Overexpression of vascular endothelial growth factor 165 (VEGF165) protects cardiomyocytes against doxorubicin-induced apoptosis

Doxorubicin (Dox) has been employed in cancer chemotherapy for a few decades. However its clinical application became restricted because of dose-dependent cardiomyopathy. Recent studies suggest that Dox-induced cardiomyocyte apoptosis is a primary cause of cardiac damage. Vascular endothelial growth factor (VEGF) is a major factor for endothelial cell survival and angiogenesis. We have previously shown that VEGF165 significantly attenuates oxidative stress-induced cardiomyocytes apoptosis. We hypothesized that VEGF165 will protect the cardiomyocytes from Dox-induced apoptosis. to evaluate our hypothesis, we transfected cardiomyocytes H9c2 with adenovirus expressing VEGF165 24 hours before the cells were challenged with Dox at a concentration of 2 μm. Cardiomyocyte apoptosis was evaluated by Annexin V-FITC staining and by Western blot detection of cleaved caspase-3. The hypothesis was confirmed, and the protective mechanisms involve the inhibition of death receptor-mediated apoptosis and up-regulation of the prosurvival Akt/Nf-κb/bcl-2 signaling pathway.     

PTEN基因转染对白血病细胞VEGF调控作用的影响

目的:探讨在白血病细胞中与张力蛋白同源的10号染色体缺失的磷酸酶基因(phosphatase and tensin hemology deleted on chromosome ten gene,PTEN)对血管内皮生长因子(vascular endothelial growth factor,VEGF)及其受体1(VEGF receptor 1,VEGFR1)调控作用的影响.方法:将携带有野生型PTEN及绿色荧光蛋白(green fluorescent protein,GFP)基因的腺病毒(Ad-PTEN-GFP)及空载体腺病毒(Ad-GFP)转染人慢性粒细胞白血病急变细胞株K562,实时荧光定量PCR法检测不同转染组细胞中PTEN、VEGF和VEGF1 mRNA表达水平,Western印迹法检测PTEN、VEGF、Akt和磷酸化Akt蛋白的表达水平,并采用Transwell小室侵袭实验检测不同转染组细胞的侵袭性.通过MTT实验及FCM法检测PTEN基因对脐静脉内皮细胞株ECV304增殖和凋亡的影响.通过鸡胚尿囊膜(chick chorioallantoic membrane,CAM)体内血管生长实验检测PTEN基因对鸡胚血管生成的影响.结果:与空载体腺病毒(Ad-GFP)相比,Ad-PTEN-GFP 转染人白血病细胞K562后,VEGF及其受体的表达被明显抑制,并呈剂量依赖性负相关;同时,Ad-PTEN-GFP 转染后K562细胞侵袭能力明显减弱.PTEN基因转染能够抑制血管内皮细胞ECV304增殖,并促进其细胞凋亡,细胞周期阻滞在S期.PTEN基因转染可明显抑制CAM血管生长.结论:肿瘤抑制基因PTEN能够抑制内皮细胞增殖和白血病细胞侵袭,其作用机制可能是负调控白血病细胞VEGF表达以及抑制肿瘤血管新生.     

VEGF硫代反义寡核苷酸抑制U937细胞VEGF的表达

Objective： To study the effect of VEGF antisense oligodeoxynucleotide （VEGF ASODN） on VEGF expression in acute monocyte leukemic cell line U937 in vitro. Methods： U937 cells were incubated with VEGF ASODN （final concentration as follows： 10, 20 and 30 μmol/L respectively） or scrambled sequence, compared with negative control. The expression of VEGF mRNA was measured by semi-quantitative RT-PCR, VEGF protein was measured by Western blot. Results： VEGF ASODN obviously inhibited expression of VEGF mRNA in U937 cell, compared with scrambled sequence and negative control （P〈0.05）. And the inhibition effect was most remarkable after 24 h, which is related with the dose of VEGF ASODN （P〈0.05）. Scrambled sequence groups had no significant difference compared with negative control groups （P〉0.05）. VEGF ASODN obviously inhibited expression of VEGF protein, compared with scrambled sequence and negative control （P〈0.05）. Conclusion： The expressions of VEGF at mRNA and protein levels in leukemic cell line U937 are down-regulated after being treated with VEGF ASODN.     

