鸡胚绒毛尿囊膜血管生成模型的应用

本文综述了鸡胚绒毛尿囊膜血管生成模型在药物促血管生成和抑制血管生成作用研究中的应用，对药物载体的选择及药物置入时间的选择等进行了介绍。     

硫酸软骨素对鸡胚绒毛尿囊膜的血管生成抑制作用

目的观察硫酸软骨素的血管生成效应。方法将 15g/L硫酸软骨素溶液 4 μl加到孵化 4d的鸡胚绒毛尿囊膜血管网上 ,2 4h后观察对血管生长的影响。同时设有同浓度的氢化可的松组和肝素组及三蒸水组作为对照组。结果硫酸软骨素使鸡胚绒毛尿囊膜血管发生大面积褪色 ,血管密度减低 ;氢化可的松组血管结构模糊 ,分枝发生断裂 ,颜色变浅 ,血管密度减少 ;肝素组血管生长旺盛 ,血管密度增加 ;正常对照组血管网清晰 ,呈叶脉状 ,放射状生长。结论硫酸软骨素有显著的抗血管生成活性。     

珠母贝糖胺聚糖对鸡胚绒毛尿囊膜血管生成的影响

目的探讨珠母贝糖胺聚糖的抗肿瘤作用机制。方法采用鸡胚绒毛尿囊膜(CAM)模型观察珠母贝糖胺聚糖单用及与5-氟尿嘧啶(5-FU)合用对CAM血管生成的影响。结果珠母贝糖胺聚糖CPG、PGⅠ、PGⅡ在实验所选剂量(0·5、5、50、500mg·L-1)下,无明显的直接抑制CAM血管生成作用;但CPG、PGⅠ、PGⅡ在实验所选剂量(5、50mg·L-1)下与5-FU(2·5mg·L-1)合用时,均能增敏5-FU的抑制CAM血管生成作用(P<0·01),并呈一定的剂量效应关系,其中PGⅡ增敏效果较好。结论珠母贝糖胺聚糖通过抑制肿瘤血管生成而对5-FU的抗肿瘤作用起增敏效果。     

暖宫孕子胶囊对鸡胚绒毛尿囊膜血管生成的影响

目的观察暖宫孕子胶囊对鸡胚绒毛尿囊膜(CAM)血管生成的影响。方法取7日鸡胚40枚,建立CAM模型,随机分为试验组和空白组,每组20枚;试验组加暖宫孕子胶囊药液于无血管区的载体上,空白组加生理盐水,每日加药1次;3 d后检测CAM载体周围一级、二级的血管数目。结果试验组一级、二级血管数目均多于空白组,两组比较有显著性差异(P<0.05)。结论暖宫孕子胶囊能促进鸡胚绒毛尿囊膜(CAM)的血管生成。     

四种抗肝癌中药对鸡胚绒毛尿囊膜血管生成的影响

目的通过测定四种中草药对鸡胚绒毛尿囊膜（CAM）模型血管生成的作用,并与阳性药物索拉菲尼对比,探究药物作用与血管生成的关系,探究药物可能存在的靶向治疗作用。方法醇提取四种广西特色中草药．建立鸡胚绒毛尿囊膜模型测定不同药物醇提物与极性组段抗血管生成的抑制作用。结果索拉菲尼、破骨风、土半夏、血党均有不同程度的抑制鸡胚尿囊血管生长作用,与空白对照组相比,索拉菲尼与破骨风对二、三级血管均有明显抑制作用（P〈0．01）;土半夏、血党对三级血管有抑制作用（P〈0．01）,其高剂量组对二级血管也有明显抑制作用（P〈0．01）。结论索拉菲尼、破骨风、土半夏、血党均可不同程度抑制血管的生长,可能是通过与索拉菲尼相同的作用机制实现;天胡荽对血管生长的抑制作用较弱。     

鸡胚绒毛尿囊膜技术应用体会

血管新生及其治疗性应用在基础研究方面取得了巨大的进展,国外已开始早期的临床研究[1].治疗性血管生成(therapeutic angiogenesis)可以导致毛细血管的芽生(angiogenesis,血管新生)和侧支血管的发展(arteriogenesis,动脉生成),为冠心病提供了一种新的治疗选择.     

