蝎毒多肽提取物抗肿瘤血管生成作用的实验研究

目的探讨东亚钳蝎蝎毒的多肽提取物PESV的抗血管生成活性和对肿瘤生长的抑制作用。方法①用不同浓度的PESV(4～20mg·L-1)作用于人脐静脉内皮细胞(HUVEC),采用BrdU参入的ELISA法观察HUVEC增殖活性和凋亡水平变化,流式细胞术检测凋亡细胞比例,免疫组化法检测Bal和Bax表达。②观察PESV对鸡胚尿囊膜(CAM)新生血管生成的影响。③皮下注射PESV(0.3mg·kg-1),观察对S180肉瘤和H22肝癌荷瘤小鼠肿瘤生长、肿瘤血管生成和血管生成因子(VEGF和bFGF)表达的影响。结果①体外实验显示,PESV在8～20mg·L-1范围明显抑制HUVEC的增殖活性(与对照组比较,P<0.01),而对乳腺癌细胞MDAMB231的增殖无影响;PESV作用后HUVEC凋亡细胞比例较对照组增加,P<0.05,Bax表达增加;Bcl2表达降低。②0.5mg/CAM和0.8mg/CAM的PESV能明显抑制CAM新生血管的形成。③体内实验显示PESV能明显抑制小鼠S180肉瘤和H22肝癌的肿瘤生长和血管生成水平,并降低肿瘤组织内血管生成相关因子VEGF和bFGF的表达。结论PESV具有良好的体内和体外抗肿瘤血管生成活性,并籍此抑制肿瘤的生长。     

二母颗粒对小鼠Lewis肺癌模型的肿瘤血管生成的影响

目的 观察二母颗粒对C57BL/6小鼠体内Lewis肺癌生长、转移以及血管生成的影响及其作用机制.方法 制备C57 BL/6小鼠Lewis肺癌模型,随机分为二母颗粒高、中、低(3、2、1g·kg-1生药)剂量组、环磷酰胺组、沙利度胺组、模型组.分别给予相应药物后,观察各组小鼠的肿瘤生长情况及肺转移发生率,采用免疫组化方法观察瘤内血管内皮细胞生长因子(VEGF)和微血管密度(MVD)的表达.结果 二母颗粒高、中、低剂量组连续ig给药14 d,对小鼠Lewis肺癌的抑瘤率分别为30.80％、10.63％、0.其中,二母颗粒高剂量组的瘤重、抑瘤率、VEGF及MVD的表达与模型组比较均有显著差异(P＜0.05).结论 二母颗粒具有抑制小鼠Lewis肺癌的作用,其作用机制可能与抑制瘤内VEGF和MVD的生成有关.     

新的血管生成抑制肽抗肿瘤血管生成的实验研究

目的观察一种新的血管生成抑制肽对肿瘤的抑制作用，并初步探讨作用机制。方法体外药效实验，采用MTT法及血管内皮细胞迁移法，体内采用Lewis肺癌瘤株皮下接种C57BL／6N小鼠，兔眼角膜烧伤模型。观察血管生成情况。结果新的血管生成抑制肽对血管内皮细胞增殖、迁移有明显的抑制作用。体内使Lewis肺癌皮下移植瘤体积明显缩小，并对烧伤诱导的兔眼角膜新生血管也具有明显抑制作用。结论新的血管生成抑制肽能明显抑制Lewis的肿瘤生长，其机制可能与抑制肿瘤血管生成有关。     

威麦宁抑制小鼠Lewis肺癌移植瘤的生长及其血管生成的实验研究

目的:探讨威麦宁对小鼠Lewis肺癌移植瘤生长及其血管生成作用的影响。方法:建立小鼠Lewis肺癌移植瘤模型,用药后取瘤组织称重并计算抑瘤率;瘤组织切片HE染色,光镜下进行形态学观察;免疫组化法检测瘤组织血管内皮细胞CD34的表达情况并计数微血管密度(MVD)。结果:威麦宁浓度在100mg/kg.d、250mg/kg.d时,对小鼠Lewis肺癌的抑制率分别为19.14%和51.56%;对照组癌巢间微血管丰富,用药组瘤组织及其周围微血管均减少;用药组瘤组织内MVD明显低于NS组(P<0.01)。结论:威麦宁能减少肿瘤组织血管生成,从而抑制小鼠移植瘤的体内生长。     

