血管化聚-DL-乳酸/纳米羟基磷灰石复合多孔支架修复兔股骨头坏死的实验研究
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摘要
目的激素性股骨头坏死是临床最常见股骨头坏死类型,其病理机制尚未完全研究清楚,本实验旨在利用一种新型的股骨头坏死造模方法,是动物模型病理改变能接近临床常见病理改变类型,为骨坏死是实验研究提供基础,基本方案为多次小剂量内毒素联合糖皮质激素注射,同时探讨该方法的效率及可行性
材料与方法36只兔子被随机分为四组,A组单纯注射内毒素(LPS,100μg/kg), B组为联合注射内毒素与甲强龙(MPS,20μg/kg),C组为单纯注射甲强龙,D组为空白对照组。最后一次注射后6周各组动物行影像学检测、血液检测以及组织学检测,检测各组骨坏死发生率,比较各组差异。
结果组织学显示各组总骨坏死发生率为83.3%(25／30),骨坏死主要方生部位为干骺端,坏死区与正常骨区别明显,发灰发白,表面不光滑,镜下显示大量细胞碎片,骨髓细胞减少消失,骨小梁稀疏出现空泡。血液学检查联合应用内毒素和甲强龙后血小板数量明显减少,出现高凝状态与低纤溶状态。影像学核磁共振结果显示各组在T1加权像股骨头及股骨近端表现为点状或片状低信号,T2加权像表现为不规则片状高信号；Micro-CT显示坏死区骨小梁稀疏,出现致密或空泡,软骨面变薄,形状不规则。
结论联合应用内毒素和激素能有效诱导兔股骨头坏死,其病理与人早期激素性股骨头坏死类似,该方法效率高,造模时间较短。
目的尝试体外分离培养骨髓间充质干细胞,并向内皮样细胞诱导,将诱导后的内皮样细胞作为种子细胞体外与PDLLA／nHA支架复合培养,并观察细胞支架复合物在体内的成血管活性。
材料与方法分离培养兔骨髓间充质干细胞,原代开始向内皮样细胞诱导,诱导培养至第三代与PDLLA／nHA支架复合培养,观察细胞与支架生物相容性、细胞贴附率以及扫描电镜下观察支架超微结构及体外支架血管化情况。兔肌袋内植入细胞支架复合物,观察体外血管化支架在体内血管诱导活性。
结果骨髓间充质干细胞诱导培养至第三代,流式细胞术检测显示细胞表行为CD31+,CD34+,VEGFR-2+／CD45-,CD105-,免疫荧光CD31阳性,证实诱导后细胞为血管内皮样细胞。细胞支架复合培养后扫描电镜观察支架材料孔隙适中,联通性好,细胞与支架生物相容性良好。细胞支架复合物植入兔肌袋内2周后能在局部形成明显毛细血管样结构,血管诱导性良好。
结论骨髓间充质干细胞能向血管内皮样细胞转化,作为种子细胞与PDLLA／nHA支架生物相容性良好,细胞支架复合物有良好的体内血管诱导能力。
目的尝试利用体外构建的血管化聚-DL-乳酸／纳米羟基磷灰石复合多孔支架治疗兔早期股骨头坏死。
材料与方法采用多次小剂量内毒素联合激素诱导兔股骨头坏死模型,所有动物均行股骨头钻孔减压术,术后将动物随机分为3组：细胞支架复合物植入组、单纯支架植入组以及空白对照组。从血液生化、组织形态、组织病理、影像学等方面检测各组动物样本,并比较各组不同。
结果血液生化方面,细胞支架复合植入组血清降钙素明显高于单纯支架组以及对照组；MRI及Micro-CT显示细胞支架复合植入组可见明显组织修复及新骨形成,VVF及BVF均明显高于其余两组；组织学检测显示细胞支架复合植入组新生血管密度明显大于其余两组,植入区有新骨生成及大量骨髓细胞、骨矿化基质长入。
结论体外血管化聚-DL-乳酸／纳米羟基磷灰石支架,可促进骨坏死区域新生血管形成进而促进坏死骨的修复和新骨生成,是治疗早期激素性股骨头坏死值得尝试的新方法。
Objective Steroid induced osteonecrosis of femoral head is common clinical disease,its pathological mechanism is still not fully understood,this experiment try to create a rabbit osteonecrosis model which pathological symptoms is close to clinical osteonecrosis by a combination application of lipopolysaccharide and methylprednisolone.The efficiency and feasibility will be discussed.
