Spatiotemporal heterogeneity of PPARγ expression in porcine uteroplacenta for regulating of placental angiogenesis through VEGF-mediated signalling

Non-infectious prenatal mortality severely affects the porcine industry, with pathological placentation as a likely key reason. Previous studies have demonstrated that peroxisome proliferator-activated receptor gamma (PPARγ) deficiency causes defects in the uteroplacental vasculature and induces embryonic losses in mice. However, its role in porcine placental angiogenesis remains unclear. In the present study, PPARγ expression was investigated in porcine uteroplacental tissues at gestational day (GD) 25, GD40 and GD70 via quantitative polymerase chain reaction (qPCR), Western blot and immunohistochemistry (IHC). Moreover, the roles of PPARγ in porcine placental angiogenesis were investigated using a cell model of porcine umbilical vein endothelial cells (PUVECs) to conduct proliferation, migration and tube formation assays in vitro and a mouse xenograft model to assess capillary formation in vivo. The results showed that PPARγ was mainly located in the glandular epithelium, trophoblast, amniotic chorion epithelium and vascular endothelium, as indicated by the higher expression levels at GD25 and GD40 than at GD70 in endometrium and by higher expression levels at GD40 and GD70 than at GD25 in placenta. Moreover, PPARγ expression was significantly downregulated in placenta with dead foetus. In PUVECs, knocking out PPARγ significantly inhibited proliferation, migration and tube formation in vitro and inhibited capillary formation in mouse xenografts in vivo by blocking S-phase, promoting apoptosis and downregulating the angiogenic factors of VEGF and its receptors. Overall, the spatiotemporal heterogeneity of PPARγ expression in porcine uteroplacental tissue suggests its vital role in endometrial remodelling and placental angiogenesis, and PPARγ regulates placental angiogenesis through VEGF-mediated signalling.     

血管内皮生长因子对缺血性脑血管病血管生成的作用研究

血管内皮生长因子是血管特异性生长因子,有促进内皮细胞增生、转移,增加血管通透性和促进血管生成的作用.阐述了血管内皮生长因子的生物学特性和功能,以及血管内皮生长因子与缺血性脑血管病的关系.     

富硒麦芽对二乙基亚硝胺诱发大鼠肝癌血管生成的影响

试验选择雄性SD大鼠100只,体重100~120 g,随机分成3组。试验Ⅰ组日粮中添加富硒麦芽（SEM）使硒含量达到3.0 mg/kg;试验Ⅱ组为模型组;试验Ⅲ组为阴性对照组,不诱癌不加SEM;试验Ⅱ、Ⅲ日粮中分别添加亚硒酸钠使硒含量达到0.1 mg/kg。试验Ⅰ、Ⅱ组饮水中添加100 mg/L二乙基亚硝胺（diethylnitrosamine,DEN）诱癌,维持DEN摄入量每天约10 mg/kg体重,连续16周,然后改饮普通灭菌水至18周末。试验Ⅲ组始终以普通灭菌水自由饮用。18周末,处死大鼠。利用免疫组织化学法分析大鼠肝肿瘤微血管密度（microvessel density,MVD）和血管内皮生长因子（VEGF）的表达。结果表明,SEM组MVD和VEGF表达较模型组显著降低。因此,表明SEM对肝肿瘤血管生成有抑制作用,下调VEGF表达可能是其抑制肿瘤血管生成的主要机理之一。     

Heparanase and Syndecan-4 Are Involved in Low Molecular Weight Fucoidan-Induced Angiogenesis

Induction of angiogenesis is a potential treatment for chronic ischemia. Low molecular weight fucoidan (LMWF), the sulfated polysaccharide from brown seaweeds, has been shown to promote revascularization in a rat limb ischemia, increasing angiogenesis in vivo. We investigated the potential role of two heparan sulfate (HS) metabolism enzymes, exostosin-2 (EXT2) and heparanase (HPSE), and of two HS-membrane proteoglycans, syndecan-1 and -4 (SDC-1 and SDC-4), in LMWF induced angiogenesis. Our results showed that LMWF increases human vascular endothelial cell (HUVEC) migration and angiogenesis in vitro. We report that the expression and activity of the HS-degrading HPSE was increased after LMWF treatment. The phenotypic tests of LMWF-treated and EXT2- or HPSE-siRNA-transfected cells indicated that EXT2 or HPSE expression significantly affect the proangiogenic potential of LMWF. In addition, LMWF increased SDC-1, but decreased SDC-4 expressions. The effect of LMWF depends on SDC-4 expression. Silencing EXT2 or HPSE leads to an increased expression of SDC-4, providing the evidence that EXT2 and HPSE regulate the SDC-4 expression. Altogether, these data indicate that EXT2, HPSE, and SDC-4 are involved in the proangiogenic effects of LMWF, suggesting that the HS metabolism changes linked to LMWF-induced angiogenesis offer the opportunity for new therapeutic strategies of ischemic diseases.     

