Ang-1基因修饰的Nestin阳性骨髓间充质干细胞治疗脊髓损伤的实验研究
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摘要
目的：(1)分离、培养及鉴定SD大鼠BMSCs,将其诱导分化为Nestin阳性的BMSCs,并通过分子生物学技术将Ang-1基因转染入细胞内；(2)构建SD大鼠T10脊髓钳夹损伤模型,评估其可行性；(3)通过Ang-1基因修饰的Nestin阳性BMSCs局部注射对损伤脊髓进行治疗,检测Nestin阳性的BMSCs是否能够分化为神经元细胞,检测Ang-1对移植入体内的Nestin阳性的BMSCs,新生及损伤后残留的神经元细胞是否具有保护作用。
方法：(1)采用全骨髓差速贴壁分离培养法将SD大鼠双后肢股骨、胫骨骨髓内的BMSCs进行分离培养。通过显微镜下观察细胞生长性状；流式细胞仪检测其细胞表面标志物CD29、CD34、CD45和CD90的表达；向成骨样细胞、成脂肪样细胞及成神经样细胞分化证明其具有多向分化潜能；同时将BMSCs诱导分化为Nestin阳性的BMSCs,并通过腺病毒载体将Ang-1基因转染入Nestin阳性的BMSCs中,Weston blot检测转染后的细胞是否表达Ang-1。(2)构建SD大鼠T10脊髓钳夹损伤模型,造模后1周内通过运动功能评价、神经电生理检测及组织病理学检测等方法检测模型的可行性；(3)造模后1周对随机分组的大鼠实施局部注射移植细胞进行治疗,在随后的1～4周内通过运动功能评价、神经电生理检测及脊髓组织的免疫组织化学检测等方法检测Ang-1基因修饰的Nestin阳性BMSCs对损伤后脊髓的相关作用。
结果：(1)通过全骨髓差速贴壁分离培养的BMSCs增殖速度快,扩增能力强,传代培养3代后细胞可获较高纯度；通过流式细胞仪检测细胞表面标记物,细胞表达CD29和CD90,阳性率均达99%以上；在体外通过诱导分化后细胞可分化为成骨细胞、成脂肪细胞及成神经样细胞；通过Weston blot检测转染后的Nestin阳性BMSCs出现Ang-1条带。(2)造模后1周内,大鼠出现明显截瘫(BBB<4分),经神经电生理检测无法引出CSEP波形,脊髓组织出现大量炎性细胞浸润,脊髓空泡变性。(3)移植后1-4周,A组大鼠后肢神经功能无明显变化,B、C组大鼠均有不同程度的恢复,C组较B组明显(P<0.05)；神经电生理检测,C组亦明显优于B组(P<0.05)；损伤脊髓经免疫组织化学染色后,B、C两组均表达BrdU、Nestin、NSE,随着时间的延长,阳性细胞数逐渐减少,但移植4周后C组仍保持一定量的阳性细胞,且明显多于A、B两组(P<0.05)。C组还可观察到BrdU阳性细胞向血管腔周围聚集。
结论：(1)通过全骨髓差速贴壁分离培养法能够获取较高纯度的BMSCs,细胞具有多向分化潜能,经神经诱导后,细胞能分化为Nestin阳性BMSCs,通过分子生物学技术,Ang-1基因转染入Nestin阳性BMSCs中获得成功。(2)成功构建SD大鼠T10脊髓钳夹型损伤模型。(3)经Ang-1基因修饰后的Nestin阳性BMSCs移植入体内后能进一步分化为成熟的神经元细胞,同时其分泌的Ang-1对Nestin阳性的BMSCs、新生及残留神经细胞具有保护作用,对脊髓损伤具有一定的修复作用。
Objectives:(1) To isolate, culture and identify the BMSCs from the SD rats.To induce and differentiate them into Nestin-positive BMSCs. And tansfected the Ang-1 into the cells by molecular biology. (2)To establish the models of clip compressive thoracic 10 spinal cord injury in the SD rats and investigate the feasibility of the models. (3) To look forward to the possibility of the nestin-positive BMSCs differentiate into neuron in spinal cord injury, and to look forward to the protection of Ang-1 about the nestin-positive BMSCs, the neogenetic and residual neuron in spinal cord injury by tansplanted the nestin-positive BMSCs modified by Ang-1 gene.
Methods:(1)We isolated and cultured the BMSCs by whole bone marrow adherence method from tibias and femurs in SD rats,We observed the morphology of BMSCs by inverted microscope. We performed flow cytometric analysis, using a large monoclonal antibody panel:CD45, CD34, CD29, CD90,and we induced the BMSCs into osteoblast cells,adipocyte and neuron-like cells.In the end we induced the BMSCs into the other cells that the surface marker were positive of Nestin.Then we tansfected the Ang-1 gene into the cells by adenovirus vector, and detected the cells expressed Ang-1 through Weston blot technology. (2)We established graded model of clip compressive thoracic 10 spinal cord injury in the SD rats. One week later, we used the methods based on the assessment of the motor function, neurophysiology monitoring,and pathology test to investigate the feasibility of the model.(3)Rats of randomized grouping were successfully local injected by the certain suspensions a week post-operation, then we assessed the correlated function of Ang-1 and Nestin positive BMSCs to the spine by the methods based on the assessment of the motor function, neurophysiology monitoring, and pathology test after one to four weeks.
Results:(1) BMSCs which was isolated and cultured by the whole bone marrow culture and adherence method, inereased lastingly and had a fast multiplication speed in vitro.The purity quotient of BMSCs after the third generation. BMSCs phenotype was considered positive for CD29, CD90 after flow cytometric analysis, and negative for the other monoclonals.The positive rate of BMSCs phenotype was out of 99%. They could be differentiated into osteoblast cells, adipocyte and neuron-like cells in vitro. The result of weston blot manifest that the Nestin-positive BMSCs transfected by Ang-1 can secreted the angiopoitin protein. (2) Most of the rats became paraplegia after spinal cord injury(BBB<4), the wave shape of CSEP could not be evoked, a lot of inflammatory cells infiltrated the spinal cord which was cytoplasmic vacuolation.(3) Certain neurologial recovery occur in group B, C at 1,2,4 weeks after transplantation, but group A was not. BBB showed significant difference between group B and group C (p<0.05), The CSEP also showed significant difference between group B and group C (p<0.05). After transplanting in group B and C,the cells labled BrdU, Nestin, NSE could be observed in the injury spine,and the cells labled BrdU could be observed around blood vessel. The number of cells labled BrdU, Nestin, NSE reduced gradually in 4 weeks, but group C remain a certain amount of positive cells,and group C showed significant more than group B (p<0.05).
Conclusion:(1)We could harvested the BMSCs which was isolated and cultured by the whole bone marrow culture and adherence method.It was multipotentiality and could be differentiated the BMSCs into Nestin positive cells. we could successfully tansfected the Ang-1 into the nestin-positive BMSCs by molecular biology. (2)The models of SD rats spinal cord injury was successfully established. (3)The Nestin-positive BMSCs tranfected by Ang-1 had the ability to differentiate into neuron, and they restituted and protected the Nestin-positive BMSCs and the nerve cells, cured the injury spine.
