Vascular Endothelial Growth Factor and Other Signaling Pathways in Developmental and Pathologic Angiogenesis

The field of angiogenesis received a huge boost in 2003 with the announcement of positive results in a phase III clinical trial using a vascular endothelial growth factor (VEGF)-blocking antibody for the treatment of cancer. Although the VEGF pathway has emerged as a central signaling pathway in normal and pathologic angiogenesis, several other pathways are also now recognized as playing essential roles. This review focuses on 2 specific areas. First, we summarize some of the work on newly discovered angiogenic signaling pathways by primarily describing the molecular biology of the pathways and the evidence for their involvement in vascular development. Second, we describe progress in therapeutic antiangiogenesis in cancer, particularly with agents that block the VEGF pathway.     

Notch配体Delta-1ike1对小鼠骨髓细胞来源的树突状细胞分化和抗原呈递功能的影响

本研究探讨Notch配体Delta—likel（Dil1）对小鼠骨髓细胞来源的树突状细胞（dendriticcell,DC）分化及其抗原呈递功能的影响。在GM—CSF和IL4存在条件下,用OP9-Dil1和OPt）．GFP细胞分别与小鼠骨髓细胞共培养8天,经肿瘤抗原刺激成熟。用流式细胞仪检测DC表面MHCII、CD80和CD86的表达情况,ELISA法检测肿瘤抗原刺激后DC培养上清细胞因子IL-12和IL110的水平,通过混合T淋巴细胞反应观察DC对T细胞的促增殖能力。结果表明：与GFP组相比,Dll1组的小鼠骨髓来源的树突状细胞明显增多（P〈0．05）。肿瘤抗原刺激后,Dll1组DC表面MHCII、CD80和CD86表达量更高。DC分泌的IL-12水平显著高于对照组（P〈0．05）,而IL-10的水平显著低于对照组（P〈0．01）。Dll1组DC具有更强的促T细胞增殖活性。结论：OP9-Dll1促进小鼠骨髓细胞向DC分化并增强其抗原呈递功能。     

Delta-like 4在胰腺癌中的表达及其意义

目的 检测Delta-like 4(DLL4)在胰腺癌中的表达,探讨其临床意义.方法 采用免疫组化法检测60例胰腺癌组织中DLL4的表达,行血管内皮特异性标志物CD34免疫染色,计算微血管密度(MVD);分析它们的相关性及其与胰腺癌临床病理特征、患者预后的关系.结果 DLL4在胰腺癌组织中的表达明显高于正常胰腺组织(68.3％比20.0％,x2=14.239,P＜0.01),其高表达与胰腺癌的肿瘤分化程度、TNM分期、淋巴结转移及累及范围密切相关,与肿瘤大小、部位、病理分型无关.胰腺癌组织MVD明显高于正常胰腺组织(34.9±13.2比18.9 ±2.2,t=3.570,P＜0.01),其与肿瘤分化程度、TNM分期、淋巴结转移及累及范围密切相关,与肿瘤大小、部位及病理分型无关.DLL4阳性表达与预后密切相关.经Cox模型分析,TNM分期、DLL4阳性表达是胰腺癌患者预后不良的独立危险因素.结论 DLL4的高表达在胰腺癌转移、浸润中起重要作用.肿瘤DLL4表达和TNM分期对评估胰腺癌患者的预后有一定价值.     

Signaling of endothelial cytoprotection in transplantation

A better knowledge of the processes by which endothelium can resist to cell death and adapt to injury by specific intracellular signaling pathways and dedicated protein regulation is a key step to understand how vascular inflammation/injury develops and how it is regulated. This review focuses on signaling pathways and molecular effectors that trigger the balance between endothelial cell activation and dysfunction. In addition to the canonical nuclear factor-κB (NF-κB), phosphatidyl inositol 3-kinase (PI-3K) and mitogen-activated protein kinases (MAPK) that orchestrated the inflammatory response and its termination we report here additive pathways such as Notch pathway and protein C/protease activated receptor (PAR) pathway that have been also reported to play a role in the control of EC activation and apoptosis. This review also provides an update of the characteristics of some established and novel protective molecules for the endothelium, identified in transplantation.     

