Vasohibin‐2 is required for epithelial–mesenchymal transition of ovarian cancer cells by modulating transforming growth factor‐β signaling

Vasohibin‐2 (VASH2) is a homolog of VASH1, an endothelium‐derived angiogenesis inhibitor. Vasohibin‐2 is mainly expressed in cancer cells, and has been implicated in the progression of cancer by inducing angiogenesis and tumor growth. Although VASH2 has been recently reported to be involved in epithelial–mesenchymal transition (EMT), its precise roles are obscure. The aim of the present study was to clarify the role of VASH2 in the EMT of cancer cells in relation to transforming growth factor‐β (TGF‐β) signaling, which is a major stimulator of EMT. Decreased expression of VASH2 in ovarian cancer cells significantly repressed the expression of TGF‐β type I receptor, namely activin receptor‐like kinase 5. Transforming growth factor‐β1‐induced phosphorylation of Smad2 and Smad3 was markedly decreased in VASH2 knockdown cells while the expression of Smad2 and Smad3 was unchanged. Accordingly, the responses to TGF‐β1 shown by promoter assay and plasminogen activator inhibitor type 1 expression were significantly attenuated in VASH2 knockdown cells. Furthermore, knockdown of VASH2 in cancer cells abrogated the TGF‐β1‐induced reduced expression of epithelial markers including E‐cadherin, and the elevated expression of mesenchymal markers including fibronectin, ZEB2, and Snail2, suggesting that endogenous VASH2 is required for TGF‐β1‐induced EMT. In accordance with these results, the effects of TGF‐β1 on cell morphology, migration, invasion, and MMP2 expression were also abrogated when VASH2 was knocked down. These results indicate that VASH2 played a significant role in the EMT by modulating the TGF‐β signaling. We propose that VASH2 would be a novel molecular target for the prevention of EMT in cancers.     

VASH-2 与肿瘤关系的研究进展

血管生成参与肿瘤的发生、侵袭和转移,在肿瘤发展过程中起到重要作用。Vasohibin 家族是最近发现 的影响血管生成的调节因子,VASH-1 与 VASH-2 是该家族的重要成员,参与肿瘤生成和发展。VASH-1 在肿瘤 发病过程中起到负性调控作用,VASH-2 则促进血管生成。该文对 VASH-2 与肿瘤关系进行综述,探讨 VASH-2 在肿瘤中的作用,为血管靶向抗癌药物的研发提供新思路。     

Vasohibin‐1 expression inhibits advancement of ovarian cancer producing various angiogenic factors

Vasohibin‐1 (VASH1) is a negative feedback regulator of angiogenesis, the first to be discovered, and was identified in vascular endothelial growth factor (VEGF)‐stimulated vascular endothelial cells. Vasohibin‐1 inhibits abnormal vascularization induced by various angiogenic factors including fibroblast growth factor and platelet‐derived growth factor (PDGF), in addition to VEGF. By focusing on this characteristic of VASH1, we investigated the antitumor effects of VASH1 expression on ovarian cancer cells that produce different angiogenic factors. By using a high VEGF‐producing ovarian cancer cell line, SHIN‐3, and a high PDGF‐producing ovarian cancer cell line, KOC‐2S, the cells were transfected with either a VEGF antagonist, soluble VEGF receptor‐1 (sVEGFR‐1, or sFlt‐1), or VASH1 genes to establish their respective cellular expression. The characteristics of these transfectants were compared with controls. We previously reported that the expression of sFlt‐1 inhibited tumor vascularization and growth of high VEGF‐producing ovarian cancer cells, reduced peritoneal dissemination and ascites development, and prolonged the survival time of the host. However, in the current study, the expression of sFlt‐1 had no such effect on the high PDGF‐producing ovarian cancer cells used here, whereas VASH1 expression inhibited tumor vascularization and growth, not only in high VEGF‐producing cells, but also in high PDGF‐producing cells, reduced their peritoneal dissemination and ascites, and prolonged the survival time of the host. These results suggest that VASH1 is an effective treatment for ovarian cancer cells that produce different angiogenic factors.     