方法：(1)采用全骨髓差速贴壁分离培养法将SD大鼠双后肢股骨、胫骨骨髓内的BMSCs进行分离培养。通过显微镜下观察细胞生长性状；流式细胞仪检测其细胞表面标志物CD29、CD34、CD45和CD90的表达；向成骨样细胞、成脂肪样细胞及成神经样细胞分化证明其具有多向分化潜能；同时将BMSCs诱导分化为Nestin阳性的BMSCs,并通过腺病毒载体将Ang-1基因转染入Nestin阳性的BMSCs中,Weston blot检测转染后的细胞是否表达Ang-1。(2)构建SD大鼠T10脊髓钳夹损伤模型,造模后1周内通过运动功能评价、神经电生理检测及组织病理学检测等方法检测模型的可行性；(3)造模后1周对随机分组的大鼠实施局部注射移植细胞进行治疗,在随后的1～4周内通过运动功能评价、神经电生理检测及脊髓组织的免疫组织化学检测等方法检测Ang-1基因修饰的Nestin阳性BMSCs对损伤后脊髓的相关作用。
结果：(1)通过全骨髓差速贴壁分离培养的BMSCs增殖速度快,扩增能力强,传代培养3代后细胞可获较高纯度；通过流式细胞仪检测细胞表面标记物,细胞表达CD29和CD90,阳性率均达99%以上；在体外通过诱导分化后细胞可分化为成骨细胞、成脂肪细胞及成神经样细胞；通过Weston blot检测转染后的Nestin阳性BMSCs出现Ang-1条带。(2)造模后1周内,大鼠出现明显截瘫(BBB<4分),经神经电生理检测无法引出CSEP波形,脊髓组织出现大量炎性细胞浸润,脊髓空泡变性。(3)移植后1-4周,A组大鼠后肢神经功能无明显变化,B、C组大鼠均有不同程度的恢复,C组较B组明显(P<0.05)；神经电生理检测,C组亦明显优于B组(P<0.05)；损伤脊髓经免疫组织化学染色后,B、C两组均表达BrdU、Nestin、NSE,随着时间的延长,阳性细胞数逐渐减少,但移植4周后C组仍保持一定量的阳性细胞,且明显多于A、B两组(P<0.05)。C组还可观察到BrdU阳性细胞向血管腔周围聚集。
结论：(1)通过全骨髓差速贴壁分离培养法能够获取较高纯度的BMSCs,细胞具有多向分化潜能,经神经诱导后,细胞能分化为Nestin阳性BMSCs,通过分子生物学技术,Ang-1基因转染入Nestin阳性BMSCs中获得成功。(2)成功构建SD大鼠T10脊髓钳夹型损伤模型。(3)经Ang-1基因修饰后的Nestin阳性BMSCs移植入体内后能进一步分化为成熟的神经元细胞,同时其分泌的Ang-1对Nestin阳性的BMSCs、新生及残留神经细胞具有保护作用,对脊髓损伤具有一定的修复作用。
Objectives:(1) To isolate, culture and identify the BMSCs from the SD rats.To induce and differentiate them into Nestin-positive BMSCs. And tansfected the Ang-1 into the cells by molecular biology. (2)To establish the models of clip compressive thoracic 10 spinal cord injury in the SD rats and investigate the feasibility of the models. (3) To look forward to the possibility of the nestin-positive BMSCs differentiate into neuron in spinal cord injury, and to look forward to the protection of Ang-1 about the nestin-positive BMSCs, the neogenetic and residual neuron in spinal cord injury by tansplanted the nestin-positive BMSCs modified by Ang-1 gene.
Methods:(1)We isolated and cultured the BMSCs by whole bone marrow adherence method from tibias and femurs in SD rats,We observed the morphology of BMSCs by inverted microscope. We performed flow cytometric analysis, using a large monoclonal antibody panel:CD45, CD34, CD29, CD90,and we induced the BMSCs into osteoblast cells,adipocyte and neuron-like cells.In the end we induced the BMSCs into the other cells that the surface marker were positive of Nestin.Then we tansfected the Ang-1 gene into the cells by adenovirus vector, and detected the cells expressed Ang-1 through Weston blot technology. (2)We established graded model of clip compressive thoracic 10 spinal cord injury in the SD rats. One week later, we used the methods based on the assessment of the motor function, neurophysiology monitoring,and pathology test to investigate the feasibility of the model.(3)Rats of randomized grouping were successfully local injected by the certain suspensions a week post-operation, then we assessed the correlated function of Ang-1 and Nestin positive BMSCs to the spine by the methods based on the assessment of the motor function, neurophysiology monitoring, and pathology test after one to four weeks.
Results:(1) BMSCs which was isolated and cultured by the whole bone marrow culture and adherence method, inereased lastingly and had a fast multiplication speed in vitro.The purity quotient of BMSCs after the third generation. BMSCs phenotype was considered positive for CD29, CD90 after flow cytometric analysis, and negative for the other monoclonals.The positive rate of BMSCs phenotype was out of 99%. They could be differentiated into osteoblast cells, adipocyte and neuron-like cells in vitro. The result of weston blot manifest that the Nestin-positive BMSCs transfected by Ang-1 can secreted the angiopoitin protein. (2) Most of the rats became paraplegia after spinal cord injury(BBB<4), the wave shape of CSEP could not be evoked, a lot of inflammatory cells infiltrated the spinal cord which was cytoplasmic vacuolation.(3) Certain neurologial recovery occur in group B, C at 1,2,4 weeks after transplantation, but group A was not. BBB showed significant difference between group B and group C (p<0.05), The CSEP also showed significant difference between group B and group C (p<0.05). After transplanting in group B and C,the cells labled BrdU, Nestin, NSE could be observed in the injury spine,and the cells labled BrdU could be observed around blood vessel. The number of cells labled BrdU, Nestin, NSE reduced gradually in 4 weeks, but group C remain a certain amount of positive cells,and group C showed significant more than group B (p<0.05).
Conclusion:(1)We could harvested the BMSCs which was isolated and cultured by the whole bone marrow culture and adherence method.It was multipotentiality and could be differentiated the BMSCs into Nestin positive cells. we could successfully tansfected the Ang-1 into the nestin-positive BMSCs by molecular biology. (2)The models of SD rats spinal cord injury was successfully established. (3)The Nestin-positive BMSCs tranfected by Ang-1 had the ability to differentiate into neuron, and they restituted and protected the Nestin-positive BMSCs and the nerve cells, cured the injury spine.
引文
[1]Woodbury D, Reynolds K, Black I B. Adult bone marrow stromal stem cells express germline, ectodermal, endodermal, and mesodermal genes prior to neurogenesis.[J]. J Neurosci Res,2002,69(6):908-917.
[2]Brazelton T R, Rossi F M, Keshet G I, et al. From marrow to brain:expression of neuronal phenotypes in adult mice.[J]. Science,2000,290(5497):1775-1779.
[3]Mezey E, Chandross K J, Harta G, et al. Turning blood into brain:cells bearing neuronal antigens generated in vivo from bone marrow.[J]. Science,2000,290(5497):1779-1782.
[4]Hofstetter C P, Schwarz E J, Hess D, et al. Marrow stromal cells form guiding strands in the injured spinal cord and promote recovery.[J]. Proc Natl Acad Sci U S A,2002,99(4):2199-2204.
[5]Mareschi K, Novara M, Rustichelli D, et al. Neural differentiation of human mesenchymal stem cells:Evidence for expression of neural markers and eag K+channel types. [J]. Exp Hematol,2006,34(11):J563-1572.
[6]Wang L, Wang Z H, Shen C Y, et al. Differentiation of human bone marrow mesenchymal stem cells grown in terpolyesters of 3-hydroxyalkanoates scaffolds into nerve cells.[J]. Biomaterials,2010,31(7):1691-1698.
[7]Sarnowska A, Braun H, Sauerzweig S, et al. The neuroprotective effect of bone marrow stem cells is not dependent on direct cell contact with hypoxic injured tissue.[J]. Exp Neurol,2009,215(2):317-327.
[8]Davis S, Aldrich T H, Jones P F, et al. Isolation of angiopoietin-1, a ligand for the TIE2 receptor, by secretion-trap expression cloning.[J]. Cell,1996,87(7):1161-1169.
[9]Suri C, Mcclain J, Thurston G, et al. Increased vascularization in mice overexpressing angiopoietin-1.[J].Science,1998,282(5388):468-471.
[10]Carvalho R S, Einhorn T A, Lehmann W, et al. The role of angiogenesis in a murine tibial model of distraction osteogenesis.[J]. Bone,2004,34(5):849-861.
[11]Park B H, Yoon S J, Jang K Y, et al. COMP-angiopoietin-1 accelerates bone formation during distraction osteogenesis.[J]. Bone,2010,46(5):1442-1448.
[12]Ismail N S, Pravda E A, Li D, et al. Angiopoietin-1 reduces H(2)O(2)-induced increases in reactive oxygen species and oxidative damage to skin cells.[J]. J Invest Dermatol,2010,130(5):1307-1317.
[13]Kampfer H, Pfeilschifter J, Frank S. Expressional regulation of angiopoietin-1 and-2 and the tie-1 and-2 receptor tyrosine kinases during cutaneous wound healing:a comparative study of normal and impaired repair.[J]. Lab Invest,2001,81(3):361-373.
[14]Valable S, Bellail A, Lesne S, et al. Angiopoietin-1-induced PI3-kinase activation prevents neuronal apoptosis.[J]. FASEB J,2003,17(3):443-445.
[15]Friedenstein A J, Gorskaja J F, Kulagina N N. Fibroblast precursors in normal and irradiated mouse hematopoietic organs.[J]. Exp Hematol,1976,4(5):267-274.
[16]Fromigue O, Hamidouche Z, Chateauvieux S, et al. Distinct osteoblastic differentiation potential of murine fetal liver and bone marrow stroma-derived mesenchymal stem cells. [J]. J Cell Biochem,2008,104(2):620-628.
[17]Prockop D J, Sekiya I, Colter D C. Isolation and characterization of rapidly self-renewing stem cells from cultures of human marrow stromal cells.[J]. Cytotherapy,2001,3(5):393-396.
