血管内皮生长因子联合突变型低氧诱导因子1α的促血管生成作用
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摘要
背景:研究表明,血管内皮生长因子的转录翻译过程在许多缺血、缺氧的条件下都会增加,有效改善机体血管形成与侧支微循环。突变低氧诱导因子1α的表达也需要严格的缺氧条件,很大程度上限制了其实际应用。目的:观察腺病毒介导的血管内皮生长因子联合突变型低氧诱导因子1α(Ad-VEGF-IRES-HIF-1αm u)转染内皮祖细胞在激素性股骨头缺血性坏死修复中促血管生成的作用。方法:①转染Ad-VEGF-IRES-HIF-1αm u到内皮祖细胞,观察细胞活性、形态及细胞病变效应;②将转染Ad-VEGF-IRES-HIF-1αm u成功的内皮祖细胞植入激素性股骨头缺血性坏死动物模型的股骨坏死部位(实验组:转染Ad-VEGF-IRES-HIF-1αm u,对照组:内皮祖细胞细胞悬液,空白组:细胞培养液);③移植10周后,检测血管内皮生长因子、低氧诱导因子1α蛋白表达及CD34表达和微血管密度计数;④墨汁灌注透明切片血管形态学观察。结果与结论:①实验组血管内皮生长因子和低氧诱导因子1α蛋白表达高于对照组与空白组(P<0.05);②实验组的CD34表达阳性的微血管数量较多,且相互之间有连接。③实验组动物股骨头血管墨汁染色显示有新的血管生成,部分血管管径良好,有再通现象,血管间有清晰的连通脉络,有效的血管脉络均匀分布在缺损区域;实验组新生血管面积显著高于对照组和空白组(P <0.05);④结果提示,血管内皮生长因子联合突变型低氧诱导因子1α在激素性股骨头缺血性坏死修复中可以增强血管生成作用。
BACKGROUND: Transcription and translation of vascular endothelial growth factor has been shown to be increased under ischemic and hypoxic conditions, which effectively improves angiogenesis and collateral microcirculation in vivo. Expression of mutant hypoxia-inducible factor-1α(HIF-1αm u) requires hypoxic condition, so its application is limited. OBJECTIVE: To study the effect of transfection of adenovirus-mediated-vascular endothelial growth factor combined with HIF-1αm u(Ad-VEGF-IRES-HIF-1αm u) into endothelial progenitor cells on the angiogenesis in the treatment of steroid-induced avascular necrosis of the femoral head. METHODS: Ad-VEGF-IRES-HIF-1αm u was transfected into endothelial progenitor cells, and the cell viability, morphology and cytopathic effect were observed. Endothelial progenitor cells transfected with Ad-VEGF-IRES-HIF-1αm u were implanted into the necrotic site of the animal model of steroid-induced avascular necrosis of the femoral head(experimental group: transfected with Ad-VEGF-IRES-HIF-1αm u; control group: suspension of endothelial progenitor cells; blank group: cell culture medium). After 10 weeks of transfection, expression of vascular endothelial growth factor, hypoxia-inducible factor1α and CD34 as well as microvessel density were detected. The vascular morphology was observed by ink perfusion. RESULTS AND CONCLUSION: The expression levels of vascular endothelial growth factor and hypoxia-inducible factor1α in the experimental group were higher than those in the control and blank groups(P < 0.05). The number of microvessels positive for CD34 was the argest in the experimental group. In the experimental group, the ink perfusion of femoral head showed new angiogenesis, the phenomenon of recanalization occurred in some vessels, blood vessels had a clear connectivity, and effective vascular veins evenly distributed into the defect area. The area of neovascularization in the experimental group was significantly larger than that in the control and blank groups(P < 0.05). Vascular endothelial growth factor combined with mutant hypoxia-inducible factor1α can enhance angiogenesis in the repair of steroid-induced avascular necrosis of the femoral head.
BACKGROUND: Transcription and translation of vascular endothelial growth factor has been shown to be increased under ischemic and hypoxic conditions, which effectively improves angiogenesis and collateral microcirculation in vivo. Expression of mutant hypoxia-inducible factor-1α(HIF-1αm u) requires hypoxic condition, so its application is limited. OBJECTIVE: To study the effect of transfection of adenovirus-mediated-vascular endothelial growth factor combined with HIF-1αm u(Ad-VEGF-IRES-HIF-1αm u) into endothelial progenitor cells on the angiogenesis in the treatment of steroid-induced avascular necrosis of the femoral head. METHODS: Ad-VEGF-IRES-HIF-1αm u was transfected into endothelial progenitor cells, and the cell viability, morphology and cytopathic effect were observed. Endothelial progenitor cells transfected with Ad-VEGF-IRES-HIF-1αm u were implanted into the necrotic site of the animal model of steroid-induced avascular necrosis of the femoral head(experimental group: transfected with Ad-VEGF-IRES-HIF-1αm u; control group: suspension of endothelial progenitor cells; blank group: cell culture medium). After 10 weeks of transfection, expression of vascular endothelial growth factor, hypoxia-inducible factor1α and CD34 as well as microvessel density were detected. The vascular morphology was observed by ink perfusion. RESULTS AND CONCLUSION: The expression levels of vascular endothelial growth factor and hypoxia-inducible factor1α in the experimental group were higher than those in the control and blank groups(P < 0.05). The number of microvessels positive for CD34 was the argest in the experimental group. In the experimental group, the ink perfusion of femoral head showed new angiogenesis, the phenomenon of recanalization occurred in some vessels, blood vessels had a clear connectivity, and effective vascular veins evenly distributed into the defect area. The area of neovascularization in the experimental group was significantly larger than that in the control and blank groups(P < 0.05). Vascular endothelial growth factor combined with mutant hypoxia-inducible factor1α can enhance angiogenesis in the repair of steroid-induced avascular necrosis of the femoral head.
