血管内皮生长因子家族及其受体与肿瘤血管生成研究进展

血管内皮生长因子(vascular endothelial growth factor,VEGF),又名血管通透性因子(vascular permeability factor,VPF)是重要的血管生成正性调节因子,是目前抗癌治疗的研究靶点之一。现已发现的VEGF家族成员包括VEGF—A、VEGF—B、VEGF—C、VEGF—D、VEGF—E和胎盘生长因子(placenta growth factor,PLGF)。VEGF的受体有VEGFR—1(fit—1)、VEGFR-2(flk-1／KDR)、VEGFR-3(fit-4)、neuropilin(NPR1／NPR2)。该家族的成员可以选择性地增强血管和／或淋巴管内皮细胞的有丝分裂,刺激内皮细胞增殖并促进血管生成,提高血管特别是微小血管的通透性,使血浆大分子外渗沉积在血管外的基质中,促进新生毛细血管网的建立,为肿瘤细胞的生长提供营养等。作者对VEGF家族成员及其受体的理化特征、VEGF与肿瘤的关系、VEGF抑制剂的研制作一综述。     

血管内皮生长因子-C与肿瘤细胞转移

血管内皮生长因子(vascular endothelial growth factor,VEGF)C通过与其受体VEGFR2／VEGFR3结合,影响到肿瘤生长、肿瘤外周血管生成、淋巴血管形成、肿瘤表面积,促进肿瘤细胞从原发部位通过血液以及淋巴系统转移到其他器官,是引起肿瘤治疗失败的主要原因之一。阻碍VEGF-C与受体的结合可以降低肿瘤细胞的转移性,为肿瘤治疗提供了一个新的靶点。     

血管内皮生长因子受体的信号转导通路

血管内皮生长因子受体(VEGFR)是VEGF的特异性受体,由于在刺激血管内皮细胞增殖、迁移、管腔形成,促进肿瘤生长和转移过程中起着重要的作用,而成为抗肿瘤新生血管生成的热点。该主要围绕VEGF及其不同受体的信号转导通路作一综述。     

血管内皮生长因子受体信号转导通路与肿瘤血管生成

血管内皮生长因子是促进血管生成的重要调节因子.它能促进内皮细胞增殖、迁移,阻止内皮细胞凋亡、管腔网状结构退化,增加血管渗透性.所有这些作用都是通过血管内皮生长因子受体信号转导通路实现的.它们在肿瘤血管生成、肿瘤生长中起着重要的作用.以血管内皮生长因子受体信号转导通路为靶点是开发肿瘤血管生成抑制剂的理想策略.     

血管内皮生长因子的神经保护作用

血管内皮生长因子（vascular endothelial growth factor,VEGF）是内皮细胞特异性的生长因子,大多数关于VEGF的研究都是致力于其在血管生长方面的作用,而近年来有大量文献报道VEGF具有神经营养和促神经发生作用,它能够直接作用于神经元细胞和神经胶质细胞甚至是神经干细胞,促进其生长及存活。VEGF的多种功能使其和多种神经退行性疾病相关,如阿茨海默病,肌萎缩侧索硬化症,帕金森病等。导入VEGF基因能够改善肌萎缩侧索硬化症、帕金森病动物模型的病情。     

血管内皮生长因子及其受体在斑马鱼胚胎血管发育中的作用

血管内皮生长因子(vascular endothelial growth factor,VEGF)是一种多功能的细胞因子,其主要作用是促进血管内皮细胞增殖和增加血管通透性,是肿瘤及正常组织血管生成的中心调控因素。以VEGF为靶点的肿瘤血管靶向性治疗成为近几年肿瘤治疗的新途径。斑马鱼作为一种重要的模式生物,被广泛用于胚胎的分子发育机制、疾病模型的构建以及药物筛选等研究中。文章对斑马鱼作为心血管系统研究模型的优势及其血管研究方法做一阐述,重点对斑马鱼VEGF及其受体的最新研究进展做了介绍,并展望了其发展前景。     