Ribozyme approach to downregulate vascular endothelial growth factor (VEGF) 189 expression in non-small cell lung cancer (NSCLC)

The aim of this study was to further clarify the role of the cell-associated isoform of vascular endothelial growth factor (VEGF189) on tumour growth and vascularity. Five isoforms of VEGF have been identified with different biological activities. VEGF121, VEGF145, VEGF165, VEGF189, VEGF206 are generated by alternative splicing. We used a hammerhead-type ribozyme (V189Rz) to suppress VEGF189 mRNA. The V189Rz specifically cleaved exon 6 of VEGF189 mRNA, but showed no activity against the VEGF121 or VEGF165 isoforms. The V189Rz was introduced into the human non-small cell lung cancer (NSCLC) cell line (OZ-6/VR). The expression level of VEGF189 mRNA was decreased in the OZ-6/VR cells, while VEGF121 and 165 expression was unaltered. The OZ-6/VR cells xenotransplanted into nude mice showed markedly reduced vascularisation and growth, whereas the cell line did not show any decreased growth under tissue culture conditions. The OZ-6/VR cells (1×10⁵ cells/mouse) formed no tumours, whereas the parental OZ-6 cells formed large tumours within 8 weeks. The specific suppression of VEGF189 by the ribozyme decreased vascularity and xenotransplantability of the lung cancer cell line. Thus, the cell-associated isoform of VEGF, VEGF189, might have a key role in stromal vascularisation and the growth of NSCLC xenografts in vivo.     

Inhibition of K562 leukemia angiogenesis and growth by expression of antisense vascular endothelial growth factor (VEGF) sequence

Vascular endothelial growth factor (VEGF), a major angiogenic factor, plays a key role in the growth of solid tumor. Recently, expression of VEGF and its receptors has been found on leukemic cells as well as on endothelial cells. VEGF may fulfill a fundamental role in promoting tumor angiogenesis and proliferation by stimulating both endothelial cells and leukemic cells. To investigate the role of VEGF in the angiogenesis and growth of leukemic cell, we used an antisense strategy to downregulate VEGF expression in K562 cells, a human erythroleukemia cell line. Expression of antisense-VEGF in K562 cells reduced the secretion of VEGF protein and inhibited cell survival. The proliferation and migration of human umbilical vein endothelial cells were decreased in response to the conditioned medium (CM) from K562 cells expressed antisense-VEGF, compared to CM from K562 cells transfected with vector control. Moreover, subcutaneous injection of nude mice with antisense-VEGF K562 cells inhibited tumor growth with a reduction of the density of microvessels and an increased apoptosis in those tumors, compared to vector control K562 cells. These results suggest that the efficient downregulation of the VEGF production in leukemic cells using antisense-VEGF may constitute a novel strategy of treatment in leukemia.     

Imaging gastrointestinal tumours using vascular endothelial growth factor-165 (VEGF165) receptor scintigraphy.

BACKGROUND: Recent studies have shown that vascular endothelial growth factor (VEGF) receptor is overexpressed in vascular endothelial cells of various human tumours as well as in human tumour cells. The aim of this study was to evaluate the usefulness of scanning with VEGF(165) labeled with (123)I for tumor localisation in patients with gastrointestinal tumours. PATIENTS AND METHODS: Human recombinant VEGF(165) was radiolabelled with (123)I by electrophilic radioiodination using the chloramine T method. [(123)I]VEGF(165) was administered intravenously [mean dose 184 +/- 18 MBq (相似文献   