新藤黄酸干预肺腺癌血管生成的细胞学研究

目的 通过对人脐静脉内皮细胞HUVEC和人肺腺癌A549的培养,检测含新藤黄酸(GNA)条件培养基对血管内皮细胞存活率、成管和生长的影响.方法 采用甲基噻唑基四唑(MTT)法和平板克隆实验法研究GNA对HUVEC存活率和克隆形成率的影响;应用薄层胶原建立血管内皮细胞的二维培养模型,观察GN A对于血管内皮细胞成管现象的影响;采用细胞划痕愈合和小室迁移实验考察GNA对HUVEC的迁移能力影响;Westernblot检测血管内皮生长因子(VEGF)和缺氧诱导因子(HIF-1α)蛋白的表达.结果 MTT检测结果显示,HUVEC细胞存活率和克隆形成率随GNA剂量增加而降低.GNA可抑制HUVEC细胞的迁移.还可抑制HUVEC管腔样结构形成.此外,GNA可下调HUVEC中VEGF和HIF-1α蛋白的表达.结论GNA可在体外抑制血管生成,其作用机制可能与抑制肿瘤细胞分泌的HIF-1α和VEGF有关.     

福安泰-03对肿瘤血管生成的抑制作用

目的研究从赤魟组织中分离到的福安泰-03(Fuan-tai-03,FAT-03)对肿瘤血管生成的影响。方法鸡胚绒毛尿囊膜(CAM)法检测FAT-03对人低分化鼻咽癌细胞(CNE-2Z)诱导的CAM血管生成的影响;免疫组化法检查FAT-03对裸小鼠Lewis肺癌组织微血管密度(MVD)和血管生成因子表达的影响。结果FAT-03抑制CNE-2Z细胞诱导的CAM血管生成,当其用量为20、40、80、160μg/胚/d×5时,其抑制率分别为22.3%、34.3%、43.3%、46.2%。FAT-03降低小鼠Lewis肺癌组织的MVD,FAT-03组平均微血管数为9.1±1.5(对照组为18.2±3.2,P<0.05)。免疫组化检测结果显示,FAT-03明显下调小鼠Lewis肺癌组织血管内皮细胞生长因子(VEGF),碱性成纤维细胞因子(bFGF)和血小板衍生生长因子(PDGF)的表达。结论FAT-03有抗肿瘤血管生成作用,其作用与它的下调VEGF、bFGF和PDGF的表达密切相关。     

杂合肽抗肿瘤血管生成的实验研究

目的 观察杂合肽对肿瘤血管生成的抑制作用，并探讨其可能作用机制。方法 体外药效实验，采用MTT法及血管内皮细胞迁移法，体内采用Lewis肺癌瘤株于腹股沟皮下接种C57BL／6N小鼠，观察皮下移植瘤生长情况。结果 杂合肽对血管内皮细胞增殖及迁移(48小时)具有显著的抑制作用，在体内使Lewis肺癌皮下移植瘤体积明显缩小，毛细血管稀少，并表现具有一定的剂量依赖关系。结论 杂合肽能够明显抑制荷瘤鼠肿瘤的生长，其作用机制可能与抑制肿瘤血管内皮细胞生成有关。     

多西他赛抑制小鼠肺癌血管生成分析

目的探讨常用化疗药物多西他赛化疗抑制肿瘤血管生成的作用。方法选择Lewis肺癌细胞荷瘤小鼠50只,随机分成多西他赛不同剂量治疗3组、生理盐水组和对照组,观察4周后5组最终肿瘤体积、质量以及移植瘤血管生成抑制情况。结果多西他赛组小鼠皮下移植瘤质量与体积明显小于其他组,其微血管数（MVD）亦明显低于其他组。结论多西他赛能明显抑制小鼠肺癌血管生成。     