莪术油对鸡胚绒毛尿囊膜新生血管抑制作用研究

目的:探讨莪术油对鸡胚绒毛尿囊膜(CAM)新生血管形成的抑制作用.方法:制备鸡胚绒毛尿囊膜模型(CAM).水蒸馏法提取莪术油,稀释成不同浓度.平行设置NS对照(阴性)和沙利度胺片对照(阳性),每组分别8枚鸡胚.接种各药物后,37℃培养箱中继续孵育,每12小时观察一次,孵化5天.通过肉眼及体视显微镜观察,数码相机拍照留存,图像处理后进行血管面积值/面积比值测定,并作统计学处理.结果:莪术油鸡胚尿囊膜血管面积值/面积比值较生理盐水对照组显著增高,且有一定的量效关系趋势.结论:莪术油可剂量依赖性地抑制新生血管的作用,且效果明显.     

黄芪注射液促进鸡胚绒毛尿囊膜血管生成的实验研究

目的:探讨黄芪注射液对鸡胚绒毛尿囊膜(chorioallantoicmembrane,CAM)血管生成的影响。方法:种蛋随机分为5组,孵育7d后,将高、中、低剂量黄芪注射液穴Astragalusmembranaceusinjec鄄tion,AMI雪及生理盐水(NS)、成纤维细胞生长因子(EGF)对照品分别加于CAM表面的载体上,作用后制备CAM标本,解剖显微镜下计数新生血管数目。结果:黄芪注射液具有明显的促进鸡胚绒毛尿囊膜血管生成的作用,与NS阴性对照组相比有显著性差异(P﹤0.05),但药效低于EGF阳性对照组(P﹤0.05)。结论:黄芪注射液可促进鸡胚绒毛尿囊膜血管生成。     

当归注射液促进鸡胚绒毛尿囊膜血管生成的实验研究

目的:探讨当归注射液对鸡胚绒毛尿囊膜(chorioallantoicmembrane,CAM)血管生成的影响。方法:种蛋随机分为5组,孵育7d后,将高、中、低剂量当归注射液(Angelicasinensisinjection,ASI)及生理盐水(NS)、成纤维细胞生长因子(EGF)对照品分别加于CAM表面的载体上,作用后制备CAM标本,解剖显微镜下计数新生血管数目。结果:当归注射液具有明显的促进鸡胚绒毛尿囊膜血管生成的作用,与NS对照组相比有显著性差异(P<0.05),但药效低于EGF对照组(P<0.05)。结论:当归注射液可促进鸡胚绒毛尿囊膜血管生成。     

5种他汀类药物对鸡胚绒毛尿囊膜血管新生的影响

目的:观察5种他汀类药物对鸡胚绒毛尿囊膜(CAM)血管新生的影响,探讨他汀类药物是否具有独立于血脂调节作用以外的促进血管新生作用。方法:制备CAM模型,将不同浓度他汀类药物、重组牛碱性成纤维细胞生长因子和生理盐水分别通过载体加到CAM上,观察CAM特异性血管生长情况及血管数目变化。结果:不同浓度辛伐他汀(0.02,0.1,0.5,2.5,12.5μmol.L-1)中,当辛伐他汀浓度为0.1μmol.L-1时,CAM血管生成数量明显高于生理盐水对照组(P<0.05),与阳性对照组比较无统计学意义(P>0.05);而不同他汀类药物,即瑞舒伐他汀组(0.025μmol.L-1)、阿托伐他汀组(0.05μmol.L-1)、普伐他汀组(0.2μmol.L-1)、氟伐他汀组(0.4μmol.L-1),CAM血管生成数均明显高于生理盐水组(P<0.05),与阳性对照组比较无统计学意义(P>0.05)。结论:常用5种他汀类药物均有促进CAM血管新生作用;不同浓度辛伐他汀对CAM的作用不同。提示他汀类药物促进血管新生的作用可能与药物剂量有关。     

The effects of amphotericin B on angiogenesis in chick chorioallantoic membrane

Objective: Amphotericin B (AmB) is widely used as a mainstay in the treatment of sight-threatening fungal endophthalmitis. From the time that itraconazole was discovered to have a previously unknown anti-angiogenic activity, we have suspected that AmB may have possible effects on ocular angiogenesis. The purpose of this study was to evaluate the in vivo anti-angiogenic effect of AmB in the chick chorioallantoic membrane (CAM) model.

