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The Hancornia speciosa latex has shown angiogenic activity. Angiogenesis plays a major role in wound healing, and materials that stimulate this process could be used to develop drugs. This study aimed to explain the role of proteins in the H. speciosa serum fraction latex in angiogenesis. Hence, this material was treated with proteinase K and the proteins were inactivated. After protein inactivation, angiogenic activity was assessed with the chick chorioallantoic membrane assay. The result showed that the proteins in the serum fraction are responsible for angiogenic activity. Then, the total protein content in the serum fraction and its enzymatic activity were investigated. The low protein content observed in the H. speciosa serum fraction latex suggests that this biomaterial could be used to develop new drugs with a hypoallergenic response. Despite the low protein content, there was a significant enzymatic activity of at least three enzymes in the serum fraction latex: ß-1,3 glucanase, ß-glucosidase, and proteases. These enzymes seem to influence the healing process, assisting debridement, extracellular matrix remodeling, and collagen deposition, and decreasing the chances of contamination by microorganisms. In conclusion, the enzymes in the H. speciosa serum latex are associated with the angiogenic activity of this biomaterial and may be used to assist the wound healing process.
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Wound Healing , Endopeptidase K , Apocynaceae , Enzymes , LatexABSTRACT
Background: Angiogenesis is important for the typical physiological activities such as cure from injury, menstrual cycle and embryo growth. It is also plays a crucial role in several pathological conditions in cancer. Antiangiogenesis, e.g., inhibition of blood vessel growth, is being investigated as a way to prevent the growth of tumors and other angiogenesis-dependent diseases. The chick embryo chorioallantoic membrane (CAM) is commonly used as an experimental in vivo assay to study both angiogenesis and antiangiogenesis in response to tissues, cells or soluble factors. Given the high occurrence of cancer worldwide and the major source of the discovery of new lead molecules are medicinal plants. The objective of the present research was to study the antiangiogenic property of “aqueous extract of Nigella sativa seeds” using chick chorioallantoic membrane (CAM) assayMethods: The chick chorioallantoic membrane (CAM) assay for screening the effect of Nigella sativa on anti-angiogenesis was performed according to the method given by Ribatti and co-workers.Results: The results of present study significantly increased the antiangiogenic effect on CAM by decreasing the proliferation of capillary networks in a dose (50 to 300 µg/egg) dependent manner which is probably related to the inhibition of neovascularization.Conclusions: It is concluded that aqueous extract of N. sativa seeds possesses significant antiangiogenic activity, and this is a possible rationale for its folkloric use as an anticancer agent.
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ABSTRACT The phytochemical study of Galium tunetanum Lam., Rubiaceae, leaves led to the isolation of 13 compounds from the chloroform-methanol and the methanol extracts, including six iridoid glycosides, one non-glycoside iridoid, two p-coumaroyl iridoid glycosides, two phenolic acids, and two flavonoid glycosides. The structural determination of the isolated compounds was performed by mono- and bidimensional NMR spectroscopic data, as well as ESI-MS experiments. All compounds were isolated from this species for the first time. The anti-angiogenic effects of the isolated iridoids were also reported on new blood vessels formation using the chick embryo chorioallantoic membrane as in vivo model. Results showed that among the isolated iridoids tested at the dose of 2 µg/egg, asperuloside (1), geniposidic acid (2), and iridoid V1 (3) reduced microvessel formation of the chorioallantoic membrane on morphological observations using a stereomicroscope. The anti-angiogenic effects of the active compounds, expressed as percentages of inhibition versus control, were 67% (1), 59% (2), and 54% (3), respectively. In addition, the active compounds were able to inhibit angiogenesis in the chorioallantoic membrane assay, in a dose-dependent manner (0.5-2 µg/egg) as compared to the standard retinoic acid.
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Objective To compare the growth of three different cancer cell lines on chick chorioallantoic membrane (CAM), to select the best transplanted cancer cell line for establishing a transplanted tumor model and to observe the biological characteristics.Methods The human lung cancer cell line A549, human tongue cancer cell line TCA8113 and human liver cancer cell line QGY7703 were respectively inoculated into CAM at the 7th day of age.The chick embryo survival rate, tumor survival rate, tumor formation rate and induced angiogenesis were detected and the growth characteristics of the transplanted tumor model were observed.Results Compared with the groups inoculated with A549 cells and QGY7703 cells, the tumor formation rate of TCA8113 cells was the highest (P < 0.05), to be the best cancer cell line for transplanted tumor.The optimal inoculated number of cells was 8.0×106/chick embryo, the optimal growth period of the tumor was 4~8 d, and the best experiment time was 7 d after inoculation.Conclusion The TCA-CAM transplanted tumor model of tongue squamous cell cancer is successfully established for further study of the biological characteristics and mechanisms of tumor growth, angiogenesis, invasion and metastasis, and provide a good experimental animal model for anti-tumor drug screening.