λ-卡拉胶寡糖体外对血管生成的抑制作用

为探讨λ-卡拉胶寡糖体外对血管生成的抑制作用,采用鸡胚尿囊膜模型(CAM)观察λ-卡拉胶寡糖对CAM血管发育的抑制,发现该寡糖可明显抑制CAM微血管生成,在200 μg·egg-1时,抑制率达54.90%.随后采用MTT法测定λ-卡拉胶寡糖对人脐静脉内皮细胞(HUVEC)增殖的影响,发现该寡糖的细胞毒作用在不同的细胞间具有选择性,对HUVEC的抑制最强,高浓度时(1 mg·mL-1)能明显抑制其增殖,但低浓度(＜250 μg·mL-1)对细胞几乎无毒.对寡糖的细胞迁移和侵袭抑制作用进行检测,并测定其对HUVEC表达基质金属蛋白酶-2(MMP-2)的影响.结果表明,λ-卡拉胶寡糖能有效减缓HUVEC的迁移、侵袭能力,并具有抑制HUVEC中MMP-2表达的作用.该研究说明λ-卡拉胶寡糖通过抑制内皮细胞的增殖、细胞侵袭和迁移达到血管生成抑制的作用.     

人参皂苷Rh2 对荷Lewis 肺癌小鼠抗肿瘤作用

目的观察人参皂苷Rh2（ginsenoside Rh2,G-Rh2）对荷Lewis肺癌小鼠抗肿瘤作用并探讨其作用机制。方法建立Lewis肺癌实体瘤模型,观察G-Rh2的抗肿瘤作用、观察肿瘤内血管密度（MVD）,以及免疫组化法观察瘤体血管内皮生长因子（VEGF）的表达。结果 0.3 mg·kg^-1、1.0 mg·kg^-1和3.0 mg·kg^-1G-Rh2组对Lewis肺癌小鼠实体瘤均有抑制作用（P〈0.05）;1.0 mg·kg^-1和3.0 mg·kg^-1 G-Rh2组MVD明显低于对照组（P〈0.05）;免疫组化结果显示,0.3 mg·kg^-1、1.0 mg·kg^-1和3.0 mg·kg^-1 G-Rh2组瘤组织内的VEGF蛋白表达阳性率均低予对照组（P〈0.05）。结论 G-Rh2可抑制Lewis肺癌生长及抑制肿瘤新生血管生成的作用,可能是通过降低VEGF表达来实现的。     

Taspine isolated from Radix et Rhizoma Leonticis inhibits proliferation and migration of endothelial cells as well as chicken chorioallantoic membrane neovascularisation

The aim of the present study was to investigate an anti-angiogenic effect of taspine isolated from Radix et Rhizoma Leonticsi. Taspine was screened for the first time, using cell membrane chromatography (CMC). The anti-angiogeneic activity of taspine was tested by using the chicken chorioallantoic membrane (CAM) neovascularisation model in vivo and the HUVEC proliferation and migration models in vitro, respectively. The results showed that taspine could inhibit CAM angiogenesis significantly within the concentration range of 0.5-2 mug/egg, proliferation and migration of endothelial cells in a dose-dependent manner. The CAM histomorphology results indicated that taspine could inhibit blood vessels sprouts and proliferation of vascular endothelial cell. These findings suggest that taspine is a promising candidate for use as an angiogenesis inhibitor.     

芸香宁碱抑制血管生成作用的研究

目的：研究芸香宁碱对血管生成的抑制作用,并研究其初步的作用机制。方法用人脐静脉内皮细胞（HUVEC）的生长、迁移实验及体内动物模型－鸡胚绒毛尿囊膜法（CAM 法）研究化合物的体内外抗血管生成作用;用酶联免疫吸附法检测芸香宁碱在非凋亡剂量时对肿瘤细胞培养上清液中 VEGF 蛋白分泌量的影响。结果芸香宁碱对血管内皮细胞具有优先抑制作用,对 HUVEC 的半数抑制剂量为（33．2±0．4）μmol·L －1,而对其他肿瘤细胞的 IC50值均大于这一数值;芸香宁碱在体外能够明显抑制内皮细胞的迁移和对细胞外基质黏附作用,并呈剂量依赖性,体内实验显示30μmol·L －1剂量浓度时显著抑制 CAM新生血管形成;芸香宁碱在15μmol·L －1和30μmol·L －1的浓度剂量时显著抑制人肝癌 HepG2细胞培养上清液中VEGF 蛋白的分泌,具有显著性差异。结论芸香宁碱在非凋亡浓度剂量具有抑制新生血管形成作用,其机制与抑制血管内皮细胞增殖以及抑制肿瘤细胞表达 VEGF 有关。     