Materials and methods: Atak-S type fertilized eggs obtained from the Poultry Institution were used. The eggs were kept under 37?°C at 85–90% relative humidity throughout the experiment. Amphotericin B was prepared in two different concentrations (AmB 125?μg/1?mL and 0.125?μg/1?mL). The CAMs treated with sterile distilled water was specified as controls. About 0.1?mL of each containing 12.5 and 0.0125?µg of AmB, respectively, were dropped to CAM surface. Thirteen eggs were used for each group. The results were evaluated at the 48th hour of the administration of the drugs and recorded by digital camera.

Results: A reduction of angiogenesis in CAM area which treated with 125?μg/1?mL of AmB was appreciable macroscopically. The affected areas showed impaired radial arrangement of small vessels with the presence of avascular zone at periphery. The dose of 0.125?μg/1?mL AmB did not show any visible anti-angiogenic effect. Numerous blood vessels with a radially arranged pattern developed toward the periphery after 48?h of treatment. In the CAMs that treated with distilled water, physiological angiogenesis was observed in allantoic vessels. Vessel formation seems to be similar in CAMs treated with 0.125?μg/1?mL AmB with the presence of visibly non-malformed alive embryos.

Conclusions: The present study gives the impression that AmB has the capacity to serve as an anti-angiogenic treatment. As it is a preliminary CAM study only, further studies on both animals and humans are required.     

丹酚酸B对鸡胚绒毛尿囊膜血管新生作用的影响

目的探讨丹酚酸B对鸡胚绒毛尿囊膜血管新生的影响。方法将7日龄鸡胚制备鸡胚绒毛尿囊膜(chick embryo chorioallantoic membrane,CAM)模型,随机分为6个组,分别为PBS(phosphatebuffer solution)对照组,丹酚酸B 4个剂量组(150、50、16.67、5.56 mg.L-1)及VEGF(vascular endo-thelial growth factor)组,培养3 d后时采集数据进行评价。结果丹酚酸B 150、50 mg.L-1组血管新生面积明显高于PBS(phosphate buffer solution)对照组(P<0.01,P<0.05)。结论丹酚酸B可明显促进鸡胚绒毛尿囊膜血管新生。     

Mainstream and sidestream cigarette smoke inhibit growth and angiogenesis in the day 5 chick chorioallantoic membrane.

The purpose of this study was to test the hypothesis that components in mainstream (MS) and sidestream (SS) cigarette smoke inhibit growth and angiogenesis using the chick chorioallantoic membrane (CAM). Varying doses of whole or gas-phase MS and SS smoke solutions were placed on day 5 CAMs, and their effects on angiogenesis were evaluated on day 6. All parameters evaluated (CAM area, major blood vessel area, major blood vessel diameter, blood vessel pattern formation, and capillary plexus formation) were inhibited to different degrees in a dose-dependent manner by both MS and SS smoke treatment. Inhibition of growth and vessel development was correlated with inhibition of cell proliferation. Inhibition of capillary plexus formation was caused by failure of mesodermal blood vessels to migrate to the ectoderm. SS smoke solution was more inhibitory than MS smoke solution in all assays, except for capillary plexus formation. In all assays, the toxicants in SS smoke partitioned mainly with the gas phase, whereas those in MS smoke were deduced to be mainly in the particulate phase in the proliferation-dependent assays (CAM area, blood vessel area, blood vessel diameter) and in both the gas and particulate phase in the pattern formation and plexus formation assays. Some of the inhibitory doses of MS and SS smoke solutions had nicotine concentrations within the range found in human smokers. Taken together, these data demonstrate that exposure to complex mixtures of chemicals in MS and SS cigarette smoke adversely affect growth, vessel development, vessel migration, and cell proliferation.     

