Expression and function of vascular endothelial growth factor receptors (Flt-1 and Flk-1) in vascular adventitial fibroblasts

Vascular endothelial growth factor receptors (VEGFRs) are previously considered to exist exclusively in endothelial cells. However, little is known if the receptors are expressed in other non-endothelial cells. In this study, we measured activation of two VEGFRs, Flk-1 and Flt-1, and their biological functions in cultured adventitial fibroblasts and injured rat carotid injury arteries induced by balloon angioplasty. Our results indicated that Flt-1, but not Flk-1, existed in adventitial fibroblasts. Angiotensin II increased Flt-1 protein expression in a time- and concentration-dependent manner. Adventitial fibroblast migration stimulated by vascular endothelial growth factor (VEGF) and placental growth factor (PIGF) required Flt-1 expression. The Flt-1-induced adventitial fibroblast migration was blocked by anti-Flt-1 neutralizing antibody and soluble VEGFR1 protein (sFlt-1). However, Flt-1 activation did not enhance cell proliferation. In addition, Flt-1 expression was significantly increased in the neointima and adventitia in injured rat carotid arteries. We concluded that functional expression of Flt-1 in adventitial fibroblasts might be an important mediator in the pathogenesis of vascular remodeling after arterial injury.     

乙型肝炎病毒X蛋白对肝细胞癌生长因子激活作用的研究

目的研究乙型肝炎病毒Ｘ（hepatitisBvirus,HBx）蛋白对生长因子激活作用,探讨HBx蛋白对肝细胞癌生长的影响。方法构建Ｘ基因真核表达载体,转染入HepG2细胞后,细胞组织化学和免疫印迹法检测胰岛素样生长因子－Ⅰ受体（insulin-likegrowthfactorⅠreceptor,IGF-ⅠR）和血管内皮生长因子（vascularendothelialgrowthfactor,VEGF）表达。结果转染HBx基因HepG2细胞（X细胞）IGF-ⅠR表达阳性率为84%±3%,转染空载体对照细胞（X0细胞）为26%±4%;免疫印迹法检测Ｘ细胞有明显条带。VEGF表达Ｘ细胞阳性率为83%±5%,X0细胞为28%±6%,差异有非常显著意义;Ｘ细胞IGF-ⅠR表达增高近1.5倍。结论HBx蛋白可同时激活IGF-ⅠR和VEGF,在肿瘤恶性生长、转移中起有重要作用。     

Gene transfer of kringle 1-5 suppresses tumor development and improves prognosis of mice with hepatocellular carcinoma

BACKGROUND & AIMS: Recent studies indicate that kringle 1-5 has a potent and specific antiangiogenic activity. Here, we investigated the antitumor effect of kringle 1-5 gene transfer on hepatocellular carcinoma in mice. METHODS: The inhibitory effect of kringle 1-5 protein on proliferation of bovine capillary endothelial cells was evaluated by a tetrazolium-based assay. To study tumor growth, intrahepatic metastasis, and survival, liposome/kringle 1-5 complementary DNA complexes were injected intravenously in nude mice preimplanted with 1 of 3 hepatoma cell lines into the liver. Production of kringle 1-5 was tested by immunohistochemistry and Western blotting. Intratumoral vessel density was quantified. Expression of vascular endothelial growth factor, angiopoietin-1, and angiopoietin-2 in tumors was examined by Western blotting. Serum alanine aminotransferase and alpha-fetoprotein levels and body weights were measured. RESULTS: Proliferation of bovine capillary endothelial cells was inhibited by purified kringle 1-5 in a dose-dependent manner. Gene transfer of kringle 1-5 caused a significant reduction in vessel density with suppression of tumor growth of the 3 hepatoma cell lines and serum alpha-fetoprotein levels, prolonged the survival period, and reduced the number of intrahepatic metastases. Among the analyzed angiogenic factors, kringle 1-5 reduced angiopoietin-2 expression levels. Expression of kringle 1-5 protein was detected on hepatoma cells and hepatocytes in the liver. However, it did not alter serum alanine aminotransferase levels and body weights, suggesting kringle 1-5 lacks severe side effects. CONCLUSIONS: Antiangiogenic gene therapy with kringle 1-5 complementary DNA is a promising safe and effective strategy for suppression of growth of hepatocellular carcinoma.     

Endothelin inhibits cholangiocarcinoma growth by a decrease in the vascular endothelial growth factor expression

Background: Endothelins (ET‐1, ET‐2, ET‐3) are peptides with vasoactive properties interacting with ET_A and ET_B receptors. ET‐1 inhibits secretin‐stimulated ductal secretion (hallmark of cholangiocyte growth) of cholestatic rats by interaction with ET receptors. Aim: The aims of the studies were to evaluate (i) the effect of ET‐1 on cholangiocarcinoma growth in Mz‐ChA‐1 cells and nude mice and (ii) whether ET‐1 regulation of cholangiocarcinoma growth is associated with changes in the expression of vascular endothelial growth factor‐A (VEGF‐A), VEGF‐C, VEGF receptor‐2 (VEGFR‐2) and VEGFR‐3. Methods: We determined the expression of ET_A and ET_B receptors on normal and malignant (Mz‐ChA‐1) cholangiocytes and human cholangiocarcinoma tissue and the effect of ET‐1 on the proliferation and expression of VEGF‐A, VEGF‐C (regulators of tumour angiogenesis) and its receptors, VEGFR‐2 and VEGFR‐3, in Mz‐ChA‐1 cells. In vivo, Mz‐ChA‐1 cells were injected into the flanks of athymic mice and injections of ET‐1 or saline into the tumours were performed daily. The effect of ET‐1 on tumour size, cell proliferation, apoptosis, collagen quantity and the expression of VEGF‐A and VEGF‐C and VEGFR‐2 and VEGFR‐3 were measured after 73 days. Results: Higher expression of ET_A and ET_B was observed in malignant compared with normal cholangiocytes. ET‐1 inhibited proliferation and VEGF‐A, VEGF‐C, VEGFR‐2 and VEGFR‐3 expression of Mz‐ChA‐1 cells. Chronic ET‐1 treatment decreased tumour volume, tumour cell proliferation and VEGF‐A and VEGF‐C expression but increased apoptosis and collagen tissue deposition compared with controls. Conclusions: Modulation of VEGF‐A and VEGF‐C (by ET‐1) may be important for managing cholangiocarcinoma growth.     