血管内皮生长因子和抗肿瘤血管新生药物研究进展

肿瘤的生长与迁移离不开新血管的形成,这使得抗血管新生成为肿瘤治疗的重要途径之一。血管内皮生长因子（VEGF）是针对内皮细胞作用最强、特异性最高的血管新生促进因子,因而VEGF是抗肿瘤治疗的重要靶点。我们简要介绍了VEGF的一些生物学特点及肿瘤血管新生,着重介绍了一些抗血管新生药物的最新研究成果及其临床应用。     

抗VEGF/VEGFR靶向肿瘤血管药物的研究进展

研究表明,肿瘤的生长转移和新血管的生成有密切关系,其中血管内皮细胞生长因子(vascular endothelial growth factor,VEGF)及其信号途径在肿瘤血管生成中起关键作用。阻断该途径的任何环节均可有效抑制肿瘤血管的生成,进而抑制肿瘤的生长和转移。近年来,已有多种以VEGF/VEGFR为靶点的抗肿瘤血管生成药物投入临床应用,其中bevacizumab为第一个获批上市的抗肿瘤血管生成药物。继bevacizumab后,一种以基因工程手段获得的人Fc融合蛋白Zaltrap也成功在美国上市,这种杂交分子的药代动力学明显优于单克隆抗体,能更好的遏制肿瘤血管的发生并消退已形成的肿瘤血管。在肿瘤的临床治疗中,Zaltrap比bevacizumab显示出更大的优势。此外,VEGFC/D Trap及小分子酪氨酸激酶抑制剂也能有效抑制肿瘤血管的生成。在此对以VEGF/VEGFR为靶点的抗肿瘤血管生成药物进行综述。     

VEGF 家族及其在肿瘤生长中作用的研究

血管内皮生长因子(Vascular Endothelial Growth Factor,VEGF)家族是一类多功能的细胞因子,在血管生成和淋巴管生成中具有直接和间接的调控作用,可促进内皮细胞增殖、促进血管生成以及增加血管的通透性。VEGF/VEGFR轴由多重配基和受体质量叠加交错组成,并且受体与配基结合具有专一性,在不同的细胞中具有不同的细胞类型表达和功能.启动VEGF信号通路,触发了一个网状的信号过程,从而促进血管内皮细胞生长、转移和存活。进来研究发现,VEGF的一个重要作用表现为可动员内皮祖细胞从骨髓向远处转移从而形成新生血管,因而有必要设计和发展针对这一途径的抑制因子。随着研究的深入,VEGF促进肿瘤血管生成的作用和与人类癌症的发病机制的关系是确定的,因此,抑制VEGF途径被确认为是一种重要的有效的抗癌模式     

VEGF-C与肿瘤细胞转移

血管内皮生长因子C(VEGF—C)通过与其受体VEGFR2／VEGFR3结合,影响到肿瘤生长、肿瘤外周血管生成、淋巴血管形成、肿瘤表面积,促进肿瘤细胞从原发部位通过血液以及淋巴系统转移到其他器官,是引起肿瘤治疗失败的主要原因之一。阻碍VEGF—C与受体的结合可以降低肿瘤细胞的转移性,为肿瘤治疗提供了1个新的靶点。     

