沉默Rictor对肝癌细胞体外血管生成、成瘤能力及P4HB/Hedgehog表达的影响

目的探究沉默Rictor对肝癌细胞体外血管生成、成瘤能力及P4HB/Hedgehog表达的影响。方法取肝癌细胞,将其分为肝癌细胞组(LC组)、肝癌细胞+Rictor-NC组(NC组)、肝癌细胞+siRictor组(SR组),荧光显微镜下观察各组转染情况,实时定量PCR法检测Rictor相关表达,细胞三维培养观察细胞形成拟态血管能力,裸鼠成瘤实验观察成瘤能力,免疫印迹法检测血管生成相关指标及P4HB/Hedgehog表达。结果LC组无转染,而NC组、SR组均可见绿色荧光,说明转染成功,且转染率均达90%以上,说明转染的细胞较为稳定;SR组细胞Rictor mRNA表达、血管形成数量、裸鼠瘤体生长曲线、血管内皮生长因子、P4HB、Hedgehog蛋白表达明显降低(P<0.05),说明沉默Rictor可促进肝癌细胞凋亡并抑制瘤体生成。结论沉默Rictor可显著抑制肝癌细胞体外血管生成,降低裸鼠成瘤能力,并抑制P4HB/Hedgehog表达。

ets-1基因的反义寡核苷酸对胃癌体外血管生成拟态的影响

目的:研究ets鄄1基因的反义寡核苷酸对胃癌细胞株SGC鄄7901体外血管生成拟态的影响。方法:将培养的SGC鄄7901细胞分为正义、反义和对照组;应用逆转录聚合酶链式反应检测ets鄄1mRNA的变化,克隆形成实验检测细胞增殖活性的变化,Transwell小室检测癌细胞体外侵袭力的变化,体外管状形成实验检测管道形成能力的变化。结果:转染ets鄄1反义寡核苷酸后,ets鄄1mRNA表达降低。ets鄄1基因的反义寡核苷酸对SGC鄄7901细胞增殖有明显抑制作用,可以显著降低SGC鄄7901细胞的体外侵袭能力,抑制体外管道形成。结论:ets鄄1基因的反义寡核苷酸能够抑制SGC鄄7901细胞的体外血管生成拟态形成。

体外血管生成模型及其应用

血管生成即新生血管从潜在血管生长点长出。它包括在许多生理和病理过程中,目前抗血管生成药物已成为抗肿瘤治疗的新手段。体外血管生成模型作为一种研究工具,在探讨血管形成机制、发现血管活性分子等方面将发挥十分积极的作用,还可以被用作药物筛选的工具。现分别介绍二维和三维体外血管生成模型,比较它们的优缺点,以及展望今后工作的方向。

耳、鼻、咽、喉

重组人血管内皮抑制素对鼻咽癌HNE-1细胞株体外血管生成拟态的抑制作用；uPARAP／Endo180在鼻咽癌中的表达及其意义；鼻咽癌组织COX-2基因表达及其意义的研究.

骨肿瘤

颈椎骨巨细胞瘤手术治疗的中长期随访及预后分析;唑来膦酸对骨肉瘤细胞体外血管生成拟态的影响。

Extract of Meretrix meretrix Linnaeus induces angiogenesis in vitro and activates endothelial nitric oxide synthase

Meretrix meretrix Linnaeus has long been used as traditional Chinese medicine in oriental medicine.The angiogentic activity of the extract of M.meretrix was investigated in this study,using human umbilical vein endothelial cells(HUVECs).Extract of M.meretrix Linnaeus(AFG-25) was prepared with acetone and ethanol precipitation,and further separated by Sephadex G-25 column.The results show that AFG-25 promoted proliferation,migration,and capillary-like tube formation in HUVECs,and in the presence of eNOS inhibitor NMA,the tube formation induced by AFG-25 is inhibited significantly.Moreover,AFG25 could also promote the activation of endothelial nitric oxide synthase(eNOS) and the resultant elevation of nitric oxide(NO) production.The results suggested that M.meretrix contains active ingredients with angiogentic activity and eNOS/NO signal pathway is in part involved in the proangiogenesis effect induced by AFG-25.