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Objective Angiogenesis is the development of new blood vessels. The ion channels on endothelium play a vital action in cell proliferation and so in the related angiogenesis. We aimed to investigate the anti-angiogenic effects of Mefloquine (Cl− channel blocker) and 4-Aminopyridine (K+ channel blocker). Methods The anti-angiogenic activities of Mefloquine and 4-Aminopyridine (4-AP) were investigated by in-vivo (sponge implantation method), in-vitro (aortic ring assay) and in-ovo (CAM, Chick Chorioallantoic membrane) methods. The standard antiangiogenic drug used was Bevacizumab. Results In the CAM assay, both the ion channel blockers exhibited noticeable antiangiogenic activity at the concentrations of 10
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Abstract Angiogenesis plays a key role in tumor growth, invasion and metastasis of cancer diseases and therefore, the inhibition of angiogenesis can provide an important therapeutic approach in cancer diseases. This study was designed to compare the anti-angiogenic activities of the ethanolic extract of Artemisia sieberi Besser, Asteraceae, and its active substance, artemisinin in both in vitro and in vivo models. To compare cytotoxicity level of ethanolic extract of A. sieberi with artemisinin, different concentrations (1–100 µg/ml) were tested using MTT assay on human umbilical vein endothelial cells. The anti-angiogenic properties of serial concentrations of ethanolic extract of A. sieberi and artemisinin were examined on human umbilical vein endothelial cells using a three-dimensional angiogenesis assay (in vitro model) and in the chick chorioallantoic membrane assay as in vivo model. The effects of ethanolic extract of A. sieberi and artemisinin were also tested on the expression of VEGFR-1, VEGFR-2 and CD34 genes using real-time PCR. Ethanolic extract of A. sieberi and artemisinin significantly (p < 0.001) inhibited the angiogenesis in the human umbilical vein endothelial cells culture whilst the ethanolic extract of A. sieberi showed higher effect in a concentration-dependent fashion (p < 0.001). The chick chorioallantoic membrane angiogenesis was also completely inhibited by ethanolic extract of A. sieberi at concentration of 33 ng/100 µl/egg. The gene expression analysis showed that the ethanolic extract of A. sieberi and artemisinin reduced the transcription of VEGFR-1, VEGFR-2 and CD34 genes in a concentration-dependent manner. This study demonstrated that the ethanolic extract of A. sieberi is strongly able to inhibit the angiogenesis in human umbilical vein endothelial cells and chick chorioallantoic membrane models compared to the artemisinin.
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Aim To examine the inhibitory effect of re-combinant cardiac troponin fusion protein composed of subunit I and artificial peptide which was called CIS on tumor growth. Methods The CIS ’ s effect on the growth of human umbilical vein endothelial cells ( HU-VEC) was examined using MTT assay in vitro. Chick chorioallantoic membrane model was applied to study the alteration of angiogenesis treated with purified re-combinant CIS protein. The effect of tumor growth trea-ted with CIS was observed using several in vivo mice xenograft models. Results There was a statistically significant reduction in HUVEC cell proliferative rate when the cells were treated with purified CIS fusion protein, which was also shown in a dose-dependent manner. A decreased amount of new blood vessel for-mation ( angiogenesis) on chick embryo chorioallantoic membranes was observed in recombinant CIS protein treated group compared to the untreated control group. A significant inhibition of tumor growth rate was a-chieved in CIS treated mice compared to CIS untreated control mice in 6 different mouse xenograft models. Conclusions The fusion protein CIS shows the inhibi-tory effect on the tumor growth in our in vivo mouse models, and such inhibition could be mediated by the mechanism of CIS’ s effect on the decrease of HUVEC cell proliferation and further the reduction of angiogen-esis in tumor tissues. This work could provide the foundation for the in-depth investigations on the phar-maceutical application of CIS targeting anti-tumor ther-apy.
