OP-1 injection increases VEGF expression but not angiogenesis in a rabbit model of distraction osteogenesis

We have previously shown that a single injection of rhBMP-7 (OP-1) applied to the regenerate early during distraction accelerates bone consolidation in a rabbit model of distraction osteogenesis. In the present study, we hypothesised that the injection of OP-1 improves bone consolidation by increasing blood flow to the distracted site. Blood flow into the regenerate of a rabbit model was measured and vascular endothelial growth factor (VEGF) expression was tested using semi-quantitative PCR. Immunohistochemistry was used for assessing the temporal and spatial expression of platelet endothelial cell adhesion molecule (PECAM), VEGF and its receptors following OP-1 injection. We observed a higher expression of VEGF and its receptors in the regenerate with OP-1 treatment. However, there was no difference in the increase in bone blood flow nor PECAM expression between the treated and control groups of animals. Interestingly, the increased expression of VEGF and its receptors was associated with chondrocyte and fibroblast-like cells, but not with endothelial cells. These results suggest that accelerated ossification by OP-1 may depend on a non-vascular mechanism, possibly involving a non-angiogenic function of VEGF signalling.     

Early injection of OP-1 during distraction osteogenesis accelerates new bone formation in rabbits

Distraction osteogenesis (DO) is a surgical technique for generating new bone by applying controlled distraction of two bony segments post osteotomy. A limitation of the technique is the long time required for the new bone to consolidate. We investigated the effect of injecting osteogenic protein 1 (OP-1) at the beginning of distraction in a rabbit model of DO. Regenerate bone was evaluated using radiology, densitometry, micro-computed tomography (microCT) and histomorphometry. Immunohistochemsitry was used to evaluate changes in expression of various ligands, growth factors and receptors following OP-1 treatment. Compared to the control, a two-fold increase in bone volume was apparent for treated groups at 3 weeks post injection. An upregulation of almost all of the 41 genes examined was observed. Results suggested that applying OP-1 early during distraction can accelerate bone formation by the activation of numerous pathways. This study provides further insights on strategies to improve bone regeneration rate in DO.     

犬牙槽骨牵张成骨的生物力学和组织学分析

背景：牙槽骨牵张成骨是解决严重牙槽骨萎缩的重要方法,其成骨过程和生物力学对于以后的种植和修复极为重要,目前一直缺少相关的实验研究。 目的：分析犬牙槽骨牵张成骨的生物力学和组织学特点。 方法：先拔除12只杂种犬双侧下颌前磨牙,牙槽骨修整后,制作萎缩牙槽骨模型。3个月后,植入骨内型牙槽骨牵张器。经过7 d的间歇期,以1 mm/d,1次/d的频率进行牙槽骨垂直向增高。在固定期的1,2和3个月,对牵张后的牙槽骨进行临床、生物力学、放射学和组织学检测。 结果与结论：所有牵张器与周围组织愈合良好。牵张结束时,临床和放射学检查显示：萎缩牙槽骨分别增高 (4.80±0.50) mm和(5.12±0.67) mm。组织学检测发现牵张区骨小梁在固定期的1-3个月成熟,其剪切力逐步提高,固定期3个月时和自体骨的剪切力相当。结果显示牙槽骨牵张成骨的组织学和生物力学性能在固定期3个月时与自体骨相当。 中国组织工程研究杂志出版内容重点：组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程全文链接：     

富血小板纤维蛋白对兔下颌骨牵引成骨区骨形态发生蛋白-4表达的影响

目的:探讨富血小板纤维蛋白(PRF)对牵引成骨区骨形态发生蛋白-4(BMP-4)表达的影响。方法:25只大耳白兔随机分2组,分别行双侧下颌骨皮质骨切开术,一侧下颌骨牵引间隙放置PRF膜作为实验组,对侧作为对照组,分别于稳定期第3、7、14、21、28天处死一组动物,切取牵引间隙处骨痂行H-E染色和BMP-4免疫组织化学显色,细胞图像分析仪测量牵引间隙处骨痂BMP-4表达情况。结果:下颌牵引延长后牵引间隙均有新骨形成,免疫组织化学显色显示BMP-4主要定位于成骨细胞的细胞质中。实验组在稳定期3、7、14、21 d BMP-4表达的阳性细胞率和阳性面积百分比均显著高于对照组,稳定期28 d BMP-4表达的阳性细胞率和阳性面积百分比与对照组比较差异均无统计学意义。结论:PRF能促进兔下颌骨牵引成骨区新骨的生成,BMP-4可能在牵引成骨过程中调控组织细胞应力信号传递,发挥成骨作用。     