Materials and Methods 36 rabbits were randomly divided into 4 groups:group A to D used only LPS,LPS and MPS,only MPS and control group respectively.All animals were taken blood tests,imaging detection and histological tests 6 weeks after a last injection of LPS or MPS.
Results Histological results showed a total osteonecrosis rate was 83.3%(25/30),the most frequent parts of osteonecrosis is metaphysic,the osteonecrosis was well demarcated,ashcolored and irregularly shaped,the necrotic area showed an accumulation of cell debris,a disappearance of fat cells and bone marrow.Blood platelet levels was significantly decreased in B group than group A and C. Magnetic resonance imaging showed each tl-weighted images in femoral and proximal femoral performance for a patch or flake low signal, the t2-weighted images show high signal for irregular sheets, Micro-CT shows trabeculae sparse in necrotic area, dense or vacuoles, cartilage surface thins, irregular shape.
Conclusions Combined application of LPS and MPS can effectively induce osteonecrosis of femoral head,the animal model had similar pathology symptoms as human osteonecrosis,this approach is efficient.
Objective Try to separate and cultivate BMSCs,and induce them to vascular endothelial cells,seeded the endothelial cells as seed cells in PDLLA/nHA scaffold,cultivated the cells/scaffold complex,observed the biocompatibility,scaffold ultrastructue by SEM,observed the vascular induction ability in vivo.
Materials and Methods The results of flow cytometry showed that immunophenotype of the induced cells were CD31+, CD34+, VEGFR-2+/CD45-, CD105-,confirmed for vascular endothelial cells.The PDLLA/nHA scaffolds had a good biocompatibility with BMSECs,for ultrastructure it had a moderate pore,good connectivity.After 2 weeks implantation the cell/scaffold induced apparent capillary structures.
Conclusions Bone marrow stem cells can be induced into vascular endothelial cells,and can be used as seed cells implant in PDLLA/nHA scaffold,the cell/scaffold complex had a good biocompatibility,and good vascular induction ability in vivo.
Objective Try to repair a rabbit osteonecrosis of femoral head with a vascularized PDLLA/nHA scaffold fabricated in vitro.
Materials and Methods Induced a rabbit femoral head osteonecrosis model by a combination application of LPS and MPS.All animal models were first taken a core decompression,then divided randomly into 3 groups,group A was implanted a BMSECs/PDLLA/nHA scaffold,group B was implanted a PDLLA/nHA scaffold,and group C was control group.Blood samples were collected for osteocalcin detection by ELISA.MRI were taken for imaging analysis,Micro-CT were performed for bone structure and vascular system,HE stain and VWF immunohistochemistry were taken for histological analysis.
Results Blood osteocalcin in group A were significantly higher than other 2 groups;MRI and Micro-CT images showed a obvious new bone formation and tissue repairing in gourp A,VVF and BVF detected by Micor-CT in group A were higher than other 2 groups,histological analysis showed MVD in group A were higher than other 2 groups,and more bone marrow cells and mineralization matrix were seen.
Conclusion Implanting a vasularized PDLLA/nHA scaffold combined with core decompression may represent a promising approach for early stage femoral head osteonecrosis.
