促进小鼠骨髓间充质干细胞增殖和成骨分化的血管内皮生长因子

背景：血管内皮生长因子是骨髓间充质干细胞所处骨髓微环境中的重要调控因子,其可促进骨髓间充质干细胞的血管内皮方向分化,但尚无其对骨髓间充质干细胞增殖和成骨分化调控作用的报道。
　　目的：探讨血管内皮生长因子对骨髓间充质干细胞增殖和成骨分化的调控作用及其机制。
　　方法：分离、培养小鼠骨髓间充质干细胞,CCK8方法检测不同质量浓度血管内皮生长因子重组蛋白对骨髓间充质干细胞增殖的影响,选定适宜血管内皮生长因子重组蛋白质量浓度并检测其对骨髓间充质干细胞成骨分化的影响,分子生物学方法检测血管内皮生长因子干预下骨髓间充质干细胞中Osterix,Runx2,碱性磷酸酶、骨钙素和血红素氧化酶1的表达。
　　结果与结论：①血管内皮生长因子可以促进骨髓间充质干细胞增殖,且具有浓度依赖性,100μg/L血管内皮生长因子重组蛋白的促增殖作用最显著。②成骨诱导剂存在条件下,血管内皮生长因子促进骨髓间充质干细胞成骨标志基因Osterix、Runx2、碱性磷酸酶和骨钙素的表达;血管内皮生长因子干预组骨髓间充质干细胞成骨分化的钙结节数量较对照组增加。③血管内皮生长因子可诱导骨髓间充质干细胞中血红素氧化酶1 mRNA和蛋白水平的高表达。以上结果表明血管内皮生长因子促进骨髓间充质干细胞增殖和向成骨细胞分化的调控机制可能与骨髓间充质干细胞内血红素氧化酶1表达增加有关。     

血管内皮生长因子对人骨髓和脐带间充质干细胞中内皮细胞组分的影响

本研究旨在比较人骨髓间充质干细胞(BM-MSC)和脐带间充质干细胞(UC-MSC)中的内皮细胞(EC)比例,并探讨血管内皮生长因子(VEGF)对MSC中EC比例的影响。利用流式细胞术检测EC标志CD34+CD133+和vWF+CD31+双阳性细胞的比例,用瑞氏染色观察经VEGF作用的MSC细胞形态变化,免疫荧光技术观察CD34、CD133、CD31、VWF的表达,最后采用实时荧光定量PCR检测VEGF对EC标志性基因表达的影响。结果表明:BM-MSC和UC-MSC中存在少量EC及内皮祖细胞(EPC),经VEGF 10 ng/ml作用24 h后,MSC的长宽比变大,EC比例上升而EPC比例下降,EC相关标志基因Tie-2和ecNOS等的表达均上调,其中UC-MSC对VEGF的反应更明显。结论:BM-MSC和UC-MSC中存在少量的EC及EPC,VEGF可以提高EC的比例和细胞功能。     

VEGF与肿瘤的相关性及其对间充质干细胞分化为血管内皮细胞诱导作用的研究

目的研究血管内皮生长因子（VEGF）在肿瘤患者的表达水平及与愈后的相关性，并探讨VEGF对间充质干细胞（MSCs）分化为血管内皮细胞的诱导作用。方法用ELISA法检测初发及愈后出院的肿瘤患者血清VEGF含量；分离骨髓单个核细胞，经贴壁法获取MSCs，体外经高浓度VEGF诱导分化，通过流式分析表面分子CD34、CD31和vWF以及体外成血管实验对分化细胞进行鉴定。结果初发患者与正常对照相比高水平表达VEGF（P〈0．01），而愈后明显降低（P〈0．01）。MSCs经VEGF诱导后呈现“纤维细胞”样长梭形，内皮系表面分子CD34、CD31、vWF均有不同程度表达，在半固体培养基中可形成“血管管腔样”结构。结论VEGF与肿瘤的发生及其愈后有明显的相关性；在体外培养条件下可诱导MSCs分化为血管内皮细胞。     