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Objective To establish the xenotransplanted tumor model of Craniopharyngioma in chick chorioallan?toic membrane (CAM) and detect the angiogenesis ability, microvessel density (MVD) and cell proliferation of the xeno?graft. Method Craniopharyngioma tissues from surgical craniopharyngioma patients were transplanted on the CAM. An?giogenesis assay was performed and the MVD and PCNA were evaluated using immunohistochemistry following the trans?plantation. Results The tumor formation rate of adamantinomatous craniopharyngioma (ACP) and squamous papillary cra?niopharyngioma (SPCP) was 47.14% and 43.33%, respectively. There was no significant difference in tumor formation rate between ACP and SPCP(χ2=0.123,P=0.726). The CAM angiogenesis, MVD and expression of PCNA were higher in ACP than in SPCP. The expression of PCNA was positively correlated with MVD (Pearson r=0.639,P<0.001) and CAM assay score (Spearman r=0.490,P=0.001 ) in CP. Conclusion The model of human craniopharyngioma can be es?tablished in the CAM. The angiogenesis of the xenograft in the CAM can be evaluated and the craniopharyngioma xeno?graft of CAM possesses a new blood circulation and cell proliferation ability.
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Objective To evaluate the antitumor effect of corosolic acid and its impact on CAM and YSM angiogenesis.Methods The effects of CRA on A549 proliferation was studied by MTT method in vitro.Tumor-bruden nude mice model were established by injecting A549 lung cancer cell marked with bioluminescent to nude mice,and the growth of tumor in mice were detected by IVIS small animal in vivo imaging system.Experiment of chicken embryos eggs are used to observe the role of drugs on CAMand YSMblood vessels. Results The value of IC50 of CRA for A549 in vitro was 26.8μg/mL.A tumor-burdened animal experiment results showed that the CRA to A549 solid tumor has a certain therapeutic effect.CAM and YSM blood vessels of chicken embryos eggs treated with CRA were decreased significantly than negative control group.Conclusion CRA has certain therapeutic effect for A549,which may be related to the inhibition of angiogenesis in tumor tissues.
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OBJECTIVE: To explore the anti-tumor activities of a new compound 3β,12β,20(S)-trihydroxy dammarane-3-O-β-D-glucopyranosyl (1-2)-β-D-glucopyranoside (HRG), which is a substance of ginsenoside structure modification, in vivo. METHODS: Liver cancer H22 tumor model on the chick chorioallantoic membrane (CAM) was established to observe the effects of HRG on tumor growth, the number of induced vessels and the growth inhibition rate. The implanted tumors were dyed by hematoxylin-eosin (HE), and the morphological properties were studied with light microscope. Immunohistochemistry (SP method) was used to detect the expressions of the implanted tumor's MVD and VEGF. RESULTS: HRG inhibited the tumor growth at 25, 50 and 100 μg · mL-1. Compared with the model control group, the inhibitory rates of the tumor growth were 27.73%, 50.02%, and 64.21%, respectively. HRG significantly reduced the number of tumor-induced vessels. At the same time, there existed different degree necrosis among the treatment groups, and the degree of necrosis had an inverse relationship with the number of tumor-induced vessels. In addition, HRG reduced the MVD of the implanted tumors, and decreased the expression of VEGF. CONCLUSION: HRG has good anti-tumor activities in vivo, and can significantly inhibit the growth of liver cancer H22-CAM tumor and the induction of angiogenesis. The mechanisms may be associated with lower tumor MVD and VEGF expression.
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Synadenium umbellatum Pax, popularly known as "cola-nota", is a medicinal plant that grows in tropical regions. Latex of this plant is used to treat various diseases such as diabetes mellitus, Hansen´s disease, tripanosomiases, leukemia and several malignant tumors. In the present study, the angiogenic activity of S. umbellatum latex was evaluated using the chick embryo chorioallantoic membrane (CAM) assay. Results showed significant increase of the vascular net (p < 0.05) compared to the negative control (H2O). The histological analysis was in accordance with the results obtained. In conclusion, our data indicate that S. umbellatum latex, under the conditions of this research, presented angiogenic effect.
Synadenium umbellatum Pax, popularmente conhecida como "cola-nota", é uma planta medicinal que cresce em regiões tropicais. O látex desta planta tem sido utilizado no tratamento de várias doenças, como diabetes mellitus, hanseníase, tripanossomíases, leucemia e vários tumores malignos. No presente estudo, a atividade angiogênica do látex de S. umbellatum foi avaliada pelo ensaio da membrana corio-alantóide (MCA) de ovo embrionado de galinha. Os resultados mostraram aumento significativo da rede vascular (p < 0.05) em relação ao controle negativo (H2O). A análise histológica está em concordância com os resultados obtidos. Em conclusão, os dados indicaram que, nas condições experimentais deste estudo, o látex de S. umbellatum exibiu efeito angiogênico.