Strategies of enhancing bone regenerate formation in distraction osteogenesis

Distraction osteogenesis (DO) is a commonly used technique in multiple orthopedic sub-specialties, including trauma, oncology and pediatrics. This technique aims to produce new bone formation in the distraction gap in a controlled manner. The issue with this technique has been the high risk of complications, one of which is poor regenerate formation during the distraction process. Although several factors (including patient and operative factors) and techniques (including surgical, mechanical and pharmacological) have been described to ensure successful regenerate formation during the process of DO, these factors are sometimes difficult to control clinically. Our aim from this review is to highlight the different factors that affect DO, modalities to assess the regenerate and review treatment options for poor regenerate in the distraction gap. In addition, we propose a management protocol derived from the available literature that can be used to facilitate the management of inadequate regenerate formation.     

牵张延长下颌骨对二腹肌纤维型构成影响的研究

目的 :研究牵张延长下颌骨前后二腹肌的组织化学特征。方法 :应用肌球蛋白ATP酶 (pH9.4)染色法 ,观测了牵张前后 8只狗的二腹肌肌纤维类型 ,并用VIDAS图像分析仪测量其横截面积。结果 :ATP酶染色 ,二腹肌肌纤维分为Ⅰ型和Ⅱ型 ,Ⅰ型纤维数量较Ⅱ型纤维少。牵张延长下颌骨后 ,Ⅰ型纤维数量显著性增多 ,Ⅱ型纤维数量显著性减少。牵张成骨后 ,Ⅰ型纤维、、Ⅱ型纤维的横截面积与术前比较没有显著性差异。结论 :牵张延长下颌骨后 ,与牵张方向平行的二腹肌肌纤维型分布发生了与功能变化相适应的改变     

Early injection of OP-1 during distraction osteogenesis accelerates new bone formation in rabbits

Distraction osteogenesis (DO) is a surgical technique for generating new bone by applying controlled distraction of two bony segments post osteotomy. A limitation of the technique is the long time required for the new bone to consolidate. We investigated the effect of injecting osteogenic protein 1 (OP-1) at the beginning of distraction in a rabbit model of DO. Regenerate bone was evaluated using radiology, densitometry, micro-computed tomography (microCT) and histomorphometry. Immunohistochemsitry was used to evaluate changes in expression of various ligands, growth factors and receptors following OP-1 treatment. Compared to the control, a two-fold increase in bone volume was apparent for treated groups at 3 weeks post injection. An upregulation of almost all of the 41 genes examined was observed. Results suggested that applying OP-1 early during distraction can accelerate bone formation by the activation of numerous pathways. This study provides further insights on strategies to improve bone regeneration rate in DO.     

Differential brain,but not serum VEGF levels in a genetic rat model of depression

Compared to the classical monoamine hypotheses focus on neuroplasticity is a major new approach in studies of depression and antidepressants. Recent studies have demonstrated that vascular endothelial growth factor (VEGF) is regulated by antidepressant treatment in rodents. However, in depressive patients no significant changes were found in the serum VEGF levels compared to control subjects. To our knowledge, brain and serum VEGF levels have never been reported in parallel for any psychiatric disease model. That prompted us to examine the levels of VEGF in serum, hippocampus, frontal cortex, corpus striatum, and hypothalamus in male Flinders Sensitive Line (FSL) and Flinders Resistant Line (FRL), a genetic rat model of depression. The VEGF levels were identical in the FSL and the FRL rats in serum, corpus striatum, and hypothalamus. In hippocampus and frontal cortex, the VEGF levels were significantly decreased in the FSL rats compared to the FRL rats. The results may add to the hypothesis that altered expression of growth factors/neurotrophic factors are related to the pathophysiology of depression.     