Delta-like ligand 4 in hepatocellular carcinoma intrinsically promotes tumour growth and suppresses hepatitis B virus replication

AIM To investigate the role of Delta-like ligand 4(DLL4) on tumour growth in hepatitis B virus(HBV)-associated hepatocellular carcinoma(HCC) in vivo.METHODS We suppressed DLL4 expression in an HBV expressing HCC cell line, HepG2.2.15 and analysed the growth ability of cells as subcutaneous tumours in nude mice. The expression of tumour angiogenesis regulators, VEGF-A and VEGF-R2 in tumour xenografts were examined by western blotting. The tumour proliferation and neovasculature were examined by immunohistochemistry. The viral replication and viral protein expression were measured by quantitative PCR and western blotting, respectively.RESULTS Eighteen days after implantation, tumour volume in mice implanted with sh DLL4 HepG2.2.15 was significantly smaller than in mice implanted with control HepG2.2.15(P 0.0001). The levels of angiogenesis regulators, VEGF-A and VEGF-R2 were significantly decreased in implanted tumours with suppressed DLL4 compared with the control group(P 0.001 and P 0.05, respectively). Furthermore, the suppression of DLL4 expression in tumour cells reduced cell proliferation and the formation of new blood vessels in tumours. Unexpectedly, increased viral replication was observed after suppression of DLL4 in the tumours.CONCLUSION This study demonstrates that DLL4 is important in regulating the tumour growth of HBV-associated HCC as well as the neovascularization and suppression of HBV replication.     

Notch-Dll4在激光诱导的大鼠脉络膜新生血管生成中的作用

目的探讨Notch信号通路中Delta样配体4(Delta-like ligand4,Dll4)在激光诱导的大鼠脉络膜新生血管(choroidal neovascularization,CNV)生成中的作用机制。方法将雄性棕色挪威(brown-Norway,BN)大鼠分为正常组、光凝组、实验组和对照组。光凝组、实验组、对照组利用532nm激光诱导建立双眼CNV模型。实验组在建立CNV模型后立刻玻璃体内注射25g.L-1Avastin10μL,对照组注射0.01mmol.L-1PBS10μL。采用免疫荧光染色检测Dll4和血管内皮生长因子(vascular endo-thelial growth factor,VEGF)在正常组和光凝组光凝后7d中的表达;采用Western-blotting和RT-PCR检测正常组和光凝组(光凝后1d、3d、7d、14d)实验组、对照组(注射后7d)Dll4和VEGF的蛋白和mRNA表达;通过HE染色和视网膜色素上皮-脉络膜-巩膜铺片检测实验组和对照组(注射后14d)CNV生成的厚度和面积。结果Dll4和VEGF在CNV区域有共表达。Dll4和VEGF蛋白及其mRNA的表达趋势...     

Dynamic maternal and fetal Notch activity and expression in placentation

IntroductionMurine placentation requires trophoblast Notch2, while the Notch ligand, JAGGED1, is reduced in invasive trophoblasts from women with preeclampsia. However, the placental cells with active Notch signaling and expression of other Notch proteins and ligands in placentation have yet to be defined. We sought to identify endothelial cell and trophoblast subtypes with canonical Notch signaling in the decidua and placenta and correlate this to expression of Notch proteins and ligands.MethodsNotch reporter transgenic mice were used to define canonical Notch activity and immunofluorescence staining performed to characterize expression of Notch1, 2, 3, 4 and ligands, Delta-like 4 (Dll4) and Jagged1 (Jag1) during early placentation and in the mature placenta.ResultsNotch signaling is active in maternal and fetal endothelial cells and trophoblasts during early placentation and in the mature placenta. Dll4, Jag1, Notch1, and Notch4 are expressed in maternal vasculature in the decidua. Dll4, Jag1 and Notch1 are expressed in fetal vasculature in the labyrinth. Dll4, Notch2 and Notch4 are co-expressed in the ectoplacental cone. Notch2 and Notch4 are expressed in parietal-trophoblast giant cells and junctional zone trophoblasts with active canonical Notch signaling and in labyrinthine syncytiotrophoblasts and sinusoidal-trophoblast giant cells.DiscussionCanonical Notch activity and distinct expression patterns for Notch proteins and ligands was evident in endothelium and trophoblasts, suggesting Notch1, Notch2, Notch4, Dll4, and Jag1 have distinct and overlapping functions in placentation. Characterization of Notch signaling defects in existing mouse models of preeclampsia may shed light on the role of Notch in developing the preeclampsia phenotype.     