骨髓间充质干细胞来源的成骨细胞与血管内皮生长因子基因整合的研究

目的 大鼠骨髓间充质干细胞(MSCs)在药物诱导下定向分化为成骨细胞,对其进行血管内皮生长因子(VEGF)基因片段转染,观察其生物学特性的变化.方法 选择MSCs体外培养的第2代细胞诱导其向成骨细胞分化,实验分为对照组和地塞米松浓度1×10-8mol/L诱导组.对诱导分化后的成骨细胞进行碱性磷酸酶(ALP)染色和活性测定及Ⅰ型胶原的免疫组化染色、茜素红钙节结染色的鉴定.对诱导后的成骨细胞进行已构建的VEGF基因片段(ad5-h-VEGF)的人5型腺病毒载体进行转染,并进行免疫组化鉴定.结果 诱导培养第3、6、9、12天的ALP活性(A405)测定,地塞米松浓度1×10-8mol/L组测定值(A值)分别为0.1 44±0.004、0.175±0.003、0.207±0.010、0.224±0.004;对照组分别为0.101±0.003、0.124±0.016、0.133±0.005、0.139±0.005.两组在组间、不同时点以及组间和不同时点的交互作用差异均有统计学意义(P＜0.05).MSCs经地塞米松的成骨诱导剂诱导7～9天可出现碱性磷酸酶染色阳性,连续培养20天后可出现茜素红钙节结染色阳性.转染后细胞VEGF免疫组化染色阳性.结论 对骨髓间充质干细胞来源的成骨细胞作ad5-h-VEGF基因转染,可使诱导的成骨细胞胞浆中VEGF阳性表达.     

β2-glycoprotein I inhibits vascular endothelial growth factor and basic fibroblast growth factor induced angiogenesis through its amino terminal domain

Summary.  Background:  Beta-2 glycoprotein I (β₂GPI) is a plasma glycoprotein which interacts with various proteins of the coagulation and fibrinolysis system. β₂GPI has recently been shown to have anti-angiogenic properties. Objectives:  We undertook this study to investigate the specific domain of β₂GPI involved in the anti-angiogenic function and its effect on downstream signaling of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF). Methods:  Various preparations of β₂GPI were used on human umbilical vein endothelial cells (HUVECs) in the absence or presence of VEGF and bFGF. The effect on HUVECs' proliferation, migration and tubule formation in Matrigel matrix was investigated. The effect of β₂GPI on the mRNA expression of VEGF receptors and phosphorylation of signaling molecules was also studied. Results:  β₂GPI is shown in this study to be an anti-angiogenic molecule in vitro by inhibiting VEGF and bFGF-induced proliferation, migration and papillary-like tubule formation of HUVECs. This inhibition was achieved by native, proteolytically clipped and domain deletion mutants, domain I-IV (DI-IV) but not domain II-V (DII-V) of β₂GPI. Native β₂GPI was found to downregulate the expression of the VEGF receptor KDR/Flk-1 on endothelial cells and to block the phosphorylation of VEGF's downstream effector molecules in the MAPK/ERK and PI3K/Akt/GSK3β pathways. Conclusions:  These results indicate that β₂GPI has anti-angiogenic functions which depend on the presence of domain I. This anti-angiogenic activity may have important implications for the therapeutic manipulation of angiogenesis in various disease states.     