Osteogenic response of human adipose-derived stem cells to BMP-6, VEGF,and combined VEGF plus BMP-6 in vitro

Exogenous addition of three factors—mesenchymal stem cells (MSCs), vascular endothelial growth factor (VEGF), and bone morphogenetic proteins (BMPs)—has proven to be more beneficial than delivery of any single factor for fracture repair in animal models. We studied the osteogenic differentiation of human adipose-derived stem cells (hADSCs) in the presence of VEGF, BMP-6, or VEGF plus BMP-6 to better understand their enhancement of osteoblastic differentiation of MSCs. The VEGF plus BMP-6 group demonstrated an additive effect on the enhancement of mineralization and expression of ALP and Msx2 genes. Unlike VEGF or BMP-6 alone, the combination of VEGF and BMP-6 significantly enhanced the expression of COL1A1, osterix, and Dlx5 genes. The data indicate that a cross-talk between VEGF and BMP-6 signaling pathways enhances osteogenic differentiation of hADSCs.     

AG490对HEL细胞VEGF和HIF-1α表达的影响

 目的: 探讨JAK2抑制剂AG490对人红白血病(HEL)细胞迁移及对VEGF和HIF-1α表达的影响。方法: 用不同浓度的AG490处理HEL细胞,CCK-8法检测细胞活力,Hoechst 33342荧光染色检测细胞凋亡,流式细胞术检测细胞凋亡及周期,Transwell小室检测细胞迁移能力,RT-PCR检测JAK2的mRNA水平,Western blot检测p-JAK2、VEGF和HIF-1α的蛋白水平。结果: AG490能够抑制HEL细胞的活力,不同浓度(20、40、60、80和100 μmol/L)AG490作用HEL细胞48 h后,细胞活力分别为88%、75%、48%、10%和0.12%(P<0.05);Hoechst 33342凋亡细胞染色显示80 μmol/L AG490处理细胞48 h后,亮蓝色凋亡细胞较对照组明显增多(P<0.05);流式结果显示80 μmol/L AG490作用细胞48 h后,凋亡率上升;细胞迁移实验结果显示20 μmol/L AG490处理细胞24 h后漏出细胞明显低于对照组(P<0.05);RT-PCR结果显示不同浓度AG490处理HEL细胞48 h后JAK2 mRNA呈剂量依赖性减低;Western blot结果显示实验组细胞的p-JAK2、VEGF和HIF-1α蛋白水平较对照组明显减低(P<0.05)。结论: AG490可通过抑制JAK2信号通路,抑制HEL细胞血管新生因子VEGF和HIF-1α的表达。     

杜仲醇提取物对兔下颌牵张成骨的影响

背景：目前牵张成骨由于治疗周期长、并发症较多成为临床广泛运用的瓶颈,不能满足临床推广需要。 目的：以兔下颌骨牵张动物模型为实验平台,观察全身运用杜仲醇提取物对牵张新骨再生的影响。 方法：24只新西兰大白兔随机均分为对照组及实验组,建立兔下颌单侧牵张模型,牵张速率为每12 h 1 mm。在牵张期2次/d,实验组及对照组分别予以灌胃杜仲醇提取物及等量生理盐水,牵张结束后6周处死动物收集标本行成骨检测。 结果与结论：两组动物牵张间隙内均可观察到新骨生成。牵张成骨结束后6周下颌骨CT图像显示实验组兔牵张间隙舌侧骨皮质生成良好,颊侧骨皮质连续,牵张间隙可见均匀骨质生成桥接;下颌骨核素扫描显示实验组兔下颌骨牵张间隙表现为核浓集,强度明显强于对照组;X射线显示实验组牵张间隙完全桥接,新生成骨质密度均匀,可见上下侧骨皮质形成良好;Micro-CT图像显示实验组骨皮质部分形成较好,骨小梁分布较为均匀,骨小梁明显较对照组粗壮,对照组部分区域仍有囊性变;Micro-CT微结构参数检测显示实验组骨体积分数、骨小梁厚度、骨小梁数量和连接密度均显著高于对照组;组织学观察显示与对照组比较,实验组牵张间隙中有较为成熟的束状骨,骨小梁方向较为规律。实验结果显示全身运用杜仲醇提取物可有效促进兔下颌快速骨牵张的新骨再生。     

LHT7, a chemically modified heparin,inhibits multiple stages of angiogenesis by blocking VEGF,FGF2 and PDGF-B signaling pathways