Chrysin inhibits lipopolysaccharide-induced angiogenesis via down-regulation of VEGF/VEGFR-2(KDR) and IL-6/IL-6R pathways

The relationship between chrysin and inflammation-induced angiogenesis remains unclear. The aim of this study was to evaluate the suppressive effects of chrysin on lipopolysaccharide (LPS)-induced angiogenesis in chicken chorioallantoic membrane (CAM) as well as in human umbilical endothelial cells (HUVEC). The IN VIVO CAM model was applied to evaluate the percentage of new vessels formation, followed by measuring endothelial migration and tube formation in HUVEC cultures. The mechanisms of the suppressive effect of chrysin on LPS-induced angiogenesis, in terms of VEGF, VEGF receptors (VEGFR), interleukin 6 (IL-6) and IL-6 receptor gene expressions, were analyzed by Western blot, ELISA cytokine assay, and quantitative real time PCR. The results showed that chrysin (10(-8) - 10(-5) M) inhibited LPS-induced CAM neovascular density. There was a significant down-regulation of VEGF and VEGFR-2 (KDR) but not VEGFR-1 (Flt-1) gene expression by chrysin in LPS-treated HUVEC cultures. Besides, chrysin concentration-dependently inhibited the auto-regulation loop of IL-6/IL-6R in LPS-treated HUVEC cells. We conclude that chrysin suppresses both IN VITRO and IN VIVO LPS-induced angiogenesis.     

Berberine inhibits HIF-1alpha expression via enhanced proteolysis

We have studied the antiangiogenic property of berberine. We showed that berberine could directly inhibit in vitro human umbilical vein endothelial cell (HUVEC) tube formation and migration. In addition, to determine whether berberine could influence the cross-talk between the gastric adenocarcinoma cell line SC-M1 and vascular endothelial cells, we performed modified confrontation culture experiments and showed that berberine (7.5 microM, 16 h) could inhibit the capacity of hypoxic SC-M1 cells to stimulate HUVEC migration. These results demonstrated berberine's antiangiogenic property and its clinical potential as an inhibitor of tumor angiogenesis. Parallel Western blot analyses revealed that berberine prevented hypoxic SC-M1 cultures from expressing vascular endothelial growth factor (VEGF) and hypoxia-inducible factor (HIF)-1alpha, two key factors in mediating tumor angiogenesis. However, overexpression of HIF-1alpha in SC-M1 cells dramatically reversed the inhibitory effect of berberine on SC-M1-induced in vitro HUVEC migration. These data indicated that HIF-1alpha repression is a critical step in the inhibitory effect of berberine on tumor-induced angiogenesis. Northern blot analyses plus pulse-chase assays revealed that berberine did not down-regulate HIF-1alpha mRNA but destabilized HIF-1alpha protein. We found that berberine-induced HIF-1alpha degradation was blocked by a 26S proteasome inhibitor. Moreover, immunoprecipitation and Western blot analyses showed that berberine increased the lysine-acetylated HIF-1alpha in hypoxic SC-M1 cultures. These data indicated that a proteasomal proteolytic pathway and lysine acetylation were involved in berberine-triggered HIF-1alpha degradation. In conclusion, our data provided molecular evidence to support berberine as a potent antiangiogenic agent in cancer therapy.     