Anti-angiogenesis effect of Demethyl bryoanthrathiophene(DBT) in vitro

Objective:The aim of the study was to investigate the effect of Demethyl bryoanthrathiophene(DBT) on proliferations of human umbilical vein endothelial cells(HUVECs) and human lung adenocarcinoma cell line A549,and antiangiogenic effect of DBT on HUVECs in vitro.Methods:MTT assay was used to observe the effect of DBT on proliferations of HUVECs and A549 cells,flat plate scarification assay and tube formation in vitro test were used to observe the impact of DBT on migration and vaso-formed ability of HUVECs.The effects of DBT on apoptosis and cell cycle of HUVECs were calculated by flow cytometry.Results:MTT assay showed that treatment with DBT resulted in strong inhibition to the growth of HUVECs and A549 cells.The inhibition effects of DBT on HUVECs and A549 cells were related to dosage and times of dependency.In different doses of DBT(0.16,0.32 and 0.48 μmol/L) of flat plate scarification for 24 h,inhibition rates of DBT to migration of HUVECs were 14.70%,38.23% and 58.82%,respectively.In dose of DBT from 0.04,0.20 to 0.40 μmol/L for 24 h in tube formation,there were significance differences(P < 0.01) in the decreasing number of angiogenesis and incomplete blood vessel compared with control groups.All results showed that DBT promoted the apoptosis rate of HUVECs,and the increase of concentration of DBT accompanied the acceleration of apoptosis rate.Conclusion:DBT could inhibit the proliferations of HUVECs and A549 cells,and effectively suppress angiogenesis in vitro.

LOW MOLECULAR WEIGHT HEPARIN ENHANCES THE EFFECT OF aFGF IN ACCELERATING NEOVASCULA-RIZATION

Objective To explore the potential of low molecular weight heparin (LMWH) in combination cooperated with aFGF in accelerating neovascularization in vivo. Methods Ischemic model was set up in the right hindlimbs of 28 New Zealand white rabbits. Four groups of animals treated with saline, LMWH, aFGF and aFGF plus LMWH were allocated equally in group Ⅰ, group Ⅱ, group Ⅲ and group Ⅳ respectively. Vascular neovascularization and smooth muscular thickness of the ischemic hindlimb vessels of each animal in different groups were compared with each other on the 28th day postoperatively by angiography with DSA and the standard immunoperoxidase technique. Results No significant neovascularization was seen when aFGF adiministered in low dosage by venous infusion. But when the same dosage of aFGF plus LMWH were administered by venous infusion, a significant neovascularization was observed. Conclusion LMWH can potentiate aFGF in accelerating neovascularization.

Anti-angiogenesis effect of generation 4 polyamidoamine/vascular endothelial growth factor antisense oligodeoxynucleotide on breast cancer in vitro

Objective:To study the effects of the generation 4 polyamidoamine/vascular endothelial growth factor antisense oligodeoxynucleotide(G4PAMAM/VEGFASODN) compound on the expressions of vascular endothelial growth factor(VEGF) and its mRNA of breast cancer cells and on the inhibition of vascular endothelial cells. Methods:We examined the morphology of G4PAMAM/VEGFASODN compound and its pH stability,in vitro transfection efficiency and toxicity,and the expressions of VEGF and its mRNA. Methyl thiazolyl tetrazolium assay was used to detect the inhibitory function of the compound on vascular endothelial cells. Results:The compound was about 10 nm in diameter and was homogeneously netlike. From pH 5 to 10,it showed quite a buffered ability. The 48-h transfection rate in the charge ratio of 1:40 was 98.76%,significantly higher than that of the liposome group(P<0.05). None of the transfection products showed obvious toxicity on the cells. The expressions of both VEGF protein and its mRNA after G4PAMAM/VEGFASODN transfection decreased markedly. Conclusion:With a low toxicity,high safety,and high transfection rate,G4PAMAM/VEGFASODN could be a promising gene vector. Specifically,it inhibits VEGF gene expression efficiently,laying a basis for further in vivo animal studies.

A novel synthetic small molecule YF-452 inhibits tumor growth through antiangiogenesis by suppressing VEGF receptor 2 signaling

Tumor angiogenesis is characterized by abnormal vessel morphology, endowing tumor with highly hypoxia and unresponsive toward treatment. To date, mounting angiogenic factors have been discovered as therapeutic targets in antiangiogenic drug development. Among them, vascular endothelial growth factor receptor 2 （VEGFR2） inhibitors exerts potent antiangiogenic activity in tumor therapy. Therefore, it may provide a valid strategy for cancer treatment through targeting the tumor angiogenesis via VEGFR2 pathway. In this study, we established a high-profile compounds library and certificated a novel compound named N-（N-pyrrolidylacetyl）-9-（4-bromobenzyl）-l,3,4,9-tetrahydro-^-carboline （YF-452）, which remarkably inhibited the migration, invasion and tube-like structure formation of human umbilical vein endothelial cells （HUVECs） with little toxicity invitro. Rat thoracic aorta ring assay indicated that YF-452 significantly blocked the formation ofmicrovascular exvivo. In addition, YF-452 inhibited angiogenesis in chick chorioallantoic membrane （CAM） and mouse corneal micropocket assays. Moreover, YF-452 remarkably suppressed tumor growth in xenografts mice model. Furthermore, investigation of molecular mechanism revealed that YF-452 inhibited VEGF-induced phosphorylation of VEGFR2 kinase and the downstream protein kinases including extracellular signal regulated kinase （ERK）, focal adhesion kinase （FAK） and Src. These results indicate that YF-452 inhibits angiogenesis and may be a potential antiangiogenic drug candidate for cancer therapy.