材料与方法36只兔子被随机分为四组,A组单纯注射内毒素(LPS,100μg/kg), B组为联合注射内毒素与甲强龙(MPS,20μg/kg),C组为单纯注射甲强龙,D组为空白对照组。最后一次注射后6周各组动物行影像学检测、血液检测以及组织学检测,检测各组骨坏死发生率,比较各组差异。
结果组织学显示各组总骨坏死发生率为83.3%(25／30),骨坏死主要方生部位为干骺端,坏死区与正常骨区别明显,发灰发白,表面不光滑,镜下显示大量细胞碎片,骨髓细胞减少消失,骨小梁稀疏出现空泡。血液学检查联合应用内毒素和甲强龙后血小板数量明显减少,出现高凝状态与低纤溶状态。影像学核磁共振结果显示各组在T1加权像股骨头及股骨近端表现为点状或片状低信号,T2加权像表现为不规则片状高信号；Micro-CT显示坏死区骨小梁稀疏,出现致密或空泡,软骨面变薄,形状不规则。
结论联合应用内毒素和激素能有效诱导兔股骨头坏死,其病理与人早期激素性股骨头坏死类似,该方法效率高,造模时间较短。
目的尝试体外分离培养骨髓间充质干细胞,并向内皮样细胞诱导,将诱导后的内皮样细胞作为种子细胞体外与PDLLA／nHA支架复合培养,并观察细胞支架复合物在体内的成血管活性。
材料与方法分离培养兔骨髓间充质干细胞,原代开始向内皮样细胞诱导,诱导培养至第三代与PDLLA／nHA支架复合培养,观察细胞与支架生物相容性、细胞贴附率以及扫描电镜下观察支架超微结构及体外支架血管化情况。兔肌袋内植入细胞支架复合物,观察体外血管化支架在体内血管诱导活性。
结果骨髓间充质干细胞诱导培养至第三代,流式细胞术检测显示细胞表行为CD31+,CD34+,VEGFR-2+／CD45-,CD105-,免疫荧光CD31阳性,证实诱导后细胞为血管内皮样细胞。细胞支架复合培养后扫描电镜观察支架材料孔隙适中,联通性好,细胞与支架生物相容性良好。细胞支架复合物植入兔肌袋内2周后能在局部形成明显毛细血管样结构,血管诱导性良好。
结论骨髓间充质干细胞能向血管内皮样细胞转化,作为种子细胞与PDLLA／nHA支架生物相容性良好,细胞支架复合物有良好的体内血管诱导能力。
目的尝试利用体外构建的血管化聚-DL-乳酸／纳米羟基磷灰石复合多孔支架治疗兔早期股骨头坏死。
材料与方法采用多次小剂量内毒素联合激素诱导兔股骨头坏死模型,所有动物均行股骨头钻孔减压术,术后将动物随机分为3组：细胞支架复合物植入组、单纯支架植入组以及空白对照组。从血液生化、组织形态、组织病理、影像学等方面检测各组动物样本,并比较各组不同。
结果血液生化方面,细胞支架复合植入组血清降钙素明显高于单纯支架组以及对照组；MRI及Micro-CT显示细胞支架复合植入组可见明显组织修复及新骨形成,VVF及BVF均明显高于其余两组；组织学检测显示细胞支架复合植入组新生血管密度明显大于其余两组,植入区有新骨生成及大量骨髓细胞、骨矿化基质长入。
结论体外血管化聚-DL-乳酸／纳米羟基磷灰石支架,可促进骨坏死区域新生血管形成进而促进坏死骨的修复和新骨生成,是治疗早期激素性股骨头坏死值得尝试的新方法。
Objective Steroid induced osteonecrosis of femoral head is common clinical disease,its pathological mechanism is still not fully understood,this experiment try to create a rabbit osteonecrosis model which pathological symptoms is close to clinical osteonecrosis by a combination application of lipopolysaccharide and methylprednisolone.The efficiency and feasibility will be discussed.
Materials and Methods 36 rabbits were randomly divided into 4 groups:group A to D used only LPS,LPS and MPS,only MPS and control group respectively.All animals were taken blood tests,imaging detection and histological tests 6 weeks after a last injection of LPS or MPS.
Results Histological results showed a total osteonecrosis rate was 83.3%(25/30),the most frequent parts of osteonecrosis is metaphysic,the osteonecrosis was well demarcated,ashcolored and irregularly shaped,the necrotic area showed an accumulation of cell debris,a disappearance of fat cells and bone marrow.Blood platelet levels was significantly decreased in B group than group A and C. Magnetic resonance imaging showed each tl-weighted images in femoral and proximal femoral performance for a patch or flake low signal, the t2-weighted images show high signal for irregular sheets, Micro-CT shows trabeculae sparse in necrotic area, dense or vacuoles, cartilage surface thins, irregular shape.