低氧环境下骨髓间充质干细胞中血管内皮生长因子的表达

背景：作为种子细胞来源的骨髓间充质干细胞移植后能否在相对缺氧的体内环境下生存,是移植成功与否的重要环节。因此研究体外低氧环境下骨髓间充质干细胞的生长状态,可以为体内细胞移植提供实验依据。目的：观察体外低氧环境对大鼠骨髓间充质干细胞生长及血管内皮生长因子表达的影响。方法：分离培养大鼠骨髓间充质干细胞,光镜观察细胞的形态;取第3代骨髓间充质干细胞,按氧浓度分为两组,分别在常氧条件下（体积分数为21%氧气）和低氧条件下（体积分数为3%氧气）培养72 h。用CCK-8法和流式细胞术检测两组细胞的增殖情况,Western-blot印迹法检测常氧组和低氧组细胞中缺氧诱导因子1α与血管内皮生长因子蛋白的表达。结果与结论：①成功分离和培养了骨髓间充质干细胞,光镜下细胞呈长梭形,形态均一。②CCK-8法结果显示各时间点低氧组的细胞数目均高于常氧组,在36,48 h时活力增加显著（P〈0.05）。③流式细胞术结果显示,与常氧组比较,低氧组处于S期的细胞数量增加,且细胞增殖指数也显著增加（P〈0.05）。④Western-blot印迹法结果显示,常氧组细胞缺氧诱导因子1α及血管内皮生长因子蛋白仅有少量的表达,而低氧组两蛋白表达量均呈时间依赖性上调（P〈0.05）。以上结果说明低氧可诱导体外培养的大鼠骨髓间充质干细胞的增殖,又上调了缺氧诱导因子1α与血管内皮生长因子蛋白的表达,随着低氧时间的延长呈升高趋势。     

Notch信号和血管内皮生长因子基因对大鼠骨髓间充质干细胞诱导分化后内皮细胞功能的影响

目的 探讨Notch信号和血管内皮生长因子(VEGF165)基因对大鼠骨髓间充质干细胞(MSCs)诱导分化后内皮细胞功能的影响.方法 分离、培养大鼠MSCs,用含VEGF165和碱性成纤维细胞生长因子(bFGF)的细胞培养液培养大鼠MSCs 2周诱导其向内皮细胞分化;用脂质体将携带有VEGF165基因的质粒转染诱导内皮细胞并对转染效果进行鉴定,逆转录-聚合酶链反应(RT-PCR)检测转染前后细胞上Notch信号受体Notch 1和配体Jagged 1的表达变化;用γ-内分泌酶抑制剂L-685458阻断细胞Notch信号通路的转导,划痕实验检测细胞迁移能力;将细胞接种在半固体培养基上,观察其形成毛细血管样结构的能力.结果 转染VEGF165基因的内皮细胞上表达有VEGF165 mRNA,说明实验成功地将VEGF165基因导入诱导后内皮细胞中.转染VEGF165基因后,细胞上Notch信号配体Jagged1 mRNA表达增强(1.08±0.01比1.01±0.02,P＜0.01),Notch1 mRNA表达无明显变化(0.60±0.02比0.59±0.01,P＞0.05);细胞的迁移能力增强(划痕空白处细胞个数:46.45±4.46比41.61±1.42,P＜0.05),形成毛细血管样结构能力无明显变化(细胞分级:3.00±0.89比2.00±0.89,P＞0.05).内皮细胞转染VEGF165基因后,以γ-内分泌酶抑制剂L-685458阻断细胞Notch信号通路的转导,则细胞迁移能力(划痕空白处细胞个数:51.72±3.47比46.45±4.46)和形成毛细血管样结构能力(细胞分级:4.17±0.75比3.00±0.89)均进一步增强(均P＜0.05).结论 转染VEGF165基因可增强大鼠MSCs诱导分化内皮细胞的功能,在此基础上阻断Notch信号通路转导可进一步增强细胞功能.     