[18]Huang A H, Chen Y K, Chan A W, et al. Isolation and characterization of normal hamster buccal pouch stem/stromal cells--a potential oral cancer stem/stem-like cell model.[J]. Oral Oncol,2009,45(11):e189-e195.
[19]Van Vlasselaer P, Falla N, Snoeck H, et al. Characterization and purification of osteogenic cells from murine bone marrow by two-color cell sorting using anti-Sca-1 monoclonal antibody and wheat germ agglutinin.[J]. Blood,1994,84(3):753-763.
[20]杨丽,张荣华,谢厚杰,等.建立大鼠骨髓间充质干细胞稳定分离培养体系与鉴定[J].中国组织工程研究与临床康复,2009,13(6)：1064-1068.
[21]裴雪涛.干细胞与再生医学研究及其产业化前景[J].生物产业技术,2009(3)：14-16.
[22]Delorme B, Chateauvieux S, Charbord P. The concept of mesenchymal stem cells. [J]. Regen Med,2006,1(4):497-509.
[23]Lazarus H M, Koc O N, Devine S M, et al. Cotransplantation of HLA-identical sibling culture-expanded mesenchymal stem cells and hematopoietic stem cells in hematologic malignancy patients.[J]. Biol Blood Marrow Transplant,2005,11(5):389-398.
[24]Shahdadfar A, Fronsdal K, Haug T, et al. In vitro expansion of human mesenchymal stem cells: choice of serum is a determinant of cell proliferation, differentiation, gene expression, and transcriptome stability.[J]. Stem Cells,2005,23(9):1357-1366.
[25]Dvorakova J, Hruba A, Velebny V, et al. Isolation and characterization of mesenchymal stem cell population entrapped in bone marrow collection sets.[J]. Cell Biol Int,2008,32(9):1116-1125.
[26]Abdallah B M, Kassem M. Human mesenchymal stem cells:from basic biology to clinical applications.[J]. Gene Ther,2008,15(2):109-116.
[27]Mansilla E, Marin G H, Drago H, et al. Bloodstream cells phenotypically identical to human mesenchymal bone marrow stem cells circulate in large amounts under the influence of acute large skin damage:new evidence for their use in regenerative medicine.[J]. Transplant Proc,2006,38(3):967-969.
[28]Hergeth S P, Aicher W K, Essl M, et al. Characterization and functional analysis of osteoblast-derived fibulins in the human hematopoietic stem cell niche.[J]. Exp Hematol,2008,36(8):1022-1034.
[29]Huang A H, Chen Y K, Chan A W, et al. Isolation and characterization of human dental pulp stem/stromal cells from nonextracted crown-fractured teeth requiring root canal therapy.[J]. J Endod,2009,35(5):673-681.
[30]Majore I, Moretti P, Hass R, et al. Identification of subpopulations in mesenchymal stem cell-like cultures from human umbilical cord.[J]. Cell Commun Signal,2009,7:6.
[31]Hergeth S P, Aicher W K, Essl M, et al. Characterization and functional analysis of osteoblast-derived fibulins in the human hematopoietic stem cell niche.[J]. Exp Hematol,2008,36(8):1022-1034.
[32]Bocchi E A, Bacal F, Guimaraes G, et al. Granulocyte-colony stimulating factor or granulocyte-colony stimulating factor associated to stem cell intracoronary infusion effects in non ischemic refractory heart failure.[J]. Int J Cardiol,2010,138(1):94-97.
[33]Zou X F, Zhu J, Li J M, et al. [Bone marrow stem cell-derived astrocytes are involved in glia limitans formation in rats after brain injury] [J]. Nan Fang Yi Ke Da Xue Xue Bao,2009,29(12):2391-2393.
[34]Kuh S U, Zhu Y, Li J, et al. Can TGF-betal and rhBMP-2 act in synergy to transform bone marrow stem cells to discogenic-type cells?[J]. Acta Neurochir (Wien),2008,150(10):1073-1079,1079.
[35]Shimizu K, Ito A, Yoshida T, et al. Bone tissue engineering with human mesenchymal stem cell sheets constructed using magnetite nanoparticles and magnetic force.[J]. J Biomed Mater Res B Appl Biomater,2007,82(2):471-480.
[36]Lee J H, Kim B G,Ahn J M, et al. Role of PI3K on the regulation of BMP2-induced beta-Catenin activation in human bone marrow stem cells.[J]. Bone,2010,46(6):1522-1532.
[37]Cheng M T, Liu C L, Chen T H, et al. Comparison of Potentials Between Stem Cells Isolated from Human Anterior Cruciate Ligament and Bone Marrow for Ligament Tissue Engineering.[J]. Tissue Eng Part A,2010.
[38]Marie P J, Fromigue O. Osteogenic differentiation of human marrow-derived mesenchymal stem cells.[J]. Regen Med,2006,1(4):539-548.
[39]Duan R P, Wu L, Lin Y F, et al. [The gene expression patterns of bone-marrow mesenchymal stem cells under different osteogenic induction][J]. Sichuan Da Xue Xue Bao Yi Xue Ban,2006,37(6):856-859.
[40]Cui Q, Wang G J, Balian G. Steroid-induced adipogenesis in a pluripotential cell line from bone marrow.[J]. J Bone Joint Surg Am,1997,79(7):1054-1063.
[41]Rosen E D, Walkey C J, Puigserver P, et al. Transcriptional regulation of adipogenesis. [J]. Genes Dev,2000,14(11):1293-1307.
[42]Klemm D J, Roesler W J, Boras T, et al. Insulin stimulates cAMP-response element binding protein activity in HepG2 and 3T3-L1 cell lines.[J]. J Biol Chem,1998,273(2):917-923.
[43]Tang Q Q, Jiang M S, Lane M D. Repressive effect of Spl on the C/EBPalpha gene promoter:role in adipocyte differentiation.[J]. Mol Cell Biol,1999,19(7):4855-4865.
[44]Woodbury D, Schwarz E J, Prockop D J, et al. Adult rat and human bone marrow stromal cells differentiate into neurons.[J]. J Neurosci Res,2000,61(4):364-370.
[45]Crigler L, Robey R C, Asawachaicharn A, et al. Human mesenchymal stem cell subpopulations express a variety of neuro-regulatory molecules and promote neuronal cell survival and neuritogenesis.[J]. Exp Neurol,2006,198(1):54-64.
[46]Qian X, Davis A A, Goderie S K, et al. FGF2 concentration regulates the generation of neurons and glia from multipotent cortical stem cells.[J]. Neuron,1997,18(1):81-93.
[47]王鹏,沈忆新,陆政峰,等.全骨髓法分离培养骨髓间充质干细胞及向神经样细胞分化的研究[J].中国现代医药杂志,2009,11(1)：10-13.
[48]Li Y S, Mahadik S P, Rapport M M, et al. Acute effects of GM1 ganglioside:reduction in both behavioral asymmetry and loss of Na+, K+-ATPase after nigrostriatal transection.[J]. Brain Res,1986,377(2):292-297.
[49]Mahadik S P, Makar T K, Murthy J N, et al. Temporal changes in superoxide dismutase, glutathione peroxidase, and catalase levels in primary and peri-ischemic tissue. Monosialoganglioside (GM1) treatment effects.[J]. Mol Chem Neuropathol,1993,18(1-2):1-14.
[50]Doherty P, Ashton S V, Skaper S D, et al. Ganglioside modulation of neural cell adhesion molecule and N-cadherin-dependent neurite outgrowth.[J]. J Cell Biol,1992,117(5):1093-1099.
[51]Manev H, Favaron M, Vicini S, et al. Ganglioside-mediated protection from glutamate-induced neuronal death.[J]. Acta Neurobiol Exp (Wars),1990,50(4-5):475-488.
[52]Panni M K, Cooper J D, Sofroniew M V. Ganglioside GM1 potentiates NGF action on axotomised medial septal cholinergic neurons.[J]. Brain Res,1998,812(1-2):76-80.
[53]Basso D M, Beattie M S, Bresnahan J C. Graded histological and locomotor outcomes after spinal cord contusion using the NYU weight-drop device versus transection.[J]. Exp Neurol,1996,139(2):244-256.
[54]Allen A R, Marcy G E, Yu J K. The continuous and concurrent use of streptomycin, para-aminosalicylic acid, isoniazid plus early surgery in the treatment of tuberculosis.[J]. Dis Chest,1954,26(1):41-46.
[55]Pu Y, Guo Q S, Wang A M, et al. Repair of acutely injured spinal cord through constructing tissue-engineered neural complex in adult rats.[J]. Chin J Traumatol,2007,10(3):171-176.
[56]田代实,王伟,徐运兰,等.细胞周期素依赖性激酶抑制剂olomoucine对大鼠脊髓损伤后轴突再生微环境的影响及其意义[J].中华医学杂志,2006,86(13)：901-905.