Wogonin inhibits IL-6-induced angiogenesis via down-regulation of VEGF and VEGFR-1, not VEGFR-2

So far, no antiangiogenic activity of wogonin, a flavonoid, on human umbilical vein endothelial cells (HUVECs) has been demonstrated. The aim of this study was to investigate the effects of wogonin on IL-6-induced angiogenesis in HUVEC cultures and chorioallantoic membrane (CAM) neovascularization. The in vivo CAM model was applied to evaluate the percentage of new vessel formations, followed by measurement of endothelial migration and tube formation in HUVEC cultures. The results revealed that wogonin (10(-8) approximately 10(-5)M) concentration-dependently inhibited IL-6-induced angiogenesis. The signaling pathway was through down-regulation of the autocrine loop of VEGF and VEGFR-1, but not of VEGFR-2. Furthermore, the regulating loop of the IL-6 receptor complex was also attenuated via expression of sIL-6Ralpha and gp130, but not of IL-6/IL-6R binding density. We conclude that wogonin is a suppressive agent of the autoregulated loop of VEGF, VEGFR-1 and the IL-6 receptor complex.     

毛萼乙素抑制人正常及肝癌血管生成的作用研究

目的:研究毛萼乙素对人脐静脉内皮细胞(HUVEC)增殖的影响以及对人肝癌血管生成的影响。方法:采用改良MTT法测试6个浓度的毛萼乙素在体外对原代培养的HUVEC增殖的影响;用人肝癌鸡胚尿囊膜血管复制模型,测定毛萼乙素在器官水平对血管生成的抑制作用。结果:毛萼乙素对HUVEC增殖具有显著的抑制作用,半数抑制浓度(IC50)=7.27μg·mL-1;抑制效应呈剂量和时间依赖性。在剂量为25、50、100μg·mL-1时对人肝癌鸡胚尿囊膜血管面积比的抑制率分别为14.15%、48.72%、60.63%,抑制作用中等;其中剂量在50μg·mL-1时与阳性对照反应停相当(P>0.05)。结论:毛萼乙素对人脐静脉内皮细胞的增殖有显著的抑制作用,并能显著抑制人肝癌细胞诱导的鸡胚尿囊膜血管生成。     

Inhibition of tumor-induced angiogenesis and its mechanism by ardipusilloside I purified from Ardisia pusilla

The aim of this study was to evaluate the effects of ardipusilloside I isolated from Ardisia pusilla on tumor angiogenesis and its mechanism of action. The anti-angiogenic effect in vivo was evaluated on xenograft in the athymic mice model and the chicken chorioallantoic membrane (CAM) neovascularization model, the inhibition of growth in vitro was detected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, and the mechanism was demonstrated through detecting microvessel density (MVD), vascular endothelial growth factor (VEGF), VEGF receptor 2 (VEGFR2) and P-VEGFR2 protein expressions, as well as mRNA expressions of VEGF and VEGFR2. The results showed that ardipusilloside I had a good inhibitory effect on A549 xenografted tumor growth, angiogenesis of CAM, and A549 cell growth. Compared to the negative control, MVD protein and mRNA expressions of VEGF and VEGFR were significantly inhibited by ardipusilloside I in a dose-dependent manner. These findings suggested that ardipusilloside I might be a promising candidate as angiogenesis inhibitors.     

槲皮素抑制血管生成作用的实验研究

目的　研究槲皮素 (Quercetin)对血管生成和培养的人脐静脉内皮细胞 (HUVEC)的影响。方法　采用生长因子 (血管内皮细胞生长因子VEGF、碱性成纤维细胞生长因子bFGF)诱导的鸡胚绒毛尿囊膜 (CAM)血管增生模型观察槲皮素对血管生成的影响 ;利用培养的HUVEC ,用MTT法观察槲皮素抑制内皮细胞增殖的作用 ;流式细胞仪观察槲皮素对HUVEC细胞周期的影响。结果　槲皮素 (0 1、0 0 5和 0 0 2 5mmol·L-1)能明显抑制VEGF诱导的CAM小血管生成 ;槲皮素 (0 1和 0 0 5mmol·L-1)能明显抑制bFGF诱导的CAM小血管生成 ;槲皮素 (2 4 0、12 0 μmol·L-1和 6 0 μmol·L-1)对内皮细胞增殖有抑制作用 ,抑制率分别为 6 7 0 %、5 8 1%和39 7% ;槲皮素 (2 4 0、12 0 μmol·L-1)能显著导致HUVEC的S、G2 期阻滞。结论　槲皮素能抑制VEGF和bFGF诱导的血管生成 ,且对内皮细胞增殖具有抑制作用。     