神经生长因子可促进骨折愈合过程中血管内皮生长因子的表达

背景：骨折愈合过程十分复杂,需要多种细胞因子的参与.目前研究较多的细胞因子有骨形态发生蛋白、成纤维细胞生长因子、转化生长因子β、血管内皮细胞生长因子和胰岛素样生长因子,但神经生长因子在骨折愈合过程中对血管内皮细胞生长因子的作用尚不明确.目的：观察神经生长因子对兔骨折愈合中血管内皮生长因子表达的影响.方法：实验建立标准兔桡骨骨折模型,分别用神经生长因子、神经生长因子单克隆抗体和生理盐水进行干预,即应用神经生长因子组、拮抗神经生长因子组和对照组.结果与结论：损伤后24,48 h和损伤后1,3,6,8周Western blot检测骨折端组织血管内皮生长因子蛋白的表达分析结果显示,3组各时间点血管内皮生长因子表达的关系为：应用神经生长因子组＞对照组＞拮抗神经生长因子组（P＜0.05）.结果证实,在骨折愈合过程当中,应用神经生长因子可以促进血管内皮生长因子表达.     

Controlled release of vascular endothelial growth factor from spray‐dried alginate microparticles in collagen–hydroxyapatite scaffolds for promoting vascularization and bone repair

A major limitation with current tissue‐engineering approaches is creating functionally vascularized constructs that can successfully integrate with the host; this often leads to implant failure, due to avascular necrosis. In order to overcome this, the objective of the present work was to develop a method to incorporate growth factor‐eluting alginate microparticles (MPs) into freeze‐dried, collagen‐based scaffolds. A collagen–hydroxyapatite (CHA) scaffold, previously optimized for bone regeneration, was functionalized for the sustained delivery of an angiogenic growth factor, vascular endothelial growth factor (VEGF), with the aim of facilitating angiogenesis and enhancing bone regeneration. VEGF was initially encapsulated in alginate MPs by spray‐drying, producing particles of < 10 µm in diameter. This process was found to effectively encapsulate and control VEGF release while maintaining its stability and bioactivity post‐processing. These VEGF‐MPs were then incorporated into CHA scaffolds, leading to homogeneous distribution throughout the interconnected scaffold pore structure. The scaffolds were capable of sustained release of bioactive VEGF for up to 35 days, which was proficient at increasing tubule formation by endothelial cells in vitro. When implanted in vivo in a rat calvarial defect model, this scaffold enhanced vessel formation, resulting in increased bone regeneration compared to empty‐defect and VEGF‐free scaffolds. This biologically functionalized scaffold, composed entirely of natural‐based materials, may offer an ideal platform to promote angiogenesis and tissue regeneration. Copyright © 2015 John Wiley & Sons, Ltd.     

重度子痫前期患者血管内皮生长因子及其受体-1水平变化及意义

目的检测早发型和晚发型重度子痫前期患者血清和胎盘中血管内皮生长因子（vascular endothelial growth factor,VEGF）及血管内皮生长因子受体-1（soluble vascular endothelial growth factor receptors-1,sFlt-1）水平,探讨其在重度子痫前期发病中的作用机制。方法 20例早发型重度子痫前期患者为早发重度组,20例晚发型重度子痫前期患者为晚发重度组,20例正常晚期妊娠者为正常妊娠组,采用ELISA法检测3组血清VEGF及sFlt-1水平,同时应用免疫组织化学SP法检测胎盘组织中VEGF及sFlt-1的阳性表达率。结果血清VEGF水平及胎盘VEGF阳性表达率早发重度组（（27.5±3.6）ng/L和15%）、晚发重度组（（36.2±3.2）ng/L和30%）均低于正常妊娠组（（45.5±3.7）ng/L和45%）（P〈0.05）,早发重度组低于晚发重度组（P〈0.05）;血清sFlt-1水平及胎盘sFlt-1阳性表达率早发重度组（（42.2±3.2）ng/L和60%）、晚发重度组（（30.2±2.5）ng/L和45%）均高于正常妊娠组（（14.7±2.7）ng/L和10%）（P〈0.05）,早发重度组高于晚发重度组（P〈0.05）。结论 VEGF水平低表达及sFlt-1水平高表达可能与重度子痫前期的严重程度相关,在重度子痫前期发生、发展中起重要作用。     

Vascular endothelial growth factor‐transfected adipose‐derived stromal cells enhance bone regeneration and neovascularization from bone marrow stromal cells