Despite the therapeutic benefits of the angiogenesis inhibitors shown in the clinics, they have encountered an unexpected limitation by the occurrence of acquired resistance. Although the mechanism of the resistance is not clear so far, the upregulation of alternative angiogenic pathways and stabilization of endothelium by mural cells were reported to be responsible. Therefore, blocking multiple angiogenic pathways that are crucial in tumor angiogenesis has been highlighted to overcome such limitations. To develop an angiogenesis inhibitor that could block multiple angiogenic factors, heparin is an excellent lead compound since wide array of angiogenic factors are heparin-binding proteins. In previous study, we reported a heparin-derived angiogenesis inhibitor, LHT7, as a potent angiogenesis inhibitor and showed that it blocked VEGF signaling pathway. Here we show that LHT7 could block the fibroblast growth factor 2 (FGF2) and platelet-derived growth factor B (PDGF-B) in addition to VEGF. Simultaneous blockade of these angiogenic factors resulted in inhibition of multiple stages of the angiogenic process, including initial angiogenic response to maturation of the endothelium by pericyte coverage in vitro. In addition, the treatment of LHT7 in vivo did not show any sign of vascular normalization and directly led to decreased blood perfusion throughout the tumor. Our findings show that LHT7 could effectively inhibit tumor angiogenesis by blocking multiple stages of the angiogenesis, and could potentially be used to overcome the resistance.     

The expression of receptor for advanced glycation end products is associated with angiogenesis in human oral squamous cell carcinoma

The receptor for advanced glycation end products (RAGE) is associated with cancer progression in several human cancers. In this study, we examined the roles of RAGE in the angiogenesis of oral squamous cell carcinoma (OSCC). RAGE concentration was examined in 20 OSCC tumors by enzyme-linked immunosorbent assay (ELISA). The microvessel density (MVD) and lymph vessel density (LVD) were examined by immunostaining. Concentrations of vascular endothelial growth factor (VEGF) and VEGF-C were examined in tumor tissues by ELISA. Tumoral RAGE concentration was associated with higher tumor MVD (P = 0.0123) and tumor VEGF concentration (P = 0.0344), but not with LVD and VEGF-C concentration. Treatment with RAGE ligand, high-mobility group box (HMGB)-1 increased the secretion of VEGF but not that of VEGF-C in human OSCC cell lines, HSC-3 and HSC-4. The effect of HMGB-1 was abrogated by RAGE down-regulation by antisense S-oligodeoxynucleic acid. These results suggest that RAGE expression is closely associated to angiogenesis in OSCC.     

树鼩脑缺血后适应升高海马区rCBF及VEGF的变化

目的 探讨缺血后适应(PC)缓解海马rCBF与血管内皮生长因子(VEGF)的变化及其机制.方法 建立树鼩血栓性局部脑缺血模型,通过激光多普勒血流计测量海马CA1区rCBF含量;用免疫组化法测定海马VEGF的表达.结果 树鼩脑缺血时海马rCBF逐渐降低,以24 h的改变最显著,脑缺血后海马CA1区VEGF阳性细胞数增多,12 h表达最强(P<0.01);缺血PC可显著影响缺血所致的改变:rCBF逐渐增加,72 h最显著(P<0.01),与此同时VEGF的表达除8 h外均比血栓性缺血组增强(P<0.01),12 h组最明显;电镜显示缺血24 h血栓性缺血组的海马线粒体应激及内质网池形成最明显,给予PC后得以缓解.结论 缺血12 h内PC通过明显增强VEGF的表达可能与其改善rCBF有关,从而延长治疗的时间窗.     

Hypoxia-inducible factor-1alpha expression in experimental cirrhosis: correlation with vascular endothelial growth factor expression and angiogenesis

Angiogenesis progresses together with fibrogenesis during chronic liver injury. Hypoxia-inducible factor-1alpha (HIF-1alpha), a master regulator of homeostasis, plays a pivotal role in hypoxia-induced angiogenesis through its regulation of vascular endothelial growth factor (VEGF). The association between hypoxia, angiogenesis and VEGF expression has been demonstrated in experimental cirrhosis. However, expression of HIF-1alpha has yet to be reported. The aim of this study was to investigate the significance of HIF-1alpha expression during experimental liver fibrosis and the relationships between HIF-1alpha expression, VEGF expression and angiogenesis. Cirrhosis was induced in male Wistar rats by intraperitoneal administration of diethyl nitrosamine (DEN) (100 mg/kg, once a week). The serial sections from liver tissues were stained with anti-HIF-1alpha, anti-VEGF and anti-CD34 antibodies before being measured by light microscopy. Our results showed that HIF-1alpha expression gradually increases according to the severity of fibrosis (p<0.01). Moreover, its expression was found to be correlated with angiogenesis (r=0.916) and VEGF expression (r=0.969). The present study demonstrates that HIF-1alpha might have a role in the development of angiogenesis via regulation of VEGF during experimental liver fibrogenesis and suggests that this factor could be a potential target in the manipulation of angiogenesis in chronic inflammatory diseases of the liver.     