Vascular endothelial growth factor (VEGF) expression in plasmacytoma

Vascular endothelial growth factor (VEGF) plays an important role in angiogenesis. Although the role and importance of angiogenic factors such as VEGF have been established in various solid tumors, this has not been widely evaluated in hemopoietic neoplasias. In this trial, VEGF was studied in plasmacytoma and VEGF expression was compared with histopathologic grade. Forty-seven samples have taken from cases with plasmacytoma (Pm) (33 bones and 14 soft tissue plasmacytomas) were used as study material. Pm was the initial presentation in all cases, and bone marrow (BM) involvement was detected in 19 cases with systemic evaluation. Twenty-seven of the cases were male (age range was between 19 and 81 years). Histopathologically 27 cases had mature and 20 cases had immature morphology. Immunohistochemical analysis was used to detect VEGF expression and this was scored according to the percentage of the VEGF stained cells. VEGF expression was detected in 32 cases and in eight cases this expression was strong. In 11 cases expression was moderate and 13 cases showed mild expression. When we compared VEGF expression with pathologic grade, 17 of 20 immature samples showed VEGF expression while 15 of 27 mature samples showed VEGF expression. There was a statistically significant association between immature morphology and VEGF expression (p < 0.0264). Additionally all the samples, except one, with strong VEGF expression showed immature morphology. In conclusion two thirds of the cases with Pm showed VEGF expression and this was associated with immature morphology. Increased expression of VEGF was seen in plasmacytomas, and additional studies are needed to determine whether this translates to increased microvasculature or increased risk of progression to myeloma.     

All three receptors for vascular endothelial growth factor (VEGF) are expressed on B-chronic lymphocytic leukemia (CLL) cells

B-chronic lymphocytic leukemia (B-CLL) cells have a long survival owing to an alteration in the normal pathways of apoptosis. CLL cells have been found to produce and secrete vascular endothelial growth factor (VEGF). In addition to its major role in angiogenesis, VEGF affects cell survival by interfering with apoptosis. The aim of the present study was to investigate the expression of the VEGF receptors VEGFR-1, VEGFR-2, and VEGFR-3 on B-CLL cells, singly and combined. B-CLL cells were isolated from peripheral blood drawn from patients with CLL. Total VEGF receptor, examined in 13 samples by flow cytometry was present in all cases with mean CD19+/VEGF+ expression of 76% (range 52-92%). Specific receptor expression, examined in 27 samples by immunocytochemical methods, was positive for VEGFR-1 in all 27 patients and for VEGFR-2 and VEGFR-3 in 26 (96%). These findings suggest that the VEGF transduction pathway may be very active in CLL cells, and both its paracrine and autocrine pathways may contribute to their enhanced survival.     

Progestins suppress estrogen-induced expression of vascular endothelial growth factor (VEGF) subtypes in uterine endometrial cancer cells.

Vascular endothelial growth factor (VEGF) contributes to the early advancement of uterine endometrial cancers that conserve hormone dependency via angiogenic activity. This process prompted us to study sex steroidal suppression of VEGF expression in Ishikawa cells (a line of well-differentiated uterine endometrial cancer cells). Estrogen transiently induced VEGF subtype (VEGF165 and VEGF121) secretion from Ishikawa cells. Progestins (progesterone, medroxyprogesterone acetate (MPA) and 17 alpha-hydroxyprogesterone) suppressed the estrogen-induced events. In conclusion, progestins could suppress VEGF-related angiogenic potential, which contributes to tumor growth in the early stage of uterine endometrial cancers that conserve estrogen dependency.     

Use of soluble recombinant decoy receptor vascular endothelial growth factor trap (VEGF Trap) to inhibit vascular endothelial growth factor activity

Primary tumors and metastases require blood vessel formation to support their continued growth and eventual metastasis. They use existing vasculature during initial growth but eventually must orchestrate the development and maintenance of new vessels--a process termed angiogenesis--to grow beyond a small size and spread. Angiogenesis is regulated by a number of soluble factors, the relative proportions of which can exacerbate or inhibit the process. Vascular endothelial growth factor (VEGF) is a potent stimulator of angiogenesis, produced by the majority of human solid tumors. Inhibitors of VEGF might have an impact on the growth and metastasis of these cancers. The relevance of this strategy to the treatment of colorectal cancer was first successfully demonstrated in human clinical trials using a monoclonal antibody against VEGF. A potent antiangiogenic soluble recombinant decoy, VEGF Trap is a protein constructed from VEGF receptor-binding domains linked to an immunoglobulin G(1) constant region. It possesses an affinity for VEGF that is significantly higher than that of the monoclonal antibody. VEGF Trap has demonstrated marked efficacy in halting angiogenesis and shrinking tumors in preclinical animal models and is currently being studied in phase I clinical trials in humans with advanced solid malignancies.     