Conclusions Combined application of LPS and MPS can effectively induce osteonecrosis of femoral head,the animal model had similar pathology symptoms as human osteonecrosis,this approach is efficient.
Objective Try to separate and cultivate BMSCs,and induce them to vascular endothelial cells,seeded the endothelial cells as seed cells in PDLLA/nHA scaffold,cultivated the cells/scaffold complex,observed the biocompatibility,scaffold ultrastructue by SEM,observed the vascular induction ability in vivo.
Materials and Methods The results of flow cytometry showed that immunophenotype of the induced cells were CD31+, CD34+, VEGFR-2+/CD45-, CD105-,confirmed for vascular endothelial cells.The PDLLA/nHA scaffolds had a good biocompatibility with BMSECs,for ultrastructure it had a moderate pore,good connectivity.After 2 weeks implantation the cell/scaffold induced apparent capillary structures.
Conclusions Bone marrow stem cells can be induced into vascular endothelial cells,and can be used as seed cells implant in PDLLA/nHA scaffold,the cell/scaffold complex had a good biocompatibility,and good vascular induction ability in vivo.
Objective Try to repair a rabbit osteonecrosis of femoral head with a vascularized PDLLA/nHA scaffold fabricated in vitro.
Materials and Methods Induced a rabbit femoral head osteonecrosis model by a combination application of LPS and MPS.All animal models were first taken a core decompression,then divided randomly into 3 groups,group A was implanted a BMSECs/PDLLA/nHA scaffold,group B was implanted a PDLLA/nHA scaffold,and group C was control group.Blood samples were collected for osteocalcin detection by ELISA.MRI were taken for imaging analysis,Micro-CT were performed for bone structure and vascular system,HE stain and VWF immunohistochemistry were taken for histological analysis.
Results Blood osteocalcin in group A were significantly higher than other 2 groups;MRI and Micro-CT images showed a obvious new bone formation and tissue repairing in gourp A,VVF and BVF detected by Micor-CT in group A were higher than other 2 groups,histological analysis showed MVD in group A were higher than other 2 groups,and more bone marrow cells and mineralization matrix were seen.
Conclusion Implanting a vasularized PDLLA/nHA scaffold combined with core decompression may represent a promising approach for early stage femoral head osteonecrosis.
引文
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[4]Flemming RG,Murphy CJ,Abrams GA,et al.Effects of synthetic micro-and nano-structured surfaces on cell behavior[J].Biomaterials,1999,20(6):573-588.
[5]Sato M,Webster TJ.Nanobiotechnology:implications of the future of nanotechnology in orthopedic applications [J].Expert Rev Med Devices,2004,1(1):105-114.
[6]Christenson EM,Anseth KS,van den Beucken JJ,et al.Nanobiomaterial applications in orthopedics[J].J Orthop Res,2007,25(1):11-22.
[7]Madeddu P.Therapeutic angiogenesis and vasculogenesis for tissue regeneration[J].Exp Physiol 90:315-326.
[8]Reyes M,Verfaillie OM.Characterization of multipotent adult progenitor cells,a subpopulation of mesenchymal stem cells [J].Ann NYAcad sci,2001,938:231-233.
[9]Reyes M,Dudek A,Jahagirdar B,et al.Origin of endothelial progenitors in human postnatal bone marrow[J].J Clin Invest,2002,109(3):313-315.
[10]E.Polykandriotis,A.Arkudas,et al.Autonomously vascularezed cellular constructs in tissue engineering:opening a new perspective for bilmedical science.J Cell Mol Med,2007,11(1):6-20.
[1]Duvall CL, Taylor WR, Weiss D, Guldberg RE. Quantitative microcomputed tomography analysis of collateral vessel development after ischemic injury. Am J Physiol Heart Circ Physiol 2004;287:H302-H310.
[2]Yamada M,Kubo H,Kobayashi S,et al. Bone marrow derived progenitor cells important for lung repair after lipopolysaccharide-induced lung injury.J Immunol 2004; 172(2):1266-72.
[3]Borue X,Lee S,Grove J,et al.Bone marrow-derived cells contribute to epithelial engraftment during wound healing.Am J Pathol. 2004;165(5):1767-72.