[57]Dolan E J, Transfeldt E E, Tator C H, et al. The effect of spinal distraction on regional spinal cord blood flow in cats.[J]. J Neurosurg,1980,53(6):756-764.
[58]Bunge M B, Holets V R, Bates M L, et al. Characterization of photochemically induced spinal cord injury in the rat by light and electron microscopy.[J]. Exp Neurol,1994,127(1):76-93.
[59]Kwon B K, Oxland T R, Tetzlaff W. Animal models used in spinal cord regeneration research.[J]. Spine (Phila Pa 1976),2002,27(14):1504-1510.
[60]杨迎暴,朴英杰.脊髓损伤模型的建立及其评价标准[J].中华创伤杂志,2002,18(3)：187-190.
[61]Joshi M, Fehlings M G. Development and characterization of a novel, graded model of clip compressive spinal cord injury in the mouse:Part 1. Clip design, behavioral outcomes, and histopathology.[J]. J Neurotrauma,2002,19(2):175-190.
[62]纪江峰,冯世庆.脊髓损伤动物模型研究进展[J].中华实验外科杂志,2006,23(9)：11151-1152.
[63]Rivlin A S, Tator C H. Objective clinical assessment of motor function after experimental spinal cord injury in the rat.[J]. J Neurosurg,1977,47(4):577-581.
[64]Gale K, Kerasidis H, Wrathall J R. Spinal cord contusion in the rat:behavioral analysis of functional neurologic impairment.[J]. Exp Neurol,1985,88(1):123-134.
[65]Basso D M, Beattie M S, Bresnahan J C. Graded histological and locomotor outcomes after spinal cord contusion using the NYU weight-drop device versus transection.[J]. Exp Neurol,1996,139(2):244-256.
[66]Yamada S, Zinke D E, Sanders D. Pathophysiology of "tethered cord syndrome".[J]. J Neurosurg,1981,54(4):494-503.
[67]Agrawal G, Thakor N V, All A H. Evoked potential versus behavior to detect minor insult to the spinal cord in a rat model.[J]. J Clin Neurosci,2009,16(8):1052-1055.
[68]Kida I, Yamamoto T. Comprehensive correlation between neuronal activity and spin-echo blood oxygenation level-dependent signals in the rat somatosensory cortex evoked by short electrical stimulations at various frequencies and currents.[J]. Brain Res,2010,1317:116-123.
[69]Ducker T B, Salcman M, Lucas J T, et al. Experimental spinal cord trauma, Ⅱ:Blood flow, tissue oxygen, evoked potentials in both paretic and plegic monkeys.[J]. Surg Neurol,1978,10(1):64-70.
[70]Nashmi R, Imamura H, Tator C H, et al. Serial recording of somatosensory and myoelectric motor evoked potentials:role in assessing functional recovery after graded spinal cord injury in the rat.[J]. J Neurotrauma,1997,14(3):151-159.
[71]Franceschini M A, Nissila I, Wu W, et al. Coupling between somatosensory evoked potentials and hemodynamic response in the rat.[J]. Neuroimage,2008,41(2):189-203.
[72]Hains B C, Johnson K M, Eaton M J, et al. Serotonergic neural precursor cell grafts attenuate bilateral hyperexcitability of dorsal horn neurons after spinal hemisection in rat.[J]. Neuroscience,2003,116(4):1097-1110.
[73]Willert K, Brown J D, Danenberg E, et al. Wnt proteins are lipid-modified and can act as stem cell growth factors.[J]. Nature,2003,423(6938):448-452.
[74]Grossman S D, Rosenberg L J, Wrathall J R. Temporal-spatial pattern of acute neuronal and glial loss after spinal cord contusion.[J]. Exp Neurol,2001,168(2):273-282.
[75]Kusano K, Enomoto M,Hirai T, et al. Transplanted neural progenitor cells expressing mutant NT3 promote myelination and partial hindlimb recovery in the chronic phase after spinal cord injury.[J]. Biochem Biophys Res Commun,2010,393(4):812-817.
[76]Parr A M, Kulbatski I, Zahir T, et al. Transplanted adult spinal cord-derived neural stem/progenitor cells promote early functional recovery after rat spinal cord injury.[J]. Neuroscience,2008,155(3):760-770.
[77]Ankeny D P, Mctigue D M, Jakeman L B. Bone marrow transplants provide tissue protection and directional guidance for axons after contusive spinal cord injury in rats.[J]. Exp Neurol,2004,190(1):17-31.
[78]Lee J B, Kuroda S, Shichinohe H, et al. A pre-clinical assessment model of rat autogeneic bone marrow stromal cell transplantation into the central nervous system.[J]. Brain Res Brain Res Protoc,2004,14(1):37-44.
[79]Kampfer H, Pfeilschifter J, Frank S. Expressional regulation of angiopoietin-1 and-2 and the tie-1 and-2 receptor tyrosine kinases during cutaneous wound healing:a comparative study of normal and impaired repair.[J]. Lab Invest,2001,81(3):361-373.
[80]Kim I, Kim H G, So J N, et al. Angiopoietin-1 regulates endothelial cell survival through the phosphatidylinositol 3'-Kinase/Akt signal transduction pathway.[J]. Circ Res,2000,86(1):24-29.
[81]Abdel-Malak N A, Mofarrahi M, Mayaki D, et al. Early growth response-1 regulates angiopoietin-1-induced endothelial cell proliferation, migration, and differentiation.[J]. Arterioscler Thromb Vasc Biol,2009,29(2):209-216.
[82]Gamble J R, Drew J, Trezise L, et al. Angiopoietin-1 is an antipermeability and anti-inflammatory agent in vitro and targets cell junctions.[J]. Circ Res,2000,87(7):603-607.
[83]Li X, Stankovic M, Bonder C S, et al. Basal and angiopoietin-1-mediated endothelial permeability is regulated by sphingosine kinase-1.[J]. Blood,2008,111(7):3489-3497.
[84]Arai F, Hirao A, Ohmura M, et al. Tie2/angiopoietin-1 signaling regulates hematopoietic stem cell quiescence in the bone marrow niche.[J]. Cell,2004,118(2):149-161.
[85]Valable S, Bellail A, Lesne S, et al. Angiopoietin-1-induced PI3-kinase activation prevents neuronal apoptosis.[J]. FASEB J,2003,17(3):443-445.
[86]Han S, Arnold S A, Sithu S D, et al. Rescuing vasculature with intravenous angiopoietin-1 and alpha v beta 3 integrin peptide is protective after spinal cord injury.[J]. Brain,2010,133(Pt 4):1026-1042.
[87]Rosa A I, Goncalves J, Cortes L, et al. The angiogenic factor angiopoietin-1 is a proneurogenic peptide on subventricular zone stem/progenitor cells.[J]. J Neurosci,2010,30(13):4573-4584.
[88]Liu X B, Jiang J, Gui C, et al. Angiopoietin-1 protects mesenchymal stem cells against serum deprivation and hypoxia-induced apoptosis through the PI3K/Akt pathway. [J]. Acta Pharmacol Sin,2008,29(7):815-822.
[89]Ezquer F E, Ezquer M E, Parrau D B, et al. Systemic administration of multipotent mesenchymal stromal cells reverts hyperglycemia and prevents nephropathy in type 1 diabetic mice.[J]. Biol Blood Marrow Transplant,2008,14(6).631-640.
[90]Zhang N, Li J, Luo R, et al. Bone marrow mesenchymal stem cells induce angiogenesis and attenuate the remodeling of diabetic cardiomyopathy.[J]. Exp Clin Endocrinol Diabetes,2008,116(2):104-111.
[91]Hancock A, Priester C, Kidder E, et al. Does 5-bromo-2'-deoxyuridine (BrdU) disrupt cell proliferation and neuronal maturation in the adult rat hippocampus in vivo?[J]. Behav Brain Res,2009,199(2):218-221.
[92]Sauerzweig S, Baldauf K, Braun H, et al. Time-dependent segmentation of BrdU-signal leads to late detection problems in studies using BrdU as cell label or proliferation marker.[J]. J Neurosci Methods,2009,177(1):149-159.
[1]Davis S, Aldrich T H, Jones P F, et al. Isolation of angiopoietin-1, a ligand for the TIE2 receptor, by secretion-trap expression cloning.[J]. Cell,1996,87(7):1161-1169.
[2]Maisonpierre P C, Suri C, Jones P F, et al. Angiopoietin-2, a natural antagonist for Tie2 that disrupts in vivo angiogenesis.[J]. Science,1997,277(5322):55-60.
[3]Procopio W N, Pelavin P I, Lee W M, et al. Angiopoietin-1 and-2 coiled coil domains mediate distinct homo-oligomerization patterns, but fibrinogen-like domains mediate ligand activity.[J]. J Biol Chem,1999,274(42):30196-30201.