Low-Molecular-Weight Fucoidan Induces Endothelial Cell Migration via the PI3K/AKT Pathway and Modulates the Transcription of Genes Involved in Angiogenesis

Low-molecular-weight fucoidan (LMWF) is a sulfated polysaccharide extracted from brown seaweed that presents antithrombotic and pro-angiogenic properties. However, its mechanism of action is not well-characterized. Here, we studied the effects of LMWF on cell signaling and whole genome expression in human umbilical vein endothelial cells and endothelial colony forming cells. We observed that LMWF and vascular endothelial growth factor had synergistic effects on cell signaling, and more interestingly that LMWF by itself, in the absence of other growth factors, was able to trigger the activation of the PI3K/AKT pathway, which plays a crucial role in angiogenesis and vasculogenesis. We also observed that the effects of LMWF on cell migration were PI3K/AKT-dependent and that LMWF modulated the expression of genes involved at different levels of the neovessel formation process, such as cell migration and cytoskeleton organization, cell mobilization and homing. This provides a better understanding of LMWF’s mechanism of action and confirms that it could be an interesting therapeutic approach for vascular repair.     

生肌玉红膏联合封闭负压引流术对糖尿病足患者溃疡创面血管新生及氧化应激指标的影响

目的 探讨生肌玉红膏联合封闭负压引流术（racuum sealing drainage,VSD）对糖尿病足溃疡创面血管新生及氧化应激指标的影响。方法 选取70例糖尿病足溃疡患者随机分为两组,每组35例。所有患者均给予常规清创处理及VSD,观察组加用生肌玉红膏联合负压引流术治疗。观察比较两组患者治疗后创面愈合时间、创面愈合率、踝肱指数变化情况;评价创面血管内皮生长因子（vascular endothelial growth factor,VEGF）、碱性成纤维生长因子（basic fibroblast growth factor,bFGF）、内皮抑素（endostatin,ES）表达情况及超氧化物歧化酶（superoxide dismutase,SOD）、丙二醛（malondialdehyde,MDA）、晚期蛋白氧化产物（advanced oxidation protein products,AOPP）等氧化应激指标。结果 与对照组相比,治疗后观察组患者创面愈合时间缩短,创面愈合率及踝肱指数升高,差异均有统计学意义（P<0.05）;观察组创面VEGF、bFGF表达的平均吸光度值高于对照组（P<0.05）,创面ES的水平明显低于对照组（P<0.05）;观察组SOD水平高于对照组,MDA、AOPP水平低于对照组（P<0.05）。1年随访发现对照组患者溃疡复发率高于观察组（P<0.05）,但两组患者的截肢率及病死率差异无统计学意义（P>0.05）。结论 生肌玉红膏联合VSD可以缩短糖尿病足溃疡创面愈合时间,促进创面的修复,其作用机制可能与调节VEGF、bFGF、ES、SOD、MDA、AOPP等因子表达,促进溃疡创面血管新生,改善机体氧化应激有关。     

Treatment of oral squamous cell carcinoma in a horse by surgical debulking followed by metronomic chemotherapy

A 19‐year‐old Quarter Horse gelding presented with a 6‐week history of hypersalivation, halitosis and dysmasesis. Oral examination revealed retention of food and saliva and the presence of a raised, nodular, 6 × 7 cm, ulcerated mass on the dorsal surface of the tongue base. The mass was confirmed histologically as squamous cell carcinoma. Complete resection of the mass was not possible and surgical laser debulking was followed 15 days later by chemotherapy with a combination of meloxicam and cyclophosphamide in metronomic regimen. After one week, there was a significant improvement in clinical signs and food consumption returned to normal. Therapy was well tolerated with no alteration in haematological or urinalysis parameters. After 5 months of excellent life quality, the horse showed progressive difficulties in mastication and swallowing. Endoscopic examination showed extension of the tumour to all the aboral aspect of the tongue and, with the owner's consent, the patient was subjected to euthanasia. This is believed to be the first report on the combined use of meloxicam and cyclophosphamide in a metronomic fashion for management of an oral squamous cell carcinoma in a horse. Since metronomic therapy is less expensive than conventional chemotherapy, easily administrable and well tolerated, it should be considered as a possible treatment option for nonresectable equine malignant tumours.     