Vascular endothelial growth factor (VEGF)‐transfected adipose‐derived stromal cells (ADSCs^VEGF) were devised to promote bone regeneration and neovascularization of bone marrow stromal cells (BMSCs). ADSCs^VEGF were added to BMSCs and cocultured in variable proportions. ADSCs^VEGF alone or ADSCs^VEGF with BMSCs (BMSCs:ADSCs^VEGF ratio of 1:0.025–0.5) induced significantly greater tube formation in human umbilical vein endothelial cells than untransfected ADSCs. The cocultures of BMSCs and ADSCs^VEGF at ratios of 1: 0.025–0.1 showed significantly greater osteogenic differentiation and mineralization than BMSCs alone in vitro. Osteogenic markers COL1A1, OCN and BSP were most effectively induced at the BMSC: ADSC^VEGF ratio of 1:0.05. Of angiogenesis‐related genes, upregulation of cathepsin Z and downregulation of early growth response 1 were observed while two osteogenesis‐related genes, osteoactivin and tetranectin, were upregulated in BMSCs/ADSCs^VEGF compared to BMSCs/ADSCs. When critical size calvarial defects in rats were implanted with mixture of BMSCs and ADSCs^VEGF along with hydroxyapatite/β‐tricalcium phosphate granules, BMSCs and ADSCs^VEGF at the ratio of 1:0.05 showed better bone regeneration that BMSCs alone. The cotransplantation of ADSCs^VEGF with BMSCs significantly increased neovascularization on the regenerated bone of the repaired defect than BMSCs alone. In conclusion, ADSCs^VEGF added in small proportion to BMSCs effectively promote bone regeneration and neovascularization. Copyright © 2017 John Wiley & Sons, Ltd.     

磷酸钙骨水泥复合骨形态发生蛋白 6 和血管内皮生长因子修复骨缺损简

背景：单独将骨形态发生蛋白或血管内皮生长因子植入体内易被血液冲刷掉而不能最大限度发挥诱导成骨和血管生成作用,同时缺少载体的支撑作用。 目的：观察骨形态发生蛋白6、血管内皮生长因子及磷酸钙骨水泥联合应用在骨缺损修复过程中的作用。 方法：制作新西兰兔双侧股骨内侧髁骨缺损模型,左侧分别植入磷酸钙骨水泥/骨形态发生蛋白6/血管内皮生长因子、磷酸钙骨水泥/骨形态发生蛋白6及磷酸钙骨水泥,右侧不植入任何物质作为空白对照。植入8,16周通过硬组织切片组织学观察、电镜扫描等手段观察新骨形成情况。 结果与结论：各组材料的组织相容性良好,未见明显炎症组织反应。植入8周时,磷酸钙骨水泥/骨形态发生蛋白6/血管内皮生长因子组骨水泥-骨组织交界处基本上被新生骨小梁包绕,材料进一步降解,新生骨小梁表面可见大量活跃的成骨细胞;16周时,新生骨小梁继续长入,进一步增长、增粗、增多,有大量新生编织骨成网格状长入材料中,骨水泥材料降解明显,与周围组织结合紧密,降解与骨长入同步,此组不同时间点成骨速度及成骨效果均明显优于其他两组材料（P 〈0.05）。表明3种材料联合应用可协同促进骨缺损修复。     

Differential bone‐forming capacity of osteogenic cells from either embryonic stem cells or bone marrow‐derived mesenchymal stem cells

