expression of PI3K,Akt,mTOR,HIF-1α,VEGFA,and VEGFR2 in retinal tissue.Results:Compared with the normal group,the model group exhibited significant pathological changes in retinal tissue,including the appearance of acellular capillaries,as well as significant endothelial cell(E)proliferation and pericyte

关 键 词：化瘀明目方 视网膜血管新生 磷脂酰肌醇3-激酶/蛋白激酶B/哺乳动物雷帕霉素靶蛋白(PI3K/Akt/mTOR)信号通路 缺氧诱导因子-1α(HIF-1α) 血管内皮生长因子A(VEGFA) 

分 类 号：R289[医药卫生—方剂学] R276[医药卫生—中药学] R771[医药卫生—中医学]