血管内皮细胞生长因子影响难治性急性髓系白血病发生发展的作用研究

背景和目的:难治性白血病的发生发展机制非常复杂,近年来发现白血病患者骨髓过度表达血管内皮细胞生长因子(vascularendothelialgrowthfactor,VEGF),但VEGF如何影响难治性白血病尚未明确。本研究探讨高表达VEGF对人急性髓系白血病细胞株HL-60增殖及三尖杉酯碱诱导HL-60细胞凋亡影响;观察难治性急性髓系白血病(acutemyeloidleukemia,AML)中VEGF蛋白表达与疾病发展的相关性。方法:采用脂质体转染VEGF165cDNA入HL-60细胞,通过RT-PCR及ELISA方法鉴定转染克隆HL-60/VEGF165细胞中VEGFmRNA及蛋白的表达,MTT法、集落形成实验比较HL-60/VEGF165及转染pcDNA3.1-neo空载体的HL-60/neo细胞增殖活性,流式细胞仪观察三尖杉酯碱诱导的细胞凋亡。用ELISA法检测难治性与非难治性AML患者血浆中VEGF蛋白浓度。结果:HL-60/VEGF165细胞培养上清中VEGF蛋白浓度为(399.07±12.45)ng/L,高于HL-60/neo细胞(184.45±10.53)ng/L(P<0.01)。HL-60/VEGF165细胞与HL-60/neo细胞相比生长速度明显加快,集落形成能力明显增加,集落数分别为(157.00±17.00)/500细胞和(110.00±12.90)/500细胞(P<0.05);而细胞凋亡率减少,分别为(3.18±0.33)%和(6.61±0.50)%,三尖杉酯碱诱导HL-60/VEGF165细胞凋亡率低于HL-60/neo细胞。难治性AML患者血浆中VE     

Blockage of vascular endothelial growth factor (VEGF) reduces experimental pleurodesis

Background and objective

Chemical pleurodesis controls recurrent malignant pleural effusion. The mechanism that determines pleural symphysis involves the action of vascular endothelial growth factor (VEGF). We assessed the influence of the anti-VEGF antibody (bevacizumab) on pleurodesis induced by talc or silver nitrate and analyzed the temporal development of pleural angiogenesis.

Methods

Sixty New Zealand rabbits received intrapleural injection (2 mL) of talc (400 mg/kg) or 0.5% silver nitrate. In each group, half of the animals received an intravenous injection of bevacizumab 30 min before the sclerosing agent. Five animals from each group were euthanized 7, 14, or 28 days after the procedure. Adhesions and inflammation (scores: 0-4), thickness (μm), vascular density (vessels/field), and collagen fibers (μm²) were evaluated in the visceral pleura.

Results

Antibody anti-VEGF interferes in pleurodesis induced by talc or silver nitrate. Pleural inflammation was discreet with no difference between the groups, regardless the anti-VEGF treatment. Concerning the vascular density of the visceral pleura, a smaller number of neoformed vessels was noted in the animals that received bevacizumab. In the animals receiving silver nitrate, the decrement in adhesions and vascular density was associated with reduced thick and thin collagen fibers, resulting in less pleural thickness.

Conclusion

The anti-VEGF antibody inhibits adhesions between pleural layers. Despite being an experimental study in animals with normal pleura, the results call attention to a likely lack of success in pleurodesis when VEGF blockers are used.     