[4]Stevens M E, Dhillon J, Miller C, et al. SpTiel/2 is expressed in coelomocytes, axial organ and embryos of the sea urchin Strongylocentrotus purpuratus, and is an orthologue of vertebrate Tiel and Tie2.[J]. Dev Comp Immunol,2010.
[5]Kim I, Kim H G, So J N, et al. Angiopoietin-1 regulates endothelial cell survival through the phosphatidylinositol 3'-Kinase/Akt signal transduction pathway.[J]. Circ Res,2000,86(1):24-29.
[6]Abdel-Malak N A, Mofarrahi M, Mayaki D, et al. Early growth response-1 regulates angiopoietin-1-induced endothelial cell proliferation, migration, and differentiation.[J]. Arterioscler Thromb Vase Biol,2009,29(2):209-216.
[7]Gamble J R, Drew J, Trezise L, et al. Angiopoietin-1 is an antipermeability and anti-inflammatory agent in vitro and targets cell junctions. [J]. Circ Res,2000,87(7):603-607.
[8]Li X, Stankovic M, Bonder C S, et al. Basal and angiopoietin-1-mediated endothelial permeability is regulated by sphingosine kinase-1.[J]. Blood,2008,111(7):3489-3497.
[9]Arai F, Hirao A, Ohmura M, et al. Tie2/angiopoietin-1 signaling regulates hematopoietic stem cell quiescence in the bone marrow niche.[J]. Cell,2004,118(2):149-161.
[10]Gomei Y, Nakamura Y, Yoshihara H, et al. Functional differences between two Tie2 ligands, angiopoietin-1 and-2, in regulation of adult bone marrow hematopoietic stem cells.[J]. Exp Hematol,2010,38(2):82-89.
[II]Fiedler U, Reiss Y, Scharpfenecker M, et al. Angiopoietin-2 sensitizes endothelial cells to TNF-alpha and has a crucial role in the induction of inflammation.[J]. Nat Med,2006,12(2):235-239.
[12]Scott B B, Zaratin P F, Gilmartin A G, et al. TNF-alpha modulates angiopoietin-1 expression in rheumatoid synovial fibroblasts via the NF-kappa B signalling pathway.[J]. Biochem Biophys Res Commun,2005,328(2):409-414.
[13]Abdel-Malak N A, Mofarrahi M, Mayaki D, et al. Early growth response-1 regulates angiopoietin-1-induced endothelial cell proliferation, migration, and differentiation.[J]. Arterioscler Thromb Vase Biol,2009,29(2):209-216.
[14]Jones P F. Not just angiogenesis--wider roles for the angiopoietins.[J]. J Pathol,2003,201(4):515-527.
[15]Phelps E D, Updike D L, Bullen E C, et al. Transcriptional and posttranscriptional regulation of angiopoietin-2 expression mediated by IGF and PDGF in vascular smooth muscle cells.[J]. Am J Physiol Cell Physiol,2006,290(2):C352-C361.
[16]Yacyshyn O K, Lai P F, Forse K, et al. Tyrosine phosphatase beta regulates angiopoietin-Tie2 signaling in human endothelial cells.[J]. Angiogenesis,2009,12(1):25-33.
[17]Suri C, Jones P F, Patan S, et al. Requisite role of angiopoietin-1, a ligand for the TIE2 receptor, during embryonic angiogenesis.[J]. Cell,1996,87(7):1171-1180.
[18]Thurston G, Rudge J S, Ioffe E, et al. Angiopoietin-1 protects the adult vasculature against plasma leakage.[J]. Nat Med,2000,6(4):460-463.
[19]Vates G E, Hashimoto T, Young W L, et al. Angiogenesis in the brain during development:the effects of vascular endothelial growth factor and angiopoietin-2 in an animal model. [J]. J Neurosurg,2005,103(1):136-145.
[20]Cassell O C, Hofer S O, Morrison W A, et al. Vascularisation of tissue-engineered grafts:the regulation of angiogenesis in reconstructive surgery and in disease states.[J]. Br J Plast Surg,2002,55(8):603-610.
[21]Jang J H, Shea L D. Controllable delivery of non-viral DNA from porous scaffolds. [J]. J Control Release,2003,86(1):157-168.
[22]Carvalho R S, Einhorn T A, Lehmann W, et al. The role of angiogenesis in a murine tibial model of distraction osteogenesis.[J]. Bone,2004,34(5):849-861.
[23]何剑锋,谢志坚,葛巍立,等.牵张成骨修复兔下颌骨缺损中血管生成素-1的表达及意义[J].口腔医学,2006,26(2)：130-132.
[24]余开湖,冯敢生,郑传胜,等.携带Ang-1的MSCs双介入治疗ANFH的转染与表达[J].临床放射学杂志,2005,24(9)：814-817.
[25]宋建蓉,杨志健,张馥敏,等.大鼠缺血下肢血管新生过程中人血管内皮细胞生长因子165和人血管形成素的协同效应[J].中国临床康复,2004,8(20)：3995-3997,i3页.
[26]Kampfer H, Pfeilschifter J, Frank S. Expressional regulation of angiopoietin-1 and-2 and the tie-1 and-2 receptor tyrosine kinases during cutaneous wound healing:a comparative study of normal and impaired repair.[J]. Lab Invest,2001,81(3):361-373.
[27]Valable S, Bellail A, Lesne S, et al. Angiopoietin-1-induced PI3-kinase activation prevents neuronal apoptosis.[J]. FASEB J,2003,17(3):443-445.
[28]Han S, Arnold S A, Sithu S D, et al. Rescuing vasculature with intravenous angiopoietin-1 and {alpha}v{beta}3 integrin peptide is protective after spinal cord injury.[J]. Brain,2010,133(Pt 4):1026-1042.
[29]Rosa A I, Goncalves J, Cortes L, et al. The angiogenic factor angiopoietin-1 is a proneurogenic Peptide on subventricular zone stem/progenitor cells.[J]. J Neurosci,2010,30(13):4573-4584.
[30]Liu X B, Jiang J, Gui C, et al. Angiopoietin-1 protects mesenchymal stem cells against serum deprivation and hypoxia-induced apoptosis through the PI3K/Akt pathway.[J]. Acta Pharmacol Sin,2008,29(7):815-822.
[31]Folkman J. Angiogenesis in cancer, vascular, rheumatoid and other disease.[J]. Nat Med,1995,1(1):27-31.
[32]Perry B N, Govindarajan B, Bhandarkar S S, et al. Pharmacologic blockade of angiopoietin-2 is efficacious against model hemangiomas in mice.[J]. J Invest Dermatol,2006,126(10):2316-2322.
[33]李皓桓,彭昊,方洪松,等.Ang-1,Ang-2和Tie2基因在骨肿瘤组织中的差异表达[J].武汉大学学报：医学版,2008,29(6)：780-782,791页.
[2]Brazelton T R, Rossi F M, Keshet G I, et al. From marrow to brain:expression of neuronal phenotypes in adult mice.[J]. Science,2000,290(5497):1775-1779.
[3]Mezey E, Chandross K J, Harta G, et al. Turning blood into brain:cells bearing neuronal antigens generated in vivo from bone marrow.[J]. Science,2000,290(5497):1779-1782.
[4]Hofstetter C P, Schwarz E J, Hess D, et al. Marrow stromal cells form guiding strands in the injured spinal cord and promote recovery.[J]. Proc Natl Acad Sci U S A,2002,99(4):2199-2204.
[5]Mareschi K, Novara M, Rustichelli D, et al. Neural differentiation of human mesenchymal stem cells:Evidence for expression of neural markers and eag K+channel types. [J]. Exp Hematol,2006,34(11):J563-1572.
[6]Wang L, Wang Z H, Shen C Y, et al. Differentiation of human bone marrow mesenchymal stem cells grown in terpolyesters of 3-hydroxyalkanoates scaffolds into nerve cells.[J]. Biomaterials,2010,31(7):1691-1698.
[7]Sarnowska A, Braun H, Sauerzweig S, et al. The neuroprotective effect of bone marrow stem cells is not dependent on direct cell contact with hypoxic injured tissue.[J]. Exp Neurol,2009,215(2):317-327.
[8]Davis S, Aldrich T H, Jones P F, et al. Isolation of angiopoietin-1, a ligand for the TIE2 receptor, by secretion-trap expression cloning.[J]. Cell,1996,87(7):1161-1169.
[9]Suri C, Mcclain J, Thurston G, et al. Increased vascularization in mice overexpressing angiopoietin-1.[J].Science,1998,282(5388):468-471.