引文
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[22]Balaji S,Lesaint M,Bhattacharya SS,et al.Adenoviral Mediated Gene Transfer of IGF-1 Enhances Wound Healing and Induces Angiogenesis.J Surg Res.2014;190(1):367-377.
[23]Hatanpaa KJ,Burma S,Zhao D,et al.Epidermal Growth Factor Receptor in Glioma:Signal Transduction,Neuropathology,Imaging,and Radioresistance.Neoplasia.2010;12(9):675-684.
[2]Nabi S,Rao B,Gulati R,et al.Abstracts from the 38th Annual Meeting of the Society of General Internal Medicine.J Gen Intern Med.2015;30 Suppl 2:45-551.
[3]Bai R,Liu W,Zhao A,et al.Nitric oxide content and apoptosis rate in steroid-induced avascular necrosis of the femoral head.Exp Ther Med.2015;10(2):591-597.
[4]Riemenschneider SB,Mattia DJ,Wendel JS,et al.Inosculation and perfusion of pre-vascularized tissue patches containing aligned human microvessels after myocardial infarction.Biomaterials.2016;97:51-61.
[5]Asahara T,Murohara T,Sullivan A,et al.Isolation of putative progenitor endothelial cells for angiogenesis.Science.1997;275(5230):964-967.
[6]Wang S,Chen Z,Tang X,et al.Transplantation of vascular endothelial growth factor 165-transfected endothelial progenitor cells for the treatment of limb ischemia.Mol Med Rep.2015;12(4):4967-4974.
[7]Herzog B,Pelletmany C,Britton G,et al.VEGF binding to NRP1 is essential for VEGF stimulation of endothelial cell migration,complex formation between NRP1 and VEGFR2,and signaling via FAK Tyr407 phosphorylation.Mol Biol Cell.2011;22(15):2766-2776.
[8]Müller CE,Schiedel AC,Zimmermann H.PURINES 2014:nucleotides,nucleosides and nucleobases-international conference on signalling,drugs and targets.Purinergic Signal.2014;10(4):651-656.
[9]王皓,李谌,郜玉忠.突变型低氧诱导因子1α加速骨缺损部位新血管生成的实验观察[J].西安交通大学学报(医学版),2015,36(4):455-461.
[10]Tang W,Yang F,Li Y,et al.Transcriptional Regulation of Vascular Endothelial Growth Factor(VEGF)by Osteoblast-specific Transcription Factor Osterix(Osx)in Osteoblasts*.J Biol Chem.2012;287(3):1671-1678.
[11]戚超,效军,效强,等.多壁碳纳米管改善激素性股骨头坏死模型兔的股骨头形态[J].中国组织工程研究,2014,18(16):2493-2498.
[12]Zwetsloot KA,Westerkamp LM,Holmes BF,et al.AMPKregulates basal skeletal muscle capillarization and VEGFexpression but is not necessary for the angiogenic response to exercise.J Physiol.2008;586(24):6021-6035.
[13]Fulkerson J,Lowe R,Anderson T,et al.Effects of Intraosseous Tibial vs.Intravenous Vasopressin in a Hypovolemic Cardiac Arrest Model.West J Emerg Med.2016;17(2):222-228.
[14]Neves N,Linhares D,Costa G,et al.In vivoand clinical application of strontium-enriched biomaterials for bone regeneration:A systematic review.Bone Joint Res.2017;6(6):366-375.
[15]Billaud M,Lohman AW,Johnstone SR,et al.Regulation of Cellular Communication by Signaling Microdomains in the Blood Vessel Wall.Pharmacol Rev.2014;66(2):513-569.
[16]Michan S,Juan AM,Hurst CG,et al.Sirtuin1 over-expression does not impact retinal vascular and neuronal degeneration in a mouse model of oxygen-induced retinopathy.Plos One.2014;9(1):e85031.
[17]Zhu G,Tang Y,Geng N,et al.HIF-α/MIF and NF-κB/IL-6 Axes Contribute to the Recruitment of CD11b+Gr-1+Myeloid Cells in Hypoxic Microenvironment of HNSCC 1,2.Neoplasia.2014;16(2):168-179.
[18]Biscetti F,Flex A,Pecorini G,et al.The role of High-mobility group box protein 1 in collagen antibody-induced arthritis is dependent on Vascular endothelial growth factor.Clin Exp Immunol.2016;184(1):62-72.
[19]王维军,李嗣生,牛东生,等.股骨头缺血性坏死骨质含量与VEGF、bFGF、BMP-2 mRNA表达的相关性研究[J].中国修复重建外科杂志,2011,25(8):984-991.
[20]Chang KH,Smith SE,Sullivan T,et al.Long-Term Engraftment and Fetal Globin Induction uponBCL11AGene Editing in Bone-Marrow-Derived CD34+Hematopoietic Stem and Progenitor Cells.Mol Ther Methods Clin Dev.2017;4:137-148.
[21]He MQ,He MQ,Wang JF,et al.Vascular Endothelial Growth Factor and Cluster of Differentiation 34 for Assessment of Perioperative Bleeding Risk in Gastric Cancer Patients.Chin Med J(Engl).2016;129(16):1950-1954.
[22]Balaji S,Lesaint M,Bhattacharya SS,et al.Adenoviral Mediated Gene Transfer of IGF-1 Enhances Wound Healing and Induces Angiogenesis.J Surg Res.2014;190(1):367-377.
[23]Hatanpaa KJ,Burma S,Zhao D,et al.Epidermal Growth Factor Receptor in Glioma:Signal Transduction,Neuropathology,Imaging,and Radioresistance.Neoplasia.2010;12(9):675-684.