For more than a decade, human mesenchymal stem cells (hMSCs) have been used in bone tissue‐engineering research. More recently some of the focus in this field has shifted towards the use of embryonic stem cells. While it is well known that hMSCs are able to form bone when implanted subcutaneously in immune‐deficient mice, the osteogenic potential of embryonic stem cells has been mainly assessed in vitro. Therefore, we performed a series of studies to compare the in vitro and in vivo osteogenic capacities of human and mouse embryonic stem cells to those of hMSCs. Embryonic and mesenchymal stem cells showed all characteristic signs of osteogenic differentiation in vitro when cultured in osteogenic medium, including the deposition of a mineralized matrix and expression of genes involved in osteogenic differentiation. As such, based on the in vitro results, osteogenic ES cells could not be discriminated from osteogenic hMSCs. Nevertheless, although osteogenic hMSCs formed bone upon implantation, osteogenic cells derived from both human and mouse embryonic stem cells did not form functional bone, indicated by absence of osteocytes, bone marrow and lamellar bone. Although embryonic stem cells show all signs of osteogenic differentiation in vitro, it appears that, in contrast to mesenchymal stem cells, they do not possess the ability to form bone in vivo when a similar culture method and osteogenic differentiation protocol was applied. Copyright © 2010 John Wiley & Sons, Ltd.     

磷酸三钙填充骨缺损后骨形态发生蛋白2和血管内皮生长因子的变化

背景：目前治疗骨缺损以植骨为主,其中磷酸三钙是目前广泛使用的人工骨材料,但对于磷酸三钙的植骨效果仍有争议,其治疗骨缺损的机制尚无详细报道。 目的：观察磷酸三钙植骨填充于骨缺损区后,局部骨形态发生蛋白2和血管内皮生长因子的浓度变化及骨愈合情况。 方法：取C57小鼠48只,随机均分为实验组与对照组,切除右侧股骨干中部2 mm长的骨干和骨膜制作单侧股骨缺损模型,实验组于骨缺损处植入磷酸三钙,对照组不植入任何物质。术后1-4周拍摄X射线片观察骨愈合情况,随后处死动物,植骨区取材,通过Elisa法测定样本中骨形态发生蛋白2、血管内皮生长因子的水平。 结果与结论：X射线显示,实验组术后2周时骨折大部分愈合,小部分骨皮质未完全愈合,3周时骨折基本愈合,仍有少量磷酸三钙残留,4周时骨折完全愈合,周围骨痂生长明显,磷酸三钙基本吸收;对照组术后一二周时仍可见骨折线,第3周时可见骨折线模糊,第4周时骨折部分愈合,部分骨皮质未愈合。实验组不同时间点骨形态发生蛋白2、血管内皮生长因子水平均高于对照组（P＜0.05）。结果表明磷酸三钙植骨治疗可通过上调骨形态发生蛋白2和血管内皮生长因子的表达,促进骨愈合。     

Effect of basic fibroblast growth factor in mouse embryonic stem cell culture and osteogenic differentiation

Embryonic stem cells are actively explored as a cell source in tissue engineering and regenerative medicine involving bone repair. Basic fibroblast growth factor (bFGF) has been a valuable growth factor to support the culture of human stem cells as well as their osteogenic differentiation, but the influence of bFGF on mouse embryonic stem (mES) cells is not known. Towards this goal, D3 cells were treated with bFGF during maintenance conditions and during spontaneous and osteogenic differentiation. In feeder‐free monolayers, up to 40 ng/ml of exogenous bFGF did not support self‐renewal of mES without LIF during cell expansion. During spontaneous differentiation in high‐density cultures, bFGF stimulated cell proliferation under certain conditions but did not influence differentiation, as judged by stage‐specific embryonic antigen‐1 expression. The addition of bFGF reduced the alkaline phosphatase (ALP) activity associated with osteoblast activity during differentiation induced by osteogenic supplements, although the extent of mineralization was unaffected by bFGF. The bFGF increased the mesenchymal stem cell marker Sca‐1 in an mES cell population and led to an enhanced increase in osteocalcin and runx2 expression in combination with BMP‐2. These results suggest that bFGF could be utilized to expand the cell population in high‐density cultures in addition to enriching the BMP‐2 responsiveness of mES cells. Copyright © 2012 John Wiley & Sons, Ltd.     