[10]Carvalho R S, Einhorn T A, Lehmann W, et al. The role of angiogenesis in a murine tibial model of distraction osteogenesis.[J]. Bone,2004,34(5):849-861.
[11]Park B H, Yoon S J, Jang K Y, et al. COMP-angiopoietin-1 accelerates bone formation during distraction osteogenesis.[J]. Bone,2010,46(5):1442-1448.
[12]Ismail N S, Pravda E A, Li D, et al. Angiopoietin-1 reduces H(2)O(2)-induced increases in reactive oxygen species and oxidative damage to skin cells.[J]. J Invest Dermatol,2010,130(5):1307-1317.
[13]Kampfer H, Pfeilschifter J, Frank S. Expressional regulation of angiopoietin-1 and-2 and the tie-1 and-2 receptor tyrosine kinases during cutaneous wound healing:a comparative study of normal and impaired repair.[J]. Lab Invest,2001,81(3):361-373.
[14]Valable S, Bellail A, Lesne S, et al. Angiopoietin-1-induced PI3-kinase activation prevents neuronal apoptosis.[J]. FASEB J,2003,17(3):443-445.
[15]Friedenstein A J, Gorskaja J F, Kulagina N N. Fibroblast precursors in normal and irradiated mouse hematopoietic organs.[J]. Exp Hematol,1976,4(5):267-274.
[16]Fromigue O, Hamidouche Z, Chateauvieux S, et al. Distinct osteoblastic differentiation potential of murine fetal liver and bone marrow stroma-derived mesenchymal stem cells. [J]. J Cell Biochem,2008,104(2):620-628.
[17]Prockop D J, Sekiya I, Colter D C. Isolation and characterization of rapidly self-renewing stem cells from cultures of human marrow stromal cells.[J]. Cytotherapy,2001,3(5):393-396.
[18]Huang A H, Chen Y K, Chan A W, et al. Isolation and characterization of normal hamster buccal pouch stem/stromal cells--a potential oral cancer stem/stem-like cell model.[J]. Oral Oncol,2009,45(11):e189-e195.
[19]Van Vlasselaer P, Falla N, Snoeck H, et al. Characterization and purification of osteogenic cells from murine bone marrow by two-color cell sorting using anti-Sca-1 monoclonal antibody and wheat germ agglutinin.[J]. Blood,1994,84(3):753-763.
[20]杨丽,张荣华,谢厚杰,等.建立大鼠骨髓间充质干细胞稳定分离培养体系与鉴定[J].中国组织工程研究与临床康复,2009,13(6)：1064-1068.
[21]裴雪涛.干细胞与再生医学研究及其产业化前景[J].生物产业技术,2009(3)：14-16.
[22]Delorme B, Chateauvieux S, Charbord P. The concept of mesenchymal stem cells. [J]. Regen Med,2006,1(4):497-509.
[23]Lazarus H M, Koc O N, Devine S M, et al. Cotransplantation of HLA-identical sibling culture-expanded mesenchymal stem cells and hematopoietic stem cells in hematologic malignancy patients.[J]. Biol Blood Marrow Transplant,2005,11(5):389-398.
[24]Shahdadfar A, Fronsdal K, Haug T, et al. In vitro expansion of human mesenchymal stem cells: choice of serum is a determinant of cell proliferation, differentiation, gene expression, and transcriptome stability.[J]. Stem Cells,2005,23(9):1357-1366.
[25]Dvorakova J, Hruba A, Velebny V, et al. Isolation and characterization of mesenchymal stem cell population entrapped in bone marrow collection sets.[J]. Cell Biol Int,2008,32(9):1116-1125.
[26]Abdallah B M, Kassem M. Human mesenchymal stem cells:from basic biology to clinical applications.[J]. Gene Ther,2008,15(2):109-116.
[27]Mansilla E, Marin G H, Drago H, et al. Bloodstream cells phenotypically identical to human mesenchymal bone marrow stem cells circulate in large amounts under the influence of acute large skin damage:new evidence for their use in regenerative medicine.[J]. Transplant Proc,2006,38(3):967-969.
[28]Hergeth S P, Aicher W K, Essl M, et al. Characterization and functional analysis of osteoblast-derived fibulins in the human hematopoietic stem cell niche.[J]. Exp Hematol,2008,36(8):1022-1034.
[29]Huang A H, Chen Y K, Chan A W, et al. Isolation and characterization of human dental pulp stem/stromal cells from nonextracted crown-fractured teeth requiring root canal therapy.[J]. J Endod,2009,35(5):673-681.
[30]Majore I, Moretti P, Hass R, et al. Identification of subpopulations in mesenchymal stem cell-like cultures from human umbilical cord.[J]. Cell Commun Signal,2009,7:6.
[31]Hergeth S P, Aicher W K, Essl M, et al. Characterization and functional analysis of osteoblast-derived fibulins in the human hematopoietic stem cell niche.[J]. Exp Hematol,2008,36(8):1022-1034.
[32]Bocchi E A, Bacal F, Guimaraes G, et al. Granulocyte-colony stimulating factor or granulocyte-colony stimulating factor associated to stem cell intracoronary infusion effects in non ischemic refractory heart failure.[J]. Int J Cardiol,2010,138(1):94-97.
[33]Zou X F, Zhu J, Li J M, et al. [Bone marrow stem cell-derived astrocytes are involved in glia limitans formation in rats after brain injury] [J]. Nan Fang Yi Ke Da Xue Xue Bao,2009,29(12):2391-2393.
[34]Kuh S U, Zhu Y, Li J, et al. Can TGF-betal and rhBMP-2 act in synergy to transform bone marrow stem cells to discogenic-type cells?[J]. Acta Neurochir (Wien),2008,150(10):1073-1079,1079.
[35]Shimizu K, Ito A, Yoshida T, et al. Bone tissue engineering with human mesenchymal stem cell sheets constructed using magnetite nanoparticles and magnetic force.[J]. J Biomed Mater Res B Appl Biomater,2007,82(2):471-480.
[36]Lee J H, Kim B G,Ahn J M, et al. Role of PI3K on the regulation of BMP2-induced beta-Catenin activation in human bone marrow stem cells.[J]. Bone,2010,46(6):1522-1532.
[37]Cheng M T, Liu C L, Chen T H, et al. Comparison of Potentials Between Stem Cells Isolated from Human Anterior Cruciate Ligament and Bone Marrow for Ligament Tissue Engineering.[J]. Tissue Eng Part A,2010.
[38]Marie P J, Fromigue O. Osteogenic differentiation of human marrow-derived mesenchymal stem cells.[J]. Regen Med,2006,1(4):539-548.
[39]Duan R P, Wu L, Lin Y F, et al. [The gene expression patterns of bone-marrow mesenchymal stem cells under different osteogenic induction][J]. Sichuan Da Xue Xue Bao Yi Xue Ban,2006,37(6):856-859.
[40]Cui Q, Wang G J, Balian G. Steroid-induced adipogenesis in a pluripotential cell line from bone marrow.[J]. J Bone Joint Surg Am,1997,79(7):1054-1063.
[41]Rosen E D, Walkey C J, Puigserver P, et al. Transcriptional regulation of adipogenesis. [J]. Genes Dev,2000,14(11):1293-1307.
[42]Klemm D J, Roesler W J, Boras T, et al. Insulin stimulates cAMP-response element binding protein activity in HepG2 and 3T3-L1 cell lines.[J]. J Biol Chem,1998,273(2):917-923.
[43]Tang Q Q, Jiang M S, Lane M D. Repressive effect of Spl on the C/EBPalpha gene promoter:role in adipocyte differentiation.[J]. Mol Cell Biol,1999,19(7):4855-4865.
[44]Woodbury D, Schwarz E J, Prockop D J, et al. Adult rat and human bone marrow stromal cells differentiate into neurons.[J]. J Neurosci Res,2000,61(4):364-370.
[45]Crigler L, Robey R C, Asawachaicharn A, et al. Human mesenchymal stem cell subpopulations express a variety of neuro-regulatory molecules and promote neuronal cell survival and neuritogenesis.[J]. Exp Neurol,2006,198(1):54-64.
[46]Qian X, Davis A A, Goderie S K, et al. FGF2 concentration regulates the generation of neurons and glia from multipotent cortical stem cells.[J]. Neuron,1997,18(1):81-93.
[47]王鹏,沈忆新,陆政峰,等.全骨髓法分离培养骨髓间充质干细胞及向神经样细胞分化的研究[J].中国现代医药杂志,2009,11(1)：10-13.
[48]Li Y S, Mahadik S P, Rapport M M, et al. Acute effects of GM1 ganglioside:reduction in both behavioral asymmetry and loss of Na+, K+-ATPase after nigrostriatal transection.[J]. Brain Res,1986,377(2):292-297.