Glucocorticoid effects on angiogenesis are associated with mTOR pathway activity

Glucocorticoids (GC) often are administered during pregnancy, but despite their widespread use in clinical practice, it remains uncertain how GC exposure affects pro-angiogenic factors and their receptors. We investigated the effects of GC on vascular endothelial growth factor (VEGF), placental growth factor (PIGF), vascular endothelial growth factor receptor 1 (VEGFR1) and vascular endothelial growth factor receptor 2 (VEGFR2) protein and mRNA expressions and investigated the possible association of GC with the Akt/mTOR pathway. We incubated human umbilical vein endothelial cells (HUVECs) with a synthetic GC, triamcinolone acetonide (TA). TA administration caused decreased cellular and soluble VEGF and VEGFR1 protein expressions and increased soluble VEGFR2 expression. VEGF, VEGFR1 and VEGFR2 mRNA expressions were altered in a time and dose dependent manner. PIGF protein expression was unaffected by TA treatment, but PIGF mRNA expression decreased in a dose dependent manner after incubation for 48 and 72 h. Phospho-mTOR and phospho-Akt expressions were unaffected. Phospho-p70S6K and phospho-4EBP1 protein expressions and the vascular network forming capacity of HUVECs decreased in a dose dependent manner. We found that GC exert detrimental effects on angiogenesis by altering cellular and soluble angiogenic protein and mRNA levels, and vascular network forming capacities by the Akt/mTOR pathway.     

Knockdown of VEGFR2 Inhibits Proliferation and Induces Apoptosis in Hemangioma-Derived Endothelial Cells

Angiogenesis is a process of development and growth of new capillary blood vessels from pre-existing vessels. Angiogenic growth factors play important roles in the development and maintenance of some malignancies, of which vascular endothelial growth factor (VEGF)/VEGFR2 interactions are involved in proliferation, migration, and survival of many cancer cells. The aim of this study was to investigate the function of VEGFR2 in human hemangiomas (HAs). Using immunohistochemistry assay, we examined the expression levels of VEGF, VEGFR2, Ki-67, glucose transporter-1 (Glut-1), phosphorylated protein kinase B (p-AKT) and p-ERK in different phases of human HAs. Positive expression of VEGF, VEGFR2, Ki-67, Glut-1, p-AKT and p-ERK was significantly increased in proliferating phase HAs, while decreased in involuting phase HAs (P=0.001; P=0.003). In contrast, cell apoptotic indexes were decreased in proliferating phase HAs, but increased in involuting phase HAs (P<0.01). Furthermore, we used small hairpin RNA (shRNA)-mediated VEGFR2 knockdown in primary HA-derived endothelial cells (HemECs) to understand the role of VEGF/VEGFR2 signaling. Knockdown of VEGFR2 by Lv-shVEGFR2 inhibited cell viability and induced apoptosis in primary HemECs companied with decreased expression of p-AKT, p-ERK, p-p38MAPK and Ki-67 and increased expression of caspase-3 (CAS-3); Overexpression of VEGFR2 promoted cell viability and blocked apoptosis in Lv-VEGFR2-transfected HemECs. Taken together, our findings demonstrate that, increased expression of VEGFR2 is involved in the development of primary HemECs possibly through regulation of the AKT and ERK pathways, suggesting that VEGFR2 may be a potential therapeutic target for HAs.Key words: vascular endothelial growth factor receptor 2, hemangioma, proliferation, apoptosis     

Selective Inhibition of Vascular Endothelial Growth Factor Receptor‐2 (VEGFR‐2) Identifies a Central Role for VEGFR‐2 in Human Aortic Endothelial Cell Responses to VEGF