血小板源生长因子B链的纯合二聚体与血管内皮细胞生长因子在血管网织细胞瘤中的表达及临床意义

目的研究血小板源生长因子B链的纯合二聚体（PDGF—BB）与血管内皮细胞生长因子（VEGF）在正常脑组织和血管网织细胞瘤中的表达及PDGF—BB与VEGF之间的相关性。方法应用免疫组织化学链霉菌抗生物素蛋白-过氧化物酶连接（SP）法对32例人血管网织细胞瘤和10例正常脑组织中PDGF—BB和VEGF的表达进行检测和统计学分析。结果血管网织细胞瘤组织中PDGF—BB和VEGF高于正常脑组织中PDGF—BB和VEGF表达,PDGF-BB灰度值为101．2±15．7VS180．2±14．6,VEGF灰度值为104．4±15．8VS181．5±14．6（均P〈0．01）,并且血管网织细胞瘤组织中PDGF—BB与VEGF表达呈正相关（r＝0．842,P〈0．01）。结论血管网织细胞瘤可分泌PDGF—BB和VEGF,这两种因子在作用机制上可能有密切的联系或存在因果关系。检测PDGF—BB和VEGF。可以为血管网织细胞瘤的发病机制研究及临床治疗提供新的思路。抗PDGF-BB及抗VEGF联合治疗可能成为治疗血管网织细胞瘤的新方法。     

Aflibercept (AVE0005): an alternative strategy for inhibiting tumour angiogenesis by vascular endothelial growth factors

Background: Aberrant angiogenesis is a landmark feature in cancer, which is important for proliferation, growth and metastasis, and is mediated by various pro-angiogenic factors. The VEGF pathway is one of the most important and best-studied angiogenic pathways. Inhibition of this pathway may provide clinical benefits to cancer patients. Objectives: Strategies to inhibit the VEGF pathway, including antibodies to VEGF, antibodies to the extracellular domain of VEGFR-1 or VEGFR-2, decoy receptors for VEGF and tyrosine kinase inhibitors of VEGFRs, are summarized. Methods: This review outlines and compares the latest development of these strategies, with emphasis on aflibercept, a novel decoy fusion protein of domain 2 of VEGFR-1 and domain 3 of VEGFR-2 with the Fc fragment of IgG1. Results: Aflibercept was shown to have early clinical activity. Multiple studies are ongoing to determine the clinical benefits of aflibercept in cancer patients.     

Proliferation and osteogenic differentiation of human bone marrow stromal cells on alginate–gelatine–hydroxyapatite scaffolds with anisotropic pore structure

Porous mineralized scaffolds are required for various applications in bone engineering. In particular, tube‐like pores with controlled orientation inside the scaffold may support homogeneous cell seeding as well as sufficient nutrient supply and may facilitate blood vessel ingrowth. Scaffolds with parallely orientated tube‐like pores were generated by diffusion‐controlled ionotropic gelation of alginate. Incorporation of hydroxyapatite (HA) during the gelation process yielded stable scaffolds with an average pore diameter of approximately 90 µm. To evaluate the potential use of alginate–gelatine–HA scaffolds for bone tissue engineering, in vitro tests with human bone marrow stromal cells (hBMSCs) were carried out. We analysed biocompatibility and cell penetration into the capillary pores by microscopic methods. hBMSCs were also cultivated on alginate–gelatine–HA scaffolds for 3 weeks in the presence and absence of osteogenic supplements. We studied proliferation and osteogenic differentiation in terms of total lactate dehydrogenase (LDH) activity, DNA content and alkaline phosphatase (ALP) activity and found a 10–14‐fold increase of cell number after 2 weeks of cultivation, as well as an increase of specific ALP activity for osteogenic‐induced hBMSCs. Furthermore, the expression of bone‐related genes [ALP, bone sialoprotein II (BSPII)] was analysed. We found an increase of ALP as well as BSPII expression for osteogenic‐induced hBMSCs on alginate–gelatin–HA scaffolds. Copyright © 2008 John Wiley & Sons, Ltd.