VEGF与MMP-2在食管鳞癌中的表达及其临床病理意义

目的研究血管内皮生长因子（VEGF）和基质金属蛋白酶-2（MMP-2）在食管鳞癌中的表达与其临床意义。方法用免疫组化染色的方法检测58例食管鳞癌标本及30例取非瘤正常食管组织标本中VEGF与MMP-2蛋白的表达。结果食管鳞癌中VEGF与MMP-2的表达显著高于正常组织，其表达与食管癌的组织侵润深度、淋巴结转移密切相关，两者呈正相关。结论VEGF和MMP-2在食管鳞癌中高表达，参与了食管鳞癌浸润及转移，可以作为判断食管鳞癌生物学行为的指标之一。     

COX-2、VEGF在乳腺癌组织中的表达及临床意义

目的探讨血管内皮生长因子(VEGF)和环氧化酶-2(COX-2)在乳腺癌组织中的表达及其与临床病理特征之间的关系。方法应用免疫组化S-P法检测不同乳腺组织中VEGF和COX-2的表达情况。结果VEGF、COX-2在乳腺导管癌组织中的表达明显上调,与其在正常乳腺组织、纤维腺瘤组织中的表达相比差异显著(P<0.01)。VEGF、COX-2在乳腺浸润性导管癌中的阳性表达率高于其在乳腺导管癌中的阳性表达率。两组中COX-2的表达有显著性差异(77.1%vs40%,P<0.05),VEGF的表达无显著性差异(80.0%vs70%,P>0.05)。VEGF在乳腺浸润性导管癌的阳性表达与其临床分期、淋巴结转移密切相关(P<0.05,P<0.01),与肿瘤大小无关(P>0.05)。COX-2在乳腺浸润性导管癌中的阳性表达与其临床分期、淋巴结转移及肿瘤大小均无关(P>0.05)。VEGF、COX-2在乳腺导管癌中的表达呈正相关(r=0.98,P<0.05)。结论VEGF、COX-2的高表达在乳腺癌的发生发展过程中起重要的作用,可作为重要的生物学标志。     

Comparative bone healing near eroding polylactide-polyglycolide implants of differing crystallinity in rabbit tibial bone chambers

Eroding poly(DL-lactide-co-glycolide) (PDLLG) washers and poly(L-lactide-co-glycolide) (PLLG) threads were observed chronically in vivo following loading in a bone chamber tibial implant (BCI). Images were recorded using intravital microscopy of the implanted rabbit. Erosion and bone healing, as represented by angiogenesis and osteogenesis, was determined from changes in projected area of observed polymer, vessels and bone, respectively. Erosion rates of the two polymers were significantly different. Healing adjacent to both polymers differed significantly from controls. Healing response to each polymer was also different, with the faster eroding PDLLG causing more deviation from normal osteogenesis and angiogenesis than did PLLG. It was speculated that the faster eroding polymer released macrophage-stimulating fragments earlier in the healing process, thus altering the normal macrophage-endothelial cell interaction which in turn affected angiogenesis-linked components of osteogenesis.     

Significance of survivin, caspase-3, and VEGF expression in thyroid carcinoma

To investigate the clinical significance of survivin, caspase-3, and vascular endothelial growth factor expression in a subset of thyroid carcinoma and their correlation with prognosis. Sixty-eight cases of thyroid carcinoma (TC), 12 cases of thyroid adenoma (TA) and 10 cases of normal thyroid tissue (NT) were involved in immunohistochemical and real-time RT-PCR analyses for survivin, caspase-3, and VEGF expression. Statistical analyses were performed for differential expression among NT, TA and TC, correlations of their expression with the clinicopathological parameters of TC including histological typing, clinical staging and lymphnode metastasis, and relationship of survivin with caspase-3 or VEGF in TC. We observed higher mRNA expression and positive immunostaining for survivin and VEGF in TC compared with TA and NT, with a significant positive correlation among them and significant correlations with histological typing, clinical staging and lymphnode metastasis in TC, but we could not find any significance of caspase-3 in TC and its significant relationship with survivin expression. Our results indicate that survivin and VEGF are unfavorable molecules for TC evolution and prognosis, and possess positive correlation in TC.