Abstract

Vascular endothelial growth factor receptors (VEGFR) are considered essential for angiogenesis. The VEGFR‐family proteins consist of VEGFR‐1/Flt‐1, VEGFR‐2/KDR/Flk‐1, and VEGFR‐3/Flt‐4. Among these, VEGFR‐2 is thought to be principally responsible for angiogenesis. However, the precise role of VEGFRs1–3 in endothelial cell biology and angiogenesis remains unclear due in part to the lack of VEGFR‐specific inhibitors. We used the newly described, highly selective anilinoquinazoline inhibitor of VEGFR‐2 tyrosine kinase, ZM323881 (5‐[[7‐(benzyloxy) quinazolin‐4‐yl]amino]‐4‐fluoro‐2‐methylphenol), to explore the role of VEGFR‐2 in endothelial cell function. Consistent with its reported effects on VEGFR‐2 [IC(50) < 2 nM], ZM323881 inhibited activation of VEGFR‐2, but not of VEGFR‐1, epidermal growth factor receptor (EGFR), platelet‐derived growth factor receptor (PDGFR), or hepatocyte growth factor (HGF) receptor. We studied the effects of VEGF on human aortic endothelial cells (HAECs), which express VEGFR‐1 and VEGFR‐2, but not VEGFR‐3, in the absence or presence of ZM323881. Inhibition of VEGFR‐2 blocked activation of extracellular regulated‐kinase, p38, Akt, and endothelial nitric oxide synthetase (eNOS) by VEGF, but did not inhibit p38 activation by the VEGFR‐1‐specific ligand, placental growth factor (PlGF). Inhibition of VEGFR‐2 also perturbed VEGF‐induced membrane extension, cell migration, and tube formation by HAECs. Vascular endothelial growth factor receptor‐2 inhibition also reversed VEGF‐stimulated phosphorylation of CrkII and its Src homology 2 (SH2)‐binding protein p130^Cas, which are known to play a pivotal role in regulating endothelial cell migration. Inhibition of VEGFR‐2 thus blocked all VEGF‐induced endothelial cellular responses tested, supporting that the catalytic activity of VEGFR‐2 is critical for VEGF signaling and/or that VEGFR‐2 may function in a heterodimer with VEGFR‐1 in human vascular endothelial cells.     

VEGFR2 Trafficking,Signaling and Proteolysis is Regulated by the Ubiquitin Isopeptidase USP8

Vascular endothelial growth factor A (VEGF‐A) regulates many aspects of vascular function. VEGF‐A binding to vascular endothelial growth factor receptor 2 (VEGFR2) stimulates endothelial signal transduction and regulates multiple cellular responses. Activated VEGFR2 undergoes ubiquitination but the enzymes that regulate this post‐translational modification are unclear. In this study, the de‐ubiquitinating enzyme, USP8, is shown to regulate VEGFR2 trafficking, de‐ubiquitination, proteolysis and signal transduction. USP8‐depleted endothelial cells displayed altered VEGFR2 ubiquitination and production of a unique VEGFR2 extracellular domain proteolytic fragment caused by VEGFR2 accumulation in the endosome–lysosome system. In addition, perturbed VEGFR2 trafficking impaired VEGF‐A‐stimulated signal transduction in USP8‐depleted cells. Thus, regulation of VEGFR2 ubiquitination and de‐ubiquitination has important consequences for the endothelial cell response and vascular physiology.      

Vascular endothelial growth factor (VEGF) receptor II-derived peptides inhibit VEGF

Vascular endothelial growth factor (VEGF) directly stimulates endothelial cell proliferation and migration via tyrosine kinase receptors of the split kinase domain family. It mediates vascular growth and angiogenesis in the embryo but also in the adult in a variety of physiological and pathological conditions. The potential binding site of VEGF with its receptor was identified using cellulose-bound overlapping peptides of the extracytosolic part of the human vascular endothelial growth factor receptor II (VEGFR II). Thus, a peptide originating from the third globular domain of the VEGFR II comprising residues 247RTELNVGIDFNWEYP261 was revealed as contiguous sequence stretch, which bound 125I-VEGF165. A systematic replacement with L-amino acids within the peptide representing the putative VEGF-binding site on VEGFR II indicates Asp255 as the hydrophilic key residue for binding. The dimerized peptide (RTELNVGIDFNWEYPAS)2K inhibits VEGF165 binding with an IC50 of 0.5 microM on extracellular VEGFR II fragments and 30 microM on human umbilical vein cells. VEGF165-stimulated autophosphorylation of VEGFR II as well as proliferation and migration of microvascular endothelial cells was inhibited by the monomeric peptide RTELNVGIDFNWEYPASK at a half-maximal concentration of 3-10, 0.1, and 0.1 microM, respectively. We conclude that transduction of the VEGF165 signal can be interrupted with a peptide derived from the third Ig-like domain of VEGFR II by blockade of VEGF165 binding to its receptor.     

