益气通络颗粒促气虚血瘀证脑梗大鼠血管新生的作用研究
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摘要
探讨益气通络颗粒对脑梗大鼠气虚血瘀证的作用。构建多发性脑梗气虚血瘀大鼠模型,随机分为假手术组,气虚血瘀脑梗组,金纳多组,补阳还五汤组,益气通络颗粒高、低剂量组,脑梗术后1周进行睡眠剥夺,6周后进行舌象脉象采集,取血、脑组织分别进行血液、蛋白组学及免疫组化相关指标分析。结果显示,益气通络颗粒高剂量组可明显降低脑梗神经功能评分,改善气虚血瘀大鼠的舌面色彩饱和度(G和B),益气通络颗粒低剂量组可显著改善气虚血瘀大鼠的舌面色彩饱和度B;益气通络颗粒高剂量组可明显恢复气虚血瘀大鼠的脉搏幅度,且益气通络颗粒高、低剂量对血液中活化部分凝血活酶时间(APTT)及血浆凝血酶原时间(PT)有明显的恢复。蛋白组学结果表明益气通络颗粒显著地促进了气虚血瘀脑梗大鼠脑内血管新生,免疫组化结果验证了益气通络颗粒调节了血管新生标志物VEGFR2的表达。因此该研究得出结论:益气通络颗粒对气虚血瘀脑梗大鼠有明显的缓解作用,其可能的机制是益气通络颗粒促进气虚血瘀脑梗大鼠脑血管新生。
This research was aimed to evaluate the protective effect and potential mechanism of Yiqi Tongluo Particles(YQTLs).Firstly,an animal model of multiple cerebral infarction(MCI) with Qi deficiency and blood stasis was established.Rats were randomly divided into six groups:SHAM group,Vehicle group,Buyang Huanwu decoction original group(BYHWO),EGb761 group,high and low dose of YQTLs group.Rats underwent sleep deprivation after one week of MCI and the tongues and pulses of rats after six weeks of sleep deprivation were detected,followed by collecting blood to analysis the blood coagulation.Differential expression of angiogenesis associated proteins was examined using proteomic research and verified by immunohistochemical.Results showed that neurological function score was obviously declined,G and B value of tongue surface was increased significantly and the pulse distension,the activated partial thromboplatin time(APTT) as well as prothrombin time(PT) were recovered following YQTLs 7.56 g·kg-1 treatment.Furthermore,G value of tongue surface,APTT and PT were also improved by YQTLs 3.78 g·kg-1.The results of proteomic technology showed that proteins associated with angiogenesis were reversed compared with Vehicle group.Moreover,the expression of VEGFR2 from immunohistochemical was promoted after YQTLs treatment.The MCI with Qi deficiency and blood stasis was alleviated obviously following YQTLs treatment and the possible mechanism was that YQTLs may enhance angiogenesis during cerebral ischemia.
This research was aimed to evaluate the protective effect and potential mechanism of Yiqi Tongluo Particles(YQTLs).Firstly,an animal model of multiple cerebral infarction(MCI) with Qi deficiency and blood stasis was established.Rats were randomly divided into six groups:SHAM group,Vehicle group,Buyang Huanwu decoction original group(BYHWO),EGb761 group,high and low dose of YQTLs group.Rats underwent sleep deprivation after one week of MCI and the tongues and pulses of rats after six weeks of sleep deprivation were detected,followed by collecting blood to analysis the blood coagulation.Differential expression of angiogenesis associated proteins was examined using proteomic research and verified by immunohistochemical.Results showed that neurological function score was obviously declined,G and B value of tongue surface was increased significantly and the pulse distension,the activated partial thromboplatin time(APTT) as well as prothrombin time(PT) were recovered following YQTLs 7.56 g·kg-1 treatment.Furthermore,G value of tongue surface,APTT and PT were also improved by YQTLs 3.78 g·kg-1.The results of proteomic technology showed that proteins associated with angiogenesis were reversed compared with Vehicle group.Moreover,the expression of VEGFR2 from immunohistochemical was promoted after YQTLs treatment.The MCI with Qi deficiency and blood stasis was alleviated obviously following YQTLs treatment and the possible mechanism was that YQTLs may enhance angiogenesis during cerebral ischemia.
引文
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[18]Srinivasan S,Meyer R D,Lugo R,et al.Identification of PDCL3as a novel chaperone protein involved in the generation of functional VEGF receptor 2[J].J Biol Chem,2013,288(32):23171.
[19]Hermann D M,Chopp M.Promoting brain remodelling and plasticity for stroke recovery:therapeutic promise and potential pitfalls of clinical translation[J].Lancet Neurol,2012,11(4):369.
[20]Paré-Brunet L,Sebio A,Salazar J,et al.Genetic variations in the VEGF pathway as prognostic factors in metastatic colorectal cancer patients treated with oxaliplatin-based chemotherapy[J].Pharmacogenomics J,2015,15(5):397.
[21]Zhang Q,Yu C,Peng S,et al.Autocrine VEGF signaling promotes proliferation of neoplastic Barrett's epithelial cells through a PLC-dependent pathway[J].Gastroenterology,2014,146(2):461.