[4]Geiger F,Lorenz H,Xu W,et al.VEGF producing bone marrow stromal cells(BMSC) enhance vascularization and resorption of a natural coral bone substitute[J].Bone,41(2007):516-522.
[5]Weidner N.Intratum oral microvessel density as a prognostic factor in cancer.AJP,1995,147:9.
[6]Colnot C, Romero DM, Huang S, Helms JA.Mechanismsof action of demineralized bone matrix in the repair of cortical bone defects [J]. Clin Orthop Relat Res.435:69-78
[7]Schmid J, Wallkamm B, Hammerle CH, Gogolewski S, Lang NP. The significance of angiogenesis in guided bone regeneration. A case report of a rabbit Experiment[J]. Clin Oral Implants Res 8:244-248.
[8]Pacaccio DJ, Stern SF. Demineralized bone matrix:basic science and clinical applications. Clin Podiatr Med Surg 2005;22(4):599-606
[9]Ziran BH, Smith WR, Morgan SJ. Use of calcium-based demineralized bone matrix/allograft for nonunions and posttraumatic reconstruction of the appendicular skeleton:preliminary results and complications. J Trauma 2007;63(6):1324-1328
[10]Honsawek S, Dhitiseith D, Phupong V. Effects of demineralized bone matrix on proliferation and osteogenic differentiation of mesenchymal stem cells from human umbilical cord. J Med Assoc Thai 2006;89 Suppl3:S189-195
[11]Vaccaro AR, Stubbs HA, Block JE. Demineralized bone matrix composite grafting for posterolateral spinal fusion. Orthopedics 2007;30(7):567-570
[12]Lu B,Tang TT,Yue B,et al.Zhonghua Guke Zazhi 2005;25(10):603-607陆斌,汤亭亭,岳冰,等.BMP-2基因给药修复实验性羊股骨头坏死[J].中华骨科杂志,2005,25(10)：603-607
[13]Tang TT, Lu B, Yue B, et al. Treatment of osteonecrosis of the femoral head with hBMP-2-gene-modified tissue-engineered bone in goats. J Bone Joint Surg Br 2007;89(1):127-129
[14]Schultze-Mosgau S, Lehner B, Rodel F, et al. Expression of bone morphogenic protein 2/4, transforming growth factor-beta1, and bone matrix protein expression in healing area between vascular tibia grafts and irradiated bone-experimental model of osteonecrosis. Int J Radiat Oncol Biol Phys 2005;61(4):1189-1196
[15]Kang P, Shen B, Yang J, et al. Repairing defect and preventing collapse of canine femoral head using titanium implant enhanced by autogenous bone graft and rhBMP-2. Connect Tissue Res 2007;48(4):171-179
[1]Yoshikawa T,Ohgushi H,Nakajima H,et al.In vivo osteogenic durability of cultured bone in porous ceramics:a novel method for autogenous bone graft substitution [J].Trasplantation,2000.69(1):128-134.
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[3]Abukawa H,Terai H,Hannouche D,et al.Formation of a mandibular condyle in vitro by tissue engineering[J]. J Oral Maxillofac Surg.2003 Jan;61(1):94-100.
[4]Mont MA,Ragland PS,Biggins B,et al.Use of bone morphogenetic proteins for musculoskeletal applications.An overview[J].J Bone Joint Surg Am 2004;86-A Suppl 1(Pt 2):S151-158.
[5]Li X,Gong Y,Song Y,et al.Study on the effect of composite of basic fibroblast growth factor and partially deproteinized bone on the repair of femoral head defects[J].Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi.2005 Mar 15;19(3):183-186.Chinese.
[6]Yang C,Yang SH,Du JY,et al.Basic fibroblast growth factor gene transfection to enhance the repair of avascular necrosis of the femoral head[J].Chin Med Sci J.2004 Jun;19(2):111-115.
[7]Matsumoto T,Sasaki J,Alsberg E,et al.Three-dimensional cell and tissue patterning in a strained firin gel system [J].PLoS ONE.2007 Nov 21;2(11):e1211.