Investigating the effect of excess caffeine exposure on placental angiogenesis using chicken ’functional‘ placental blood vessel network

It is now known that over‐consumption of caffeine by pregnant mothers could have detrimental effects on normal fetal development. However, it remains obscure how caffeine's harmful effect impacts directly or indirectly on the developing embryo/fetus through damaging placenta development. In this study, we demonstrated the morphological similarities between the yolk sac and chorioallantoic membranes (CAM) of chick embryos and the villi of the mammalian placenta. Using the chick yolk sac and the CAM as a model, we found that 5–15 µmol per egg of caffeine exposure inhibited angiogenesis. Under the same condition, cell proliferation in extraembryonic mesoderm was reduced while apoptosis was enhanced. Semi‐quantitative RT‐PCR analysis revealed that caffeine treatment down‐regulated VEGF, VEGFR2, PIGF, IGF2 and NRP1 expression, but up‐regulated Ang1 and Ang2 expression. We performed in situ hybridization to show VE‐cadherin expression and as to demonstrate the blood vessels in the CAM and yolk sac membranes. This distribution of the VE‐cadherin⁺ blood vessels was determined to be reduced after caffeine treatment. Furthermore, MDA activity was induced after caffeine exposure, but GSH‐PX activity was inhibited after caffeine exposure; SOD activity was unchanged as compared with the control. In summary, our results suggest that caffeine exposure could negatively impact on angiogenesis in the chick yolk sac and CAM by targeting angiogenesis‐related genes. Some of these genes are also involved in regulating excess ROS generation. The results implied that the negative impact of caffeine on fetal development was partly attributed to impaired placental angiogenesis. Copyright © 2015 John Wiley & Sons, Ltd.     

An investigation of the chick chorioallantoic membrane as an alternative model to various biological tissues for permeation studies

Objectives The chick chorioallantoic membrane (CAM) was explored as a biological membrane for use in the study of drug permeation with a Franz diffusion cell. Methods The CAM was removed from fertilized chicken eggs of embryo age 9–18 days. The permeation profiles of nicotine through the fresh CAM were first obtained with a Franz diffusion cell. The permeation profiles of nicotine through frozen CAM, snake skin, pig skin, pig retina and pig buccal mucosa were also determined and compared with those of the fresh CAM. Key findings The permeability coefficient of the CAM varied with its age. The CAM at embryo age 13 was the most robust, showing the lowest standard error in permeability. It was thus chosen for comparative studies with snake skin, pig skin, retina and buccal mucosa. The CAM was found to be most similar to the buccal mucosa in terms of permeation profile and permeability coefficient values. Frozen CAM was also found to have a higher permeability coefficient than fresh CAM. The enhanced permeability was attributed to freezing, which affected the integrity of the CAM structure. Conclusions From the findings, CAM shows potential as an alternative to the pig buccal mucosa as an in‐vitro buccal model. The robustness of the CAM for drug permeation studies is affected by its age.     

Hen's egg chorioallantoic membrane test for irritation potential

The increasingly large number of chemicals introduced onto the market and into the environment has necessitated the monitoring of environmental materials and specimen banking, as well as the development of rapid and reliable methods for the evaluation of toxicity. The Hen's Egg Test, or Hühner-Embryonen-Test (HET) is a rapid, sensitive and inexpensive toxicity test and can give information on embryotoxicity, teratogenicity, systemic and immunopathological effects, metabolic pathways and now, in developed form, on mucous-membrane irritation potencies of chemical substances. Testing with incubated hen's eggs is a borderline case between in vivo and in vitro systems and does not conflict with ethical and legal obligations especially animal protection laws. In the special field of mucous-membrane irritation testing, a specific score and classification scheme was developed for the HET, which allows risk assessments analogous to the Draize scheme. There is a good correlation between the results for HET tests on a variety of pyrithiones, phenols and isothiazolinones, and the corresponding data based on Draize tests. HET chorioallantoic membrane testing should and could not entirely replace current irritation tests in mammals, but it can diminish the number of investigations with mammals, as well as limit or eliminate pain and injury during animal experiments and allow regulators to set priority and toxicity categories.