Targeting human vasohibin‐2 by a neutralizing monoclonal antibody for anti‐cancer treatment

There are two members of the vasohibin (VASH) family, VASH1 and VASH2. VASH1 is expressed mainly in endothelial cells to inhibit angiogenesis, whereas VASH2 is expressed mainly in cancer cells to stimulate tumor growth. The aim of the present study was to establish neutralizing monoclonal antibody (mAb) against human VASH2 and apply it as an anti‐cancer treatment. We previously raised mAb against several synthetic peptides of hVASH1, and found that one of them exhibited neutralizing activity against hVASH1. Because of the similarity in the amino acid sequences between VASH1 and VASH2, we hypothesized that they shared the bioactive center. When we mutated four amino acids within the region, the mutant VASH2 lost its pro‐angiogenic activity. Therefore, we raised mAb against a synthetic peptide overlapping the mutated amino acids of hVASH2, and isolated one clone (1760) that almost completely inhibited the stimulatory effect of hVASH2 on the migration of and tube formation by endothelial cells. When we used this clone 1760 antibody for cancer treatment, the peritoneal injection of it inhibited both tumor growth and angiogenesis in a mouse xenograft model of human cancer cells. In terms of anti‐tumor activity, 25 mg/kg of clone 1760 was equivalent to 5 mg/kg of bevacizmab. From these results, we propose the targeting of human VASH2 with neutralizing mAb as a new strategy for cancer treatment.     

联合应用多项肿瘤标志诊断卵巢恶性肿瘤的价值

目的探讨CA125等7种肿瘤标志联合应用对卵巢恶性肿瘤的诊断价值。方法对430例卵巢包块手术前患者(卵巢恶性肿瘤110例,卵巢良性肿瘤320例)及50例正常妇女血清应用ELISA法进行检测。检测项目包括糖类抗原CA125(CA125)、肿瘤相关物质(TSGF)、唾液酸(SA)、癌胚抗原(CEA)、甲胎蛋白(AFP)、促性腺激素(hCG)和铁蛋白(Fer)。结果卵巢恶性肿瘤患者血清中CA125、TSGF、SA、CEA、AFP及Fer水平明显高于卵巢良性肿瘤或对照组,F=177.24,P<0.0001;F=52.49,P<0.0001;F=3.38,P=0.0347;F=6.88,P=0.0011;F=34.94,P<0.0001;F=8.23,P=0.0003;F=124.37,P<0.0001。在7项肿瘤标志中,CA125单独诊断价值最大。单独应用CA125诊断卵巢恶性肿瘤的敏感性、特异性及准确性分别为86.4%、82.8%及83.7%。联合应用7项肿瘤标志时,以任意一项及一项以上异常指标为诊断标准时,诊断卵巢恶性肿瘤的敏感性、特异性及准确性分别为95.5%、45.6%和58.4%;以任意两项及两项以上异常指标为诊断标准时,分别为93.6%、80.6%和84.0%;以任意3项及3项以上异常指标为诊断标准时,分别为87.3%、90.3%和89.5%。结论7项肿瘤标志联合检测对提高卵巢恶性肿瘤诊断的敏感性、特异性及准确性有一定意义,其中CA125、TSGF及SA3项肿瘤标志阳性诊断价值最大。     