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[9]Ochi K,Chen G,Ushida T,et al.Use of isolated matureosteoblasts in abundance acts as desired-shaped bone regenerationin combination with a modified poly-DL-lacitc-co-glycolic acid(PLGA)-collagen sponge [J].J Cell Physiol,2003,194:45-53.
[10]Hollinger JO,Winn SR.Tissue engineering of bone in the craniofacial complex[J].Ann N Y Acad Sci,1999,875(1):379-385.
[11]Alam I,Asahina I.Evaluation of ceramics composed of different hydroxyapatite to tricalcium phosphate ratios as carriers for rhBMP-2.Biomaterials,2001,22:1643-1651.
[12]Tao H,Zhang C,Zeng B,et al.Experimental study on the treatment of femur head necrosis with tricalcium phosphate and platelet-rich plasma[J].Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi.2005 Mar 15;19(3):170-173.
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[14]Ji WF,Tong PJ,Zheng WB,et al.Experimental study on treatment of femoral head necrosis with arterial perfusion of marrow stem cells [J].Zhongguo Xiufu Chongjian Waike Zazhi 2004;24(11):999-1002.
[15]Kawate K,Yajima H,Ohgushi H,et al.Tissue-engineered approach for the treatment of steroid-induced osteonecrosis of the femoral head:transplantation of autologous mesenchymal stem cells cultured with beta-tricalcium phosphate ceramics and free vascularized fibula[J].Artif Organs.2006 Dec;30(12):960-962.
[16]Qiu LY,Wang JF.Advances of studies on mesenchymal stem cells[J].Sheng Wu Gong Cheng Xue Bao.2003 Mar; 19(2):136-140.
[17]Geiger F,Lorenz H,Xu W,et al.VEGF producing bone marrow stromal cells(BMSC) enhance vascularization and resorption of a natural coral bone substitute[J].Bone.2007 Oct;41(4):516-522.Epub 2007 Jul 6.
[18]Xu XL,Lou J,Tang T,et al.Evaluation of different scaffolds for BMP-2 genetic orthopedic tissue engineering[J].J Biomed Mater Res B Appl Biomater.2005 Nov;75(2):289-303.
[19]Mont MA,Jones LC,Einhorn TA,et al.Osteonecrosis of the femoral head.Potential treatment with growth and differentiation factors[J].Clin Orthop,1998,(355) (Suppl):314-325.
[20]Schedel H,Schneller A,Vogl T,et al.Dynamic magnetic resonance tomography(MRI):a follow-up study after femur core decompression and instillation of recombination human bone morphogenetic protein-2(rhBMP-2) in avascular femur head necrosis[J].Rontgenpraxis.2000;53(1):16-24.
[21]Rosen DM,Stempien SA,Thompson AY,et al.Transforming growth factor-beta modulates the expression of osteoblast and chondroblast phenotypes in vitro[J].Cell Physiol,1988,134:337-346.
[22]Kim HJ,Kang SW,Lim HC,et al.The role of transforming growth factor-beta and bone morphogenetic protein with fibrin glue in healing of bone-tendon junction injury [J].Connect Tissue Res.2007;48(6):309-315.
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[1]Xu J, Jiang Y,Chu L,et al.MAPK/ERK signaling mediates VEGF-induced bone marrow stem cell differentiation into endothelial cell.J Cell Mol Med.2008;12(6A):2395-2406.
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[1]Colnot C, Romero DM, Huang S, Helms JA.Mechanismsof action of demineralized bone matrix in the repair of cortical bone defects[J]. Clin Orthop Relat Res.435:69-78
[2]Schmid J, Wallkamm B, Hammerle CH, Gogolewski S, Lang NP. The significance of angiogenesis in guided bone regeneration. A case report of a rabbit Experiment[J]. Clin Oral Implants Res 8:244-248.
[3]Burg KJ,Porter S,Kellam JF.Biomaterial developments for bone tissue engineering[J].Biomaterials,2000,21:2347-2359.
[4]Flemming RG,Murphy CJ,Abrams GA,et al.Effects of synthetic micro-and nano-structured surfaces on cell behavior[J].Biomaterials,1999,20(6):573-588.