[49]Mahadik S P, Makar T K, Murthy J N, et al. Temporal changes in superoxide dismutase, glutathione peroxidase, and catalase levels in primary and peri-ischemic tissue. Monosialoganglioside (GM1) treatment effects.[J]. Mol Chem Neuropathol,1993,18(1-2):1-14.
[50]Doherty P, Ashton S V, Skaper S D, et al. Ganglioside modulation of neural cell adhesion molecule and N-cadherin-dependent neurite outgrowth.[J]. J Cell Biol,1992,117(5):1093-1099.
[51]Manev H, Favaron M, Vicini S, et al. Ganglioside-mediated protection from glutamate-induced neuronal death.[J]. Acta Neurobiol Exp (Wars),1990,50(4-5):475-488.
[52]Panni M K, Cooper J D, Sofroniew M V. Ganglioside GM1 potentiates NGF action on axotomised medial septal cholinergic neurons.[J]. Brain Res,1998,812(1-2):76-80.
[53]Basso D M, Beattie M S, Bresnahan J C. Graded histological and locomotor outcomes after spinal cord contusion using the NYU weight-drop device versus transection.[J]. Exp Neurol,1996,139(2):244-256.
[54]Allen A R, Marcy G E, Yu J K. The continuous and concurrent use of streptomycin, para-aminosalicylic acid, isoniazid plus early surgery in the treatment of tuberculosis.[J]. Dis Chest,1954,26(1):41-46.
[55]Pu Y, Guo Q S, Wang A M, et al. Repair of acutely injured spinal cord through constructing tissue-engineered neural complex in adult rats.[J]. Chin J Traumatol,2007,10(3):171-176.
[56]田代实,王伟,徐运兰,等.细胞周期素依赖性激酶抑制剂olomoucine对大鼠脊髓损伤后轴突再生微环境的影响及其意义[J].中华医学杂志,2006,86(13)：901-905.
[57]Dolan E J, Transfeldt E E, Tator C H, et al. The effect of spinal distraction on regional spinal cord blood flow in cats.[J]. J Neurosurg,1980,53(6):756-764.
[58]Bunge M B, Holets V R, Bates M L, et al. Characterization of photochemically induced spinal cord injury in the rat by light and electron microscopy.[J]. Exp Neurol,1994,127(1):76-93.
[59]Kwon B K, Oxland T R, Tetzlaff W. Animal models used in spinal cord regeneration research.[J]. Spine (Phila Pa 1976),2002,27(14):1504-1510.
[60]杨迎暴,朴英杰.脊髓损伤模型的建立及其评价标准[J].中华创伤杂志,2002,18(3)：187-190.
[61]Joshi M, Fehlings M G. Development and characterization of a novel, graded model of clip compressive spinal cord injury in the mouse:Part 1. Clip design, behavioral outcomes, and histopathology.[J]. J Neurotrauma,2002,19(2):175-190.
[62]纪江峰,冯世庆.脊髓损伤动物模型研究进展[J].中华实验外科杂志,2006,23(9)：11151-1152.
[63]Rivlin A S, Tator C H. Objective clinical assessment of motor function after experimental spinal cord injury in the rat.[J]. J Neurosurg,1977,47(4):577-581.
[64]Gale K, Kerasidis H, Wrathall J R. Spinal cord contusion in the rat:behavioral analysis of functional neurologic impairment.[J]. Exp Neurol,1985,88(1):123-134.
[65]Basso D M, Beattie M S, Bresnahan J C. Graded histological and locomotor outcomes after spinal cord contusion using the NYU weight-drop device versus transection.[J]. Exp Neurol,1996,139(2):244-256.
[66]Yamada S, Zinke D E, Sanders D. Pathophysiology of "tethered cord syndrome".[J]. J Neurosurg,1981,54(4):494-503.
[67]Agrawal G, Thakor N V, All A H. Evoked potential versus behavior to detect minor insult to the spinal cord in a rat model.[J]. J Clin Neurosci,2009,16(8):1052-1055.
[68]Kida I, Yamamoto T. Comprehensive correlation between neuronal activity and spin-echo blood oxygenation level-dependent signals in the rat somatosensory cortex evoked by short electrical stimulations at various frequencies and currents.[J]. Brain Res,2010,1317:116-123.
[69]Ducker T B, Salcman M, Lucas J T, et al. Experimental spinal cord trauma, Ⅱ:Blood flow, tissue oxygen, evoked potentials in both paretic and plegic monkeys.[J]. Surg Neurol,1978,10(1):64-70.
[70]Nashmi R, Imamura H, Tator C H, et al. Serial recording of somatosensory and myoelectric motor evoked potentials:role in assessing functional recovery after graded spinal cord injury in the rat.[J]. J Neurotrauma,1997,14(3):151-159.
[71]Franceschini M A, Nissila I, Wu W, et al. Coupling between somatosensory evoked potentials and hemodynamic response in the rat.[J]. Neuroimage,2008,41(2):189-203.
[72]Hains B C, Johnson K M, Eaton M J, et al. Serotonergic neural precursor cell grafts attenuate bilateral hyperexcitability of dorsal horn neurons after spinal hemisection in rat.[J]. Neuroscience,2003,116(4):1097-1110.
[73]Willert K, Brown J D, Danenberg E, et al. Wnt proteins are lipid-modified and can act as stem cell growth factors.[J]. Nature,2003,423(6938):448-452.
[74]Grossman S D, Rosenberg L J, Wrathall J R. Temporal-spatial pattern of acute neuronal and glial loss after spinal cord contusion.[J]. Exp Neurol,2001,168(2):273-282.
[75]Kusano K, Enomoto M,Hirai T, et al. Transplanted neural progenitor cells expressing mutant NT3 promote myelination and partial hindlimb recovery in the chronic phase after spinal cord injury.[J]. Biochem Biophys Res Commun,2010,393(4):812-817.
[76]Parr A M, Kulbatski I, Zahir T, et al. Transplanted adult spinal cord-derived neural stem/progenitor cells promote early functional recovery after rat spinal cord injury.[J]. Neuroscience,2008,155(3):760-770.
[77]Ankeny D P, Mctigue D M, Jakeman L B. Bone marrow transplants provide tissue protection and directional guidance for axons after contusive spinal cord injury in rats.[J]. Exp Neurol,2004,190(1):17-31.
[78]Lee J B, Kuroda S, Shichinohe H, et al. A pre-clinical assessment model of rat autogeneic bone marrow stromal cell transplantation into the central nervous system.[J]. Brain Res Brain Res Protoc,2004,14(1):37-44.
[79]Kampfer H, Pfeilschifter J, Frank S. Expressional regulation of angiopoietin-1 and-2 and the tie-1 and-2 receptor tyrosine kinases during cutaneous wound healing:a comparative study of normal and impaired repair.[J]. Lab Invest,2001,81(3):361-373.
[80]Kim I, Kim H G, So J N, et al. Angiopoietin-1 regulates endothelial cell survival through the phosphatidylinositol 3'-Kinase/Akt signal transduction pathway.[J]. Circ Res,2000,86(1):24-29.
[81]Abdel-Malak N A, Mofarrahi M, Mayaki D, et al. Early growth response-1 regulates angiopoietin-1-induced endothelial cell proliferation, migration, and differentiation.[J]. Arterioscler Thromb Vasc Biol,2009,29(2):209-216.
[82]Gamble J R, Drew J, Trezise L, et al. Angiopoietin-1 is an antipermeability and anti-inflammatory agent in vitro and targets cell junctions.[J]. Circ Res,2000,87(7):603-607.
[83]Li X, Stankovic M, Bonder C S, et al. Basal and angiopoietin-1-mediated endothelial permeability is regulated by sphingosine kinase-1.[J]. Blood,2008,111(7):3489-3497.
[84]Arai F, Hirao A, Ohmura M, et al. Tie2/angiopoietin-1 signaling regulates hematopoietic stem cell quiescence in the bone marrow niche.[J]. Cell,2004,118(2):149-161.
[85]Valable S, Bellail A, Lesne S, et al. Angiopoietin-1-induced PI3-kinase activation prevents neuronal apoptosis.[J]. FASEB J,2003,17(3):443-445.
[86]Han S, Arnold S A, Sithu S D, et al. Rescuing vasculature with intravenous angiopoietin-1 and alpha v beta 3 integrin peptide is protective after spinal cord injury.[J]. Brain,2010,133(Pt 4):1026-1042.
[87]Rosa A I, Goncalves J, Cortes L, et al. The angiogenic factor angiopoietin-1 is a proneurogenic peptide on subventricular zone stem/progenitor cells.[J]. J Neurosci,2010,30(13):4573-4584.