Saikosaponins-b suppresses tumor growth and angiogenesis of hepatocellular carcinoma by regulating VEGF/ERK/HIF-1α signal pathway

OBJECTIVE Angiogenesis therapy has attracted interest as a potential treatment for hepatocellular carcinoma(HCC).In this study,we investigated the anti-proliferative activities and antiangiogenesis effects of saikosaponins(SS)-b on hepatocellular carcinoma(HCC)and its regulation on VEGF/ERK/HIF-1 αsignal pathway.METHODS H22 hepatoma-bearing mice model and HepG-2 cells were used to study the anti-tumor and anti-angiogenesis effects of SS-b in vivo and in vitro.Pathological change of tumor tissue was observed by HE staining,the microvascular changes were detected by immunohistochemical method.The effects of SS-b on angiogenesis were examined by using the chick embryo chorioallantoic membrane(CAM)model.The effects of SS-b on proliferation,migration and invasion were investigated by MTT assay,scratch wound healing assay and transwell assay inhuman umbilical vein endothelial cell(HUVEC)and HepG2 cells in vitro.Vascular endothelial growth factor(VEGF),matrix metalloproteinase-2/9(MMP-2/9),hypoxia-inducible factor-1α(HIF-1α)expression and the phosphorylation of extracellular regulated kinase(ERK)were analyzed using RT-PCR and Westernblot.RESULTS SS-b effectively inhibited the tumor growth of H22 mice in vivo.The inhibitory rate of tumor was 49.1%,50.7%,66.1%in SS-b 5,10 and 20 mg·kg-1group respectively.HE staining results showed that SS-b induced tumor necrosis and nuclear dissolution in H22 mice.Moreover,SS-b also reduced the number of microvessels of tumor tissue in H22 mice significantly and suppressed the angiogenesis of CAM induced by b-FGF.SS-b had an obvious inhibitory effect on cell proliferation,migration and invasion of HUVEC cells and HepG-2 cells.These effects were associated with downregulation of the expression of MMP2/9 and suppression of VEGF/ERK/HIF-1αsignaling in H22 mice and Hep-G2 cells.CONCLUSION Our findings showed that SS-b exerts anti-tumor effects by inhibiting tumor angiogenesis via regulating VEGF/ERK/HIF-1α signal pathway in vivo and in vitro.     

松萝酸抑制小鼠H22肿瘤生长及作用机制研究

摘 要 目的： 探讨松萝酸对小鼠H22肿瘤生长的影响及其作用机制。方法: 昆明种小鼠前腋下皮下接种H22细胞,第2天腹腔注射松萝酸,每3 d测量1次肿瘤体积。用免疫组化法检测微血管密度（MVD）;ELISA 法检测小鼠血清和人脐静脉血管内皮细胞（HUVEC）细胞培养基上清中血管内皮生长因子（VEGF）和碱性成纤维细胞生长因子（bFGF）水平;MTT法检测松萝酸对人脐静脉血管内皮细胞（HUVEC）体外增殖的影响。结果: 松萝酸组小鼠H22肿瘤体积、重量增长明显较模型组缓慢(P<0.05),同时松萝酸组的肿瘤微血管密度显著减少(P<0.05),松萝酸对HUVEC体外增殖有显著抑制作用。松萝酸可显著降低HUVEC细胞和小鼠血清VEGF和bFGF水平(P<0.01)。结论:松萝酸可显著抑制小鼠H22肿瘤生长及血管生成,且能抑制VEGF和bFGF表达和HUVEC体外增殖。松萝酸对VEGF和bFGF分泌的抑制作用可能是其抗H22肿瘤及抗血管生成的一个重要机制。