Identification of a Pro-Angiogenic Potential and Cellular Uptake Mechanism of a LMW Highly Sulfated Fraction of Fucoidan from Ascophyllum nodosum

Herein we investigate the structure/function relationships of fucoidans from Ascophyllum nodosum to analyze their pro-angiogenic effect and cellular uptake in native and glycosaminoglycan-free (GAG-free) human endothelial cells (HUVECs). Fucoidans are marine sulfated polysaccharides, which act as glycosaminoglycans mimetics. We hypothesized that the size and sulfation rate of fucoidans influence their ability to induce pro-angiogenic processes independently of GAGs. We collected two fractions of fucoidans, Low and Medium Molecular Weight Fucoidan (LMWF and MMWF, respectively) by size exclusion chromatography and characterized their composition (sulfate, fucose and uronic acid) by colorimetric measurement and Raman and FT-IR spectroscopy. The high affinities of fractionated fucoidans to heparin binding proteins were confirmed by Surface Plasmon Resonance. We evidenced that LMWF has a higher pro-angiogenic (2D-angiogenesis on Matrigel) and pro-migratory (Boyden chamber) potential on HUVECs, compared to MMWF. Interestingly, in a GAG-free HUVECs model, LMWF kept a pro-angiogenic potential. Finally, to evaluate the association of LMWF-induced biological effects and its cellular uptake, we analyzed by confocal microscopy the GAGs involvement in the internalization of a fluorescent LMWF. The fluorescent LMWF was mainly internalized through HUVEC clathrin-dependent endocytosis in which GAGs were partially involved. In conclusion, a better characterization of the relationships between the fucoidan structure and its pro-angiogenic potential in GAG-free endothelial cells was required to identify an adapted fucoidan to enhance vascular repair in ischemia.     

Efficacy of nano-hydroxyapatite prepared by an aqueous solution combustion technique in healing bone defects of goat

The present study was undertaken to evaluate porous hydroxyapatite (HAp), the powder of which was prepared by a novel aqueous solution combustion technique, as a bone substitute in healing bone defects in vivo, as assessed by radiologic and histopathologic methods, oxytetracycline labeling, and angiogenic features in Bengal goat. Bone defects were created in the diaphysis of the radius and either not filled (group I) or filled with a HAp strut (group II). The radiologic study in group II showed the presence of unabsorbed implants which acted as a scaffold for new bone growth across the defect, and the quality of healing of the bone defect was almost indistinguishable from the control group, in which the defect was more or less similar, although the newly formed bony tissue was more organized when HAp was used. Histologic methods showed complete normal ossification with development of Haversian canals and well-defined osteoblasts at the periphery in group II, whereas the control group had moderate fibro-collagenization and an adequate amount of marrow material, fat cells, and blood vessels. An oxytetracycline labeling study showed moderate activity of new bone formation with crossing-over of new bone trabeculae along with the presence of resorption cavities in group II, whereas in the control group, the process of new bone formation was active from both ends and the defect site appeared as a homogenous non-fluoroscent area. Angiograms of the animals in the control group showed uniform angiogenesis in the defect site with establishment of trans-transplant angiogenesis, whereas in group II there was complete trans-transplant shunting of blood vessel communication. Porous HAp ceramic prepared by an aqueous combustion technique promoted bone formation over the defect, confirming their biologic osteoconductive property.     

Serum vascular endothelial growth factor concentrations and postsurgical outcome in dogs with osteosarcoma

Vascular endothelial growth factor (VEGF) is a key angiogenic growth factor, playing putative roles in both tumour growth and metastasis. The purpose of this study was to correlate pretreatment serum concentrations of VEGF in dogs with osteosarcoma (OSA) with disease‐free interval (DFI) and overall survival (OS). Additionally, the effect of serum from dogs with OSA on ex vivo canine endothelial cell (EC) growth was determined. Pretreatment platelet‐corrected serum VEGF levels correlated significantly with DFI. No other examined variable predicted outcome. The ability of sera from dogs with OSA to stimulate canine EC proliferation did not correlate with VEGF concentration or outcome. These data support a role for VEGF in the development or progression of metastatic disease in dogs with OSA. The VEGF concentration in tested sera was not a major determinant of ex vivo canine EC proliferation in this study.