Apatinib as an alternative therapy for advanced hepatocellular carcinoma

Angiogenesis plays an important role in the occurrence and development of tumors. Registered tyrosine kinase inhibitors targeting vascular endothelial growth factor reduce angiogenesis. Apatinib, a tyrosine kinase inhibitor, can specifically inhibit vascular endothelial growth factor receptor 2, showing encouraging anti-tumor effects in a variety of tumors including advanced hepatocellular carcinoma(HCC). This article intends to review the clinical research and application prospects of apatinib in the field of HCC.     

肝细胞癌组织细胞因子信号转导抑制因子-1表达的研究

目的探讨细胞因子信号转导抑制因子(SOCS1)、血管内皮生长因子受体(VEGFR)-2和表皮生长因子受体(EGFR)在肝硬化、肝细胞癌组织中的表达变化。方法采用免疫组化法检测63例HCC组织、51例癌周肝组织、11例肝硬化组织及11例正常肝组织SOCS1蛋白的表达;另分别检测VEGFR2和EGFR在27例和41例HCC组织中的表达情况。结果癌周肝组织中SOCS1蛋白表达为阴性1例,弱阳性15例,阳性30例,强阳性5例,HCC组织中SOCS1蛋白表达为阴性11例,弱阳性38例,阳性14例,癌周肝组织中SOCS1蛋白表达强度显著高于HCC组织(P0.01);SOCS1蛋白在肝硬化组织和正常肝组织中无表达;SOCS1表达在瘤体大小间(瘤体5cm和≥5cm)有显著性差异(P0.001);SOCS1蛋白在HCC组织中的表达与VEGFR2和EGFR的表达无显著相关性(P0.05)。结论 SOCS1蛋白表达与HCC的发生发展密切相关。     

An Engineered Heparin-Binding Form of VEGF-E (hbVEGF-E). Biological effects in vitro and mobilizatiion of precursor cells

Vascular endothelial growth factor (VEGF-A) is the founding member of a family of angiogenic proteins with various binding abilities to three cognate VEGF receptors. Previously, a gene encoding from the genome of parapox orf virus (OV) with about 25% amino acid identity to mammalian VEGF-A was named VEGF-E and shown to bind and specifically activate the vascular endothelial growth factor receptor VEGFR-2 (KDR/flk-1). Here, we have generated a novel heparin-binding form of VEGF-E by introducing the heparin-domain of the human VEGF-A(165) splice variant into the viral VEGF-E protein. Recombinant heparin-binding VEGF-E (hbVEGF-E) is shown to stimulate proliferation and sprout formation of macro- and microvascular endothelial cells to a similar extent as the parental OV-VEGF-E but fails to activate peripheral mononuclear cells. However, hbVEGF-E is more potent in binding competition assays with primary human endothelial cells when compared to the OV-VEGF-E. This can be explained by our finding that binding of hbVEGF-E but not of parental OV-VEGF-E to the VEGFR-2 is strongly increased by the addition of neuropilin-1 (NP-1), a cognate co-receptor for VEGF-A. The engineered hbVEGF-E was compared with the VEGFR-1 selective and also heparin-binding form of placenta growth factor (PlGF-2) in vivo. Both heparin-binding homologues induced mobilization of endothelial progenitor cells from the bone marrow and gave rise to similar colony numbers of myeloic cells in a colony-forming assay. These findings suggest that both VEGFR-1 and VEGFR-2 are involved in stem cell mobilization.     

喉癌组织中VEGF-C及其受体VEGFR-3和D2-40的表达及意义

目的 观察血管内皮生长因子C（VEGF-C）及其受体（VEGFR-3）和D2-40在喉癌组织中的表达情况,探讨其在喉癌淋巴道转移过程中的作用及可能机制.方法 选择喉癌患者的癌组织、癌旁正常组织标本各40份,同期选择正常喉组织标本19份作对照.采用免疫组织化学法检测各标本中VEGF-C、VEGFR-3和D240的表达.结果 VEGF-C、VEGFR-3及D2-40在喉癌组织、癌旁组织及正常喉组织中均有表达,且其表达强度呈下降趋势（H分别为38.400、32.502,P均＜0.01）;在喉癌组织中,VEGF-C、VEGFR-3及D2-40的表达与淋巴转移、临床分期及临床分型有关（P均＜0.05）,与病理分级、肿瘤大小、吸烟量、年龄和性别无关（P均＞0.05）.喉癌组织中VEGF-C与VEGFRd的表达呈正相关（r=0.882,P＜0.01）.结论 VEGF-C、VEGFR-3及D2-40在喉癌组织中的表达均明显高于癌旁组织及正常喉组织,其高表达可能与喉癌的发生、发展有关.