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Animals , Chick Embryo , Chorioallantoic Membrane/drug effects , Euphorbiaceae/chemistry , Latex/pharmacology , Neovascularization, Physiologic/drug effects , Neovascularization, Physiologic/physiologyABSTRACT
Objective To investigate the therapeutic mechanism of the recipe of effective components from Yangxin Tongmai Formula (YTF) for myocardial ischemia by observing its effect on angiogenesis. Methods The breeding egges were divided into 6 groups: YT Ⅱ group treated with serum containing the recipe of effective components from YTF, YT Ⅰ group treated with serum containing YTF, SBW group treated with serum containing Shexiang Baoxin Pill, bFGF group treated with serum containing recombinant bovine basic fibroblast growth factor, KX group treated with blank control serum, and blank control group, 25 eggs in each group. The survival rate of chicken embryo, growth shape and vessel count of CAM model in each group were observed after 0, 24, 48, 72 and 96 hours. Results After treatment for 96 hours, the survival rate in b-FGF group and YT II group was higher than that in the other groups. The vasculature growth was rapidly in the medication group after culturing for 72 hours, and more apparent after 96 hours, especially in the b-FGF group and YT-Ⅱ group. The vessel count in b-FGF group, YT-Ⅱ group, YT-Ⅰ group and SBW group was in-creased as compared with that in KX group and blank control group (P < 0. 05), and ranged in the order as follows: b-FGF group > YT-Ⅱ group > YT-Ⅰ group > SBW group > KX group > blank control group. Conclusion YT-Ⅱ has the sim-ilar effect on improving angiogenesis as YT-Ⅰ, SBW and b-FGF, and its effect is better than that of YT-Ⅰ and SBW.
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Objective:To investigate the antiangiogenesis ability of 5 FU. Methods:Human umbilical vein endothelial cells were cultured primarily in vitro and influence of 5 FU on HUVEC proliferation was evaluated by MTT method. Chick chorilallantoic membranes(CAM) model was used to check whether the neovascularization of CAM could be suppressed in vivo . Results:5 FU both inhibited proliferation of HUVEC in vitro and suppressed angiogenesis of CAM in vivo at none cytotoxic doses. Conclusion: 5 FU demonstrated antiangiogenesis ability without obvious cytotoxicity.
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Objective To express the recombinant human canstatin protein, and to examine its biological activity. Methods Canstatin cDNA was cut off from the plasmid pUCm-T/canstatin with restriction enzymes BamHⅠ and Hind Ⅲ. The cDNA fragment was then ligated into the correspondence sites of plasmid pET-22b(+) by T4 DNA ligation enzyme and transformed into E.coli BL21 which was induced to express proteins with isopropyl-1-thio-b-dgalactopyranoside (IPTG). The expressed proteins were analyzed by SDS-PAGE and purified through Ni-NTA column affinity chromatography. Chick chorioallantoic membrane (CAM) assay was performed to determine the activity of the recombinant protein. Results Canstatin cDNA from pUCm-T showed one clear objective DNA band with electrophoresis. Seven of positive colonies were selected and identified by restriction enzyme analysis with BamH Ⅰ and Hind Ⅲ. Electrophoresis revealed that all selected colonies had two specific bands,one near the location of primary plamid,the other near that of objective gene fragment. After IPTG induction, there was a new protein band about 24 000 on SDS-PAGE.The induced product over total bacterial proteins in 1,2, 3. and 4 hours after induction was 18. 2%, 18. 8%, 23.0% and 23.4%, respectively, by densitometry examination. CAM assay demonstrated that the recombinant canstatin protein significantly inhibited the embryonic neovascularization in a dose-dependent manner. Conclusion The prokaryotic expression vector of human canstatin gene has been successfully constructed, laying the foundation for further clinical study.
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Objective: To screen angiogenesis inhibitor from 26 drugs for promoting blood circulation and removing blood stasis through chick chorioallantoic membrane(CAM)model in vivo.Methods: The experimental groups were 26 drugs for promoting blood circulation and removing blood stasis.Tablets,capsules,pills and powder of the drugs were adopted in the form of saturated concentration,decoction was in the form of concentration of 2g/ml,and injection,oral liquid and electuary were in their own concentration.7-day-old fertilized white chicken eggs incubated at 37℃ in chamber were prepared by creating a window in the shell.A sponge carrier was placed on CAM and treated with different liquid medicine with the dosage of 20?l per egg daily for 3 days.On day 10,the number of CAM blood vessel embranchment 5mm around the carrier was counted with a stereomicroscope.Results: 15 drugs for promoting blood circulation and removing blood stasis obviously inhibited the CAM blood vessels(P