Signal transduction by VEGF receptors in regulation of angiogenesis and lymphangiogenesis

The VEGF/VPF (vascular endothelial growth factor/vascular permeability factor) ligands and receptors are crucial regulators of vasculogenesis, angiogenesis, lymphangiogenesis and vascular permeability in vertebrates. VEGF-A, the prototype VEGF ligand, binds and activates two tyrosine kinase receptors: VEGFR1 (Flt-1) and VEGFR2 (KDR/Flk-1). VEGFR1, which occurs in transmembrane and soluble forms, negatively regulates vasculogenesis and angiogenesis during early embryogenesis, but it also acts as a positive regulator of angiogenesis and inflammatory responses, playing a role in several human diseases such as rheumatoid arthritis and cancer. The soluble VEGFR1 is overexpressed in placenta in preeclampsia patients. VEGFR2 has critical functions in physiological and pathological angiogenesis through distinct signal transduction pathways regulating proliferation and migration of endothelial cells. VEGFR3, a receptor for the lymphatic growth factors VEGF-C and VEGF-D, but not for VEGF-A, regulates vascular and lymphatic endothelial cell function during embryogenesis. Loss-of-function variants of VEGFR3 have been identified in lymphedema. Formation of tumor lymphatics may be stimulated by tumor-produced VEGF-C, allowing increased spread of tumor metastases through the lymphatics. Mapping the signaling system of these important receptors may provide the knowledge necessary to suppress specific signaling pathways in major human diseases.     

BMPs promote proliferation and migration of endothelial cells via stimulation of VEGF-A/VEGFR2 and angiopoietin-1/Tie2 signalling

The differentiation, growth, and survival of endothelial cells (ECs) are regulated by multiple signalling pathways, such as vascular endothelial growth factors (VEGFs) and angiopoietins through their receptor tyrosine kinases, VEGF receptor (VEGFR) 2 and Tie2, respectively. Bone morphogenetic proteins (BMPs), members of the transforming growth factor (TGF)-beta family, have been implicated in the development and maintenance of vascular systems. However, their effects on EC proliferation remain to be elucidated. In the present study, we show that BMPs induce the proliferation and migration of mouse embryonic stem cell (ESC)-derived endothelial cells (MESECs) and human microvascular endothelial cells (HMECs). Addition of BMP-4 to culture induced significant proliferation and migration of both types of ECs. BMP-4 also increased the expression and phosphorylation of VEGFR2 and Tie2. These findings suggest that BMP signalling activates endothelium via activation of VEGF/VEGFR2 and Angiopoietin/Tie2 signalling.     

Expression of vascular endothelial growth factor receptor 1 in bone marrow-derived mesenchymal cells is dependent on hypoxia-inducible factor 1

Bone marrow-derived cells are recruited to sites of ischemia, where they promote tissue vascularization. This response is dependent upon the expression of vascular endothelial growth factor (VEGF) receptor 1 (VEGFR1), which mediates cell migration in response to VEGF or placental growth factor (PLGF). In this study, we found that exposure of cultured mouse bone marrow-derived mesenchymal stromal cells (MSC) to hypoxia or an adenovirus encoding a constitutively active form of hypoxia-inducible factor 1 (HIF-1) induced VEGFR1 mRNA and protein expression and promoted ex vivo migration in response to VEGF or PLGF. MSC in which HIF-1 activity was inhibited by a dominant negative or RNA interference approach expressed markedly reduced levels of VEGFR1 and failed to migrate or activate AKT in response to VEGF or PLGF. Thus, loss-of-function and gain-of-function approaches demonstrated that HIF-1 activity is necessary and sufficient for basal and hypoxia-induced VEGFR1 expression in bone marrow-derived MSC.