[88]Liu X B, Jiang J, Gui C, et al. Angiopoietin-1 protects mesenchymal stem cells against serum deprivation and hypoxia-induced apoptosis through the PI3K/Akt pathway. [J]. Acta Pharmacol Sin,2008,29(7):815-822.
[89]Ezquer F E, Ezquer M E, Parrau D B, et al. Systemic administration of multipotent mesenchymal stromal cells reverts hyperglycemia and prevents nephropathy in type 1 diabetic mice.[J]. Biol Blood Marrow Transplant,2008,14(6).631-640.
[90]Zhang N, Li J, Luo R, et al. Bone marrow mesenchymal stem cells induce angiogenesis and attenuate the remodeling of diabetic cardiomyopathy.[J]. Exp Clin Endocrinol Diabetes,2008,116(2):104-111.
[91]Hancock A, Priester C, Kidder E, et al. Does 5-bromo-2'-deoxyuridine (BrdU) disrupt cell proliferation and neuronal maturation in the adult rat hippocampus in vivo?[J]. Behav Brain Res,2009,199(2):218-221.
[92]Sauerzweig S, Baldauf K, Braun H, et al. Time-dependent segmentation of BrdU-signal leads to late detection problems in studies using BrdU as cell label or proliferation marker.[J]. J Neurosci Methods,2009,177(1):149-159.
[1]Davis S, Aldrich T H, Jones P F, et al. Isolation of angiopoietin-1, a ligand for the TIE2 receptor, by secretion-trap expression cloning.[J]. Cell,1996,87(7):1161-1169.
[2]Maisonpierre P C, Suri C, Jones P F, et al. Angiopoietin-2, a natural antagonist for Tie2 that disrupts in vivo angiogenesis.[J]. Science,1997,277(5322):55-60.
[3]Procopio W N, Pelavin P I, Lee W M, et al. Angiopoietin-1 and-2 coiled coil domains mediate distinct homo-oligomerization patterns, but fibrinogen-like domains mediate ligand activity.[J]. J Biol Chem,1999,274(42):30196-30201.
[4]Stevens M E, Dhillon J, Miller C, et al. SpTiel/2 is expressed in coelomocytes, axial organ and embryos of the sea urchin Strongylocentrotus purpuratus, and is an orthologue of vertebrate Tiel and Tie2.[J]. Dev Comp Immunol,2010.
[5]Kim I, Kim H G, So J N, et al. Angiopoietin-1 regulates endothelial cell survival through the phosphatidylinositol 3'-Kinase/Akt signal transduction pathway.[J]. Circ Res,2000,86(1):24-29.
[6]Abdel-Malak N A, Mofarrahi M, Mayaki D, et al. Early growth response-1 regulates angiopoietin-1-induced endothelial cell proliferation, migration, and differentiation.[J]. Arterioscler Thromb Vase Biol,2009,29(2):209-216.
[7]Gamble J R, Drew J, Trezise L, et al. Angiopoietin-1 is an antipermeability and anti-inflammatory agent in vitro and targets cell junctions. [J]. Circ Res,2000,87(7):603-607.
[8]Li X, Stankovic M, Bonder C S, et al. Basal and angiopoietin-1-mediated endothelial permeability is regulated by sphingosine kinase-1.[J]. Blood,2008,111(7):3489-3497.
[9]Arai F, Hirao A, Ohmura M, et al. Tie2/angiopoietin-1 signaling regulates hematopoietic stem cell quiescence in the bone marrow niche.[J]. Cell,2004,118(2):149-161.
[10]Gomei Y, Nakamura Y, Yoshihara H, et al. Functional differences between two Tie2 ligands, angiopoietin-1 and-2, in regulation of adult bone marrow hematopoietic stem cells.[J]. Exp Hematol,2010,38(2):82-89.
[II]Fiedler U, Reiss Y, Scharpfenecker M, et al. Angiopoietin-2 sensitizes endothelial cells to TNF-alpha and has a crucial role in the induction of inflammation.[J]. Nat Med,2006,12(2):235-239.
[12]Scott B B, Zaratin P F, Gilmartin A G, et al. TNF-alpha modulates angiopoietin-1 expression in rheumatoid synovial fibroblasts via the NF-kappa B signalling pathway.[J]. Biochem Biophys Res Commun,2005,328(2):409-414.
[13]Abdel-Malak N A, Mofarrahi M, Mayaki D, et al. Early growth response-1 regulates angiopoietin-1-induced endothelial cell proliferation, migration, and differentiation.[J]. Arterioscler Thromb Vase Biol,2009,29(2):209-216.
[14]Jones P F. Not just angiogenesis--wider roles for the angiopoietins.[J]. J Pathol,2003,201(4):515-527.
[15]Phelps E D, Updike D L, Bullen E C, et al. Transcriptional and posttranscriptional regulation of angiopoietin-2 expression mediated by IGF and PDGF in vascular smooth muscle cells.[J]. Am J Physiol Cell Physiol,2006,290(2):C352-C361.
[16]Yacyshyn O K, Lai P F, Forse K, et al. Tyrosine phosphatase beta regulates angiopoietin-Tie2 signaling in human endothelial cells.[J]. Angiogenesis,2009,12(1):25-33.
[17]Suri C, Jones P F, Patan S, et al. Requisite role of angiopoietin-1, a ligand for the TIE2 receptor, during embryonic angiogenesis.[J]. Cell,1996,87(7):1171-1180.
[18]Thurston G, Rudge J S, Ioffe E, et al. Angiopoietin-1 protects the adult vasculature against plasma leakage.[J]. Nat Med,2000,6(4):460-463.
[19]Vates G E, Hashimoto T, Young W L, et al. Angiogenesis in the brain during development:the effects of vascular endothelial growth factor and angiopoietin-2 in an animal model. [J]. J Neurosurg,2005,103(1):136-145.
[20]Cassell O C, Hofer S O, Morrison W A, et al. Vascularisation of tissue-engineered grafts:the regulation of angiogenesis in reconstructive surgery and in disease states.[J]. Br J Plast Surg,2002,55(8):603-610.
[21]Jang J H, Shea L D. Controllable delivery of non-viral DNA from porous scaffolds. [J]. J Control Release,2003,86(1):157-168.
[22]Carvalho R S, Einhorn T A, Lehmann W, et al. The role of angiogenesis in a murine tibial model of distraction osteogenesis.[J]. Bone,2004,34(5):849-861.
[23]何剑锋,谢志坚,葛巍立,等.牵张成骨修复兔下颌骨缺损中血管生成素-1的表达及意义[J].口腔医学,2006,26(2)：130-132.
[24]余开湖,冯敢生,郑传胜,等.携带Ang-1的MSCs双介入治疗ANFH的转染与表达[J].临床放射学杂志,2005,24(9)：814-817.
[25]宋建蓉,杨志健,张馥敏,等.大鼠缺血下肢血管新生过程中人血管内皮细胞生长因子165和人血管形成素的协同效应[J].中国临床康复,2004,8(20)：3995-3997,i3页.
[26]Kampfer H, Pfeilschifter J, Frank S. Expressional regulation of angiopoietin-1 and-2 and the tie-1 and-2 receptor tyrosine kinases during cutaneous wound healing:a comparative study of normal and impaired repair.[J]. Lab Invest,2001,81(3):361-373.
[27]Valable S, Bellail A, Lesne S, et al. Angiopoietin-1-induced PI3-kinase activation prevents neuronal apoptosis.[J]. FASEB J,2003,17(3):443-445.
[28]Han S, Arnold S A, Sithu S D, et al. Rescuing vasculature with intravenous angiopoietin-1 and {alpha}v{beta}3 integrin peptide is protective after spinal cord injury.[J]. Brain,2010,133(Pt 4):1026-1042.
[29]Rosa A I, Goncalves J, Cortes L, et al. The angiogenic factor angiopoietin-1 is a proneurogenic Peptide on subventricular zone stem/progenitor cells.[J]. J Neurosci,2010,30(13):4573-4584.
[30]Liu X B, Jiang J, Gui C, et al. Angiopoietin-1 protects mesenchymal stem cells against serum deprivation and hypoxia-induced apoptosis through the PI3K/Akt pathway.[J]. Acta Pharmacol Sin,2008,29(7):815-822.
[31]Folkman J. Angiogenesis in cancer, vascular, rheumatoid and other disease.[J]. Nat Med,1995,1(1):27-31.
[32]Perry B N, Govindarajan B, Bhandarkar S S, et al. Pharmacologic blockade of angiopoietin-2 is efficacious against model hemangiomas in mice.[J]. J Invest Dermatol,2006,126(10):2316-2322.
[33]李皓桓,彭昊,方洪松,等.Ang-1,Ang-2和Tie2基因在骨肿瘤组织中的差异表达[J].武汉大学学报：医学版,2008,29(6)：780-782,791页.
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