卵巢癌组织中Vasohibin基因表达与启动子区域甲基化的关系

目的探讨卵巢上皮性肿瘤中Vaso-hibin基因mRNA表达与临床特征及其启动子甲基化的关系。方法用RT-PCR检测10例良性卵巢上皮性肿瘤、8例交界性卵巢上皮性肿瘤及30例卵巢癌组织中VasohibinmRNA表达;用甲基化特异聚合酶链反应方法(MSP)检测Vasohibin启动子甲基化。结果各组标本中都有VasohibinmRNA表达,3组Vasohib-in表达的相对值良性卵巢上皮性肿瘤为0.65±0.11,交界性卵巢上皮性肿瘤为0.47±0.13,卵巢癌为0.34±0.14。卵巢癌组织中Vasohibin的表达量较交界性卵巢上皮性肿瘤组织显著降低,t=2.479,P=0.029;交界性卵巢上皮性肿瘤组织的表达量较良性卵巢上皮性肿瘤显著下降,t=3.198,P=0.006。在卵巢癌组织中,Vasohibin的表达与不同临床分期、不同分化程度以及有无淋巴结转移无关。良性卵巢上皮性肿瘤中未发现Vasohibin基因启动子甲基化;交界性卵巢上皮性肿瘤中2例发生启动子甲基化,卵巢癌中11例发生启动子甲基化,甲基化率36.7%。卵巢癌中启动子甲基化组织较未甲基化组织VasohibinmRNA表达显著降低,t=4.671,P=0.000。结论Vasohibin与卵巢癌发生存在明显相关性。Vasohibin启动子甲基化与其mRNA表达抑制密切相关,可能是Vasohibin在卵巢癌中低表达的最主要因素之一。     

VASH2对脑胶质瘤血管新生的影响机制

目的 通过观察血管抑制素-2(VASH2)对脑胶质瘤微血管内皮细胞增殖、迁移和管形成的调控作用,探究其对脑胶质瘤血管新生的影响机制。方法 构建人脑胶质瘤微血管内皮细胞模型,通过稳定转染获得VASH2过表达和表达沉默的细胞系,在后续实验中分组,分别为空白对照组、VASH2过表达阴性对照空质粒组[VASH2(+)NC 组]、VASH2过表达组[VASH2(+)组]、VASH2表达沉默阴性对照空质粒组[VASH2 (-) NC 组]和VASH2表达沉默组[VASH2(-)组]。通过细胞生长抑制实验检测细胞增殖能力的变化;体外管形成实验检测细胞管形成能力的变化;细胞迁移实验检测细胞迁移能力的变化。应用 GraphPad Prism v5.01统计软件进行数据分析。多组间比较采用单因素方差分析,组间两两比较采用Newman-Keuls分析。结果 在人脑胶质瘤微血管内皮细胞中上调VASH2的表达,VASH2过表达组与 VASH2过表达阴性对照空质粒组相比较,细胞的增殖能力、迁移能力和管形成能力均明显增强(P<0.05) ,表明VASH2过表达能够促进胶质瘤血管的新生。在人脑胶质瘤微血管内皮细胞中下调VASH2的表达,VASH2表达沉默组与VASH2表达沉默阴性对照空质粒组相比较,细胞的增殖能力、迁移能力和管形成能力均明显减弱(P<0.05),表明VASH2沉默能够抑制胶质瘤血管的新生。结论 VASH2在胶质瘤管形成的调控方面扮演了重要的角色,可能为胶质瘤的抗血管治疗提供新策略。     

vasohibin基因重组腺病毒对胶质瘤U251移植瘤的抑制作用

目的探讨vasohibin基因重组腺病毒对胶质瘤U251移植瘤的抑制作用。方法构建vasohibin基因重组腺病毒,观察其对人脐静脉血管内皮细胞(HUVEC)体外增殖的影响。建立胶质瘤U251移植瘤模型,观察vasohibin基因重组腺病毒对胶质瘤U251移植瘤生长的抑制作用。结果 vasohibin基因重组腺病毒能明显抑制HUVEC的增殖,显著减缓胶质瘤U251移植瘤生长(P<0.05),同时明显减少了移植瘤组织微血管密度(P<0.05)。结论 vasohibin基因重组腺病毒能明显抑制胶质瘤U251移植瘤的生长与血管生成,为胶质瘤的基因治疗提供了新的思路。     

血管生成抑制蛋白vasohibin的研究进展

血管新生参与机体重要的生理和病理过程。Vasohibin是新近发现的调节血管生成的内皮源性负反馈调节因子,对抑制血管生成起重要作用。Vasohibin因参与肿瘤﹑视网膜疾病及类风湿性关节炎等血管新生异常疾病的发生发展而备受关注,相关研究可望为探索这些疾病的发生机制以及寻找新的治疗措施奠定基础。