[22]Choi S H,Lee H J,Jin Y B,et al.MMP9 processing of HSPB1regulates tumor progression[J].PLoS ONE,2014,9(1):e85509.
[23]Srinivasan S,Chitalia V,Meyer R D,et al.Hypoxia-induced expression of phosducin-like 3 regulates expression of VEGFR-2and promotes angiogenesis[J].Angiogenesis,2015,18(4):449.
[2]李莹,王磊沙,刘建勋,等.脑梗死气虚血瘀证大鼠模型表征及病理生理变化研究[J].中国中药杂志,2018,43(4):786.
[3]骆磊,章培林.益气活血中药治疗进展性脑梗死的早期疗效观察[J].中国中药杂志,2009,34(10):1295.
[4]Elisa C,Giada P,Alessandro V,et al.Cerebral collateral circulation in experimental ischemic stroke[J].Exp Trans Stroke Med,2016,8(1):1.
[5]Wang J P,Yu L,Jiang C,et al.Bone marrow mononuclear cells exert long-term neuroprotection in a rat model of ischemic stroke by promoting arteriogenesis and angiogenesis[J].Brain Behav Immun,2013,34:56.
[6]Wang J P,Fu X J,Jiang C,et al.Bone marrow mononuclear cell transplantation promotes therapeutic angiogenesis via upregulation of the VEGF VEGFR2 signaling pathway in a ratmodel of vascular dementia[J].Behav Brain Res,2014,265:171.
[7]张钢,张华健,刘军锋,等.益气通络颗粒治疗中风病(气虚血瘀证)Ⅲ期临床试验[J].临床研究,2012,28(10):743.
[8]郑咏秋,姚明江,刘建勋,等.华佗再造浸膏对大鼠局灶性脑缺血/再灌注血脑屏障损伤的保护作用及机制研究[J].中国中药杂志,2013,38(4):585.
[9]张玉琴,李鸷,李煌,等.栝楼桂枝颗粒激活Nrf2信号通路减轻脑缺血再灌注损伤大鼠氧化应激损伤作用[J].中国实验方剂学杂志,2017,23(21):112.
[10]马博,薛冰洁,陈进成,等.基于定量蛋白质组技术探索补阳还五汤治疗脑梗死气虚血瘀证的作用机制[J].中国中药杂志,2018,43(11):2199.
[11]李彦丽,武如冰,于健,等.益气活血通脉汤加味对下肢深静脉血栓后综合征(气虚血瘀证)再通的影响及作用机制[J].中国实验方剂学杂志,2018,24(3):202.
[12]Zivin J A,De Girolami U,Kochhar A,et al.A model for quantitative evaluation of embolic stroke therapy[J].Brain Res,1987,435:305.
[13]任建勋,徐立,林成仁,等.塞络通胶囊在络脉瘀阻证脑梗死大鼠缺血时间窗外给药的保护作用[J].中华中医药杂志,2012,27(12):3060.
[14]La Valley D J,Zanotelli M R,Bordeleau F,et al.Matrix stiffness enhances VEGFR-2 internalization,signaling,and proliferation in endothelial cells[J].Converg Sci Phys Oncol,2017,3:1.
[15]Zhao M,Bai H,Wang E,et al.Electrical stimulation directly induces pre-angiogenic responses in vascular endothelial cells by signaling through VEGF receptors[J].J Cell Sci,2004,117(Pt3):397.
[16]Del Valle-Pérez B,Martínez V G,Lacasa-Salavert C,et al.Filamin B plays a key role in vascular endothelial growth factor-induced endothelial cell motility through its interaction with Rac-1and Vav-2[J].J Biol Chem,2010,285(14):10748.
[17]Xu H,Czerwinski P,Hortmann M,et al.Protein kinase C alpha promotes angiogenic activity of human endothelial cells via induction of vascular endothelial growth factor[J].Cardiovasc Res,2008,78(2):349.
[18]Srinivasan S,Meyer R D,Lugo R,et al.Identification of PDCL3as a novel chaperone protein involved in the generation of functional VEGF receptor 2[J].J Biol Chem,2013,288(32):23171.
[19]Hermann D M,Chopp M.Promoting brain remodelling and plasticity for stroke recovery:therapeutic promise and potential pitfalls of clinical translation[J].Lancet Neurol,2012,11(4):369.
[20]Paré-Brunet L,Sebio A,Salazar J,et al.Genetic variations in the VEGF pathway as prognostic factors in metastatic colorectal cancer patients treated with oxaliplatin-based chemotherapy[J].Pharmacogenomics J,2015,15(5):397.
[21]Zhang Q,Yu C,Peng S,et al.Autocrine VEGF signaling promotes proliferation of neoplastic Barrett's epithelial cells through a PLC-dependent pathway[J].Gastroenterology,2014,146(2):461.
[22]Choi S H,Lee H J,Jin Y B,et al.MMP9 processing of HSPB1regulates tumor progression[J].PLoS ONE,2014,9(1):e85509.
[23]Srinivasan S,Chitalia V,Meyer R D,et al.Hypoxia-induced expression of phosducin-like 3 regulates expression of VEGFR-2and promotes angiogenesis[J].Angiogenesis,2015,18(4):449.