抗VEGF发卡状核酶抑制肝癌VEGF表达和移植瘤生长

目的:采用抗人血管内皮生长因子(vascular endothelial growth factor,VEGF)发卡状核酶基因转染肝癌细胞,观察核酶对VEGF表达及肿瘤生长的影响。方法:采用脂质体介导的方法,将人工合成的抗VEGF发卡状核酶基因转染肝癌细胞SMMC-7721,同时制备空载体和细胞对照,经G418筛选获得阳性克隆,基因组水平鉴定核酶在细胞中的表达,半定量RT-PCR法和免疫组化法观察核酶对SMMC-7721细胞VEGF表达的影响。接种各组细胞于裸鼠皮下,观察瘤体体积大小和质量,免疫组化法观测各组肿瘤微血管变化和VEGF的表达。结果:核酶基因成功导入到肿瘤细胞中,转基因细胞的增殖速率明显下降(P<0.01),转染核酶组细胞VEGF水平明显下降(P<0.01)。移植瘤成瘤率和生长速度减慢(P<0.01),肿瘤组织VEGF表达和血管形成减少(P<0.01)。结论:抗人VEGF发卡状核酶基因在体内、体外均可抑制肝癌VEGF表达,通过减少血管形成抑制肿瘤生长,为进一步开展肝癌血管靶向基因治疗提供实验依据。     

Immunohistochemical evaluation of vascular endothelial growth factor (VEGF) in colorectal carcinoma

BACKGROUND: Vascular endothelial growth factor (VEGF) is one of the most important proangiogenic factors over-expressed in many human cancers and is usually associated with an unfavorable outcome. This work was planned to assess VEGF and microvessel density (MVD) in colorectal carcinoma (CRC) and to correlate them with the available clinicopathological parameters. MATERIAL AND METHODS: Ten normal colonic mucosa, 21 adenoma and 70 CRC cases were examined by immunohistochemical staining using anti-VEGF antibody. RESULTS: Eighty-one percent (81%) of adenoma and 78.6% of CRC cases showed VEGF immunoreactivity; however, the intensity of staining was significantly in favour of malignant cases (p=0.032). VEGF immunostaining in CRC was correlated with low grade (p=0.009), early stage either by Dukes' system (p=0.03) or TNM stage (p=0.03), negative nodal status (p=0.04) and high mast cell count (p=0.04). MVD assessed by Haematoxylin and Eosin staining was associated with dense inflammatory response (p=0.003) and high mast cell count (p=0.006). CONCLUSIONS: VEGF could be an early carcinogenic factor in CRC that declines with its progression. Inflammatory cells including mast cells could play a role in CRC angiogenesis.     

Prognostic significance of microvessel density and vascular endothelial growth factor (VEGF) expression in non-Hodgkin's lymphoma

Angiogenesis has a major role in the pathogenesis of malignancies. Studies involving the role of angiogenesis have been most commonly performed in solid tumors. However, studies related to hemapoietic neoplasia and angiogenesis are relatively limited. We investigated the role of angiogenesis in non-Hodgkin's lymphomas (NHLs) and its relation with clinical and histopathologic prognostic indicators. In this respect, angiogenesis markers were evaluated in 71 patients with NHL and these were compared with other prognostic indicators including age, gender, histological grade, stage, extranodal involvement and survival. Microvessel density (MVD) using Factor VIII monoclonal antibody and vascular endothelial growth factor (VEGF) using monoclonal antibody for VEGF expression were studied in paraffin-embedded tissue samples. We did not find a significant relationship between MVD and patient characteristics including age, gender, stage, histological grade, nodal status, international prognostic index (IPI), and response to treatment. MVD was found to be greater in cases with B symptoms compared to those without B symptoms (14.6±5.7 and 11.4±5.3, respectively, p=0.002). No significant relationship was found between VEGF and age, gender, stage, histological grade, IPI, and overall survival. The complete and partial response rate to therapy was significantly higher in VEGF-negative patients than in the VEGF-positive patients (p=0.003). In conclusion, there appears to be a role for angiogenesis and angiogenic factors in NHLs. The combination of anti-angiogenic drugs with conventional anti-neoplastic treatment will probably be used in the future. Larger series of patients are needed to determine the prognostic value of angiogenesis in NHL.