Efficacy and Safety of Rovalpituzumab Tesirine Compared With Topotecan as Second-Line Therapy in DLL3-High SCLC: Results From the Phase 3 TAHOE Study

IntroductionDLL3, an atypical Notch ligand, is expressed in SCLC tumors but is not detectable in normal adult tissues. Rovalpituzumab tesirine (Rova-T) is an antibody-drug conjugate containing a DLL3-targeting antibody tethered to a cytotoxic agent pyrrolobenzodiazepine by means of a protease-cleavable linker. The efficacy and safety of Rova-T compared with topotecan as second-line therapy in patients with SCLC expressing high levels of DLL3 (DLL3-high) was evaluated.MethodsThe TAHOE study was an open-label, two-to-one randomized, phase 3 study comparing Rova-T with topotecan as second-line therapy in DLL3-high advanced or metastatic SCLC. Rova-T (0.3 mg/kg) was administered intravenously on day 1 of a 42-day cycle for two cycles, with two additional cycles available to patients who met protocol-defined criteria for continued dosing. Topotecan (1.5 mg/m²) was administered intravenously on days 1 to 5 of a 21-day cycle. The primary end point was overall survival (OS).ResultsPatients randomized to Rova-T (n = 296) and topotecan (n = 148) were included in the efficacy analyses. The median age was 64 years, and 77% had the extensive disease at initial diagnosis. The median OS (95% confidence interval) was 6.3 months (5.6–7.3) in the Rova-T arm and 8.6 months (7.7–10.1) in the topotecan arm (hazard ratio, 1.46 [95% confidence interval: 1.17–1.82]). An independent data monitoring committee recommended that enrollment be discontinued because of the shorter OS observed with Rova-T compared with topotecan. Safety profiles for both drugs were consistent with previous reports.ConclusionsCompared with topotecan, which is the current standard second-line chemotherapy, Rova-T exhibited an inferior OS and higher rates of serosal effusions, photosensitivity reaction, and peripheral edema in patients with SCLC. A considerable unmet therapeutic need remains in this population.     

Midbrain expression of Delta-like 1 homologue is regulated by GDNF and is associated with dopaminergic differentiation

Affymetrix GeneChip technology and quantitative real-time PCR (Q-PCR) were used to examine changes in gene expression in the adult murine substantia nigra pars compacta (SNc) following lentiviral glial cell line-derived neurotrophic factor (GDNF) delivery in adult striatum. We identified several genes that were upregulated after GDNF treatment. Among these, the gene encoding the transmembrane protein Delta-like 1 homologue (Dlk1) was upregulated with a greater than 4-fold increase in mRNA encoding this protein. Immunohistochemistry with a Dlk1-specific antibody confirmed the observed upregulation with increased positive staining of cell bodies in the SNc and fibers in the striatum. Analysis of the developmental regulation of Dlk1 in the murine ventral midbrain showed that the upregulation of Dlk1 mRNA correlated with the generation of tyrosine hydroxylase (TH)-positive neurons. Furthermore, Dlk1 expression was analyzed in MesC2.10 cells, which are derived from embryonic human mesencephalon and capable of undergoing differentiation into dopaminergic neurons. We detected upregulation of Dlk1 mRNA and protein under conditions where MesC2.10 cells differentiate into a dopaminergic phenotype (41.7+/-7.1% Dlk1+ cells). In contrast, control cultures subjected to default differentiation into non-dopaminergic neurons only expressed very few (3.7+/-1.3%) Dlk1-immunopositive cells. The expression of Dlk1 in MesC2.10 cells was specifically upregulated by the addition of GDNF. Thus, our data suggest that Dlk1 expression precedes the appearance of TH in mesencephalic cells and that levels of Dlk1 are regulated by GDNF.