[5]Sato M,Webster TJ.Nanobiotechnology:implications of the future of nanotechnology in orthopedic applications [J].Expert Rev Med Devices,2004,1(1):105-114.
[6]Christenson EM,Anseth KS,van den Beucken JJ,et al.Nanobiomaterial applications in orthopedics[J].J Orthop Res,2007,25(1):11-22.
[7]Madeddu P.Therapeutic angiogenesis and vasculogenesis for tissue regeneration[J].Exp Physiol 90:315-326.
[8]Reyes M,Verfaillie OM.Characterization of multipotent adult progenitor cells,a subpopulation of mesenchymal stem cells [J].Ann NYAcad sci,2001,938:231-233.
[9]Reyes M,Dudek A,Jahagirdar B,et al.Origin of endothelial progenitors in human postnatal bone marrow[J].J Clin Invest,2002,109(3):313-315.
[10]E.Polykandriotis,A.Arkudas,et al.Autonomously vascularezed cellular constructs in tissue engineering:opening a new perspective for bilmedical science.J Cell Mol Med,2007,11(1):6-20.
[1]Duvall CL, Taylor WR, Weiss D, Guldberg RE. Quantitative microcomputed tomography analysis of collateral vessel development after ischemic injury. Am J Physiol Heart Circ Physiol 2004;287:H302-H310.
[2]Yamada M,Kubo H,Kobayashi S,et al. Bone marrow derived progenitor cells important for lung repair after lipopolysaccharide-induced lung injury.J Immunol 2004; 172(2):1266-72.
[3]Borue X,Lee S,Grove J,et al.Bone marrow-derived cells contribute to epithelial engraftment during wound healing.Am J Pathol. 2004;165(5):1767-72.
[4]Geiger F,Lorenz H,Xu W,et al.VEGF producing bone marrow stromal cells(BMSC) enhance vascularization and resorption of a natural coral bone substitute[J].Bone,41(2007):516-522.
[5]Weidner N.Intratum oral microvessel density as a prognostic factor in cancer.AJP,1995,147:9.
[6]Colnot C, Romero DM, Huang S, Helms JA.Mechanismsof action of demineralized bone matrix in the repair of cortical bone defects [J]. Clin Orthop Relat Res.435:69-78
[7]Schmid J, Wallkamm B, Hammerle CH, Gogolewski S, Lang NP. The significance of angiogenesis in guided bone regeneration. A case report of a rabbit Experiment[J]. Clin Oral Implants Res 8:244-248.
[8]Pacaccio DJ, Stern SF. Demineralized bone matrix:basic science and clinical applications. Clin Podiatr Med Surg 2005;22(4):599-606
[9]Ziran BH, Smith WR, Morgan SJ. Use of calcium-based demineralized bone matrix/allograft for nonunions and posttraumatic reconstruction of the appendicular skeleton:preliminary results and complications. J Trauma 2007;63(6):1324-1328
[10]Honsawek S, Dhitiseith D, Phupong V. Effects of demineralized bone matrix on proliferation and osteogenic differentiation of mesenchymal stem cells from human umbilical cord. J Med Assoc Thai 2006;89 Suppl3:S189-195
[11]Vaccaro AR, Stubbs HA, Block JE. Demineralized bone matrix composite grafting for posterolateral spinal fusion. Orthopedics 2007;30(7):567-570
[12]Lu B,Tang TT,Yue B,et al.Zhonghua Guke Zazhi 2005;25(10):603-607陆斌,汤亭亭,岳冰,等.BMP-2基因给药修复实验性羊股骨头坏死[J].中华骨科杂志,2005,25(10)：603-607
[13]Tang TT, Lu B, Yue B, et al. Treatment of osteonecrosis of the femoral head with hBMP-2-gene-modified tissue-engineered bone in goats. J Bone Joint Surg Br 2007;89(1):127-129
[14]Schultze-Mosgau S, Lehner B, Rodel F, et al. Expression of bone morphogenic protein 2/4, transforming growth factor-beta1, and bone matrix protein expression in healing area between vascular tibia grafts and irradiated bone-experimental model of osteonecrosis. Int J Radiat Oncol Biol Phys 2005;61(4):1189-1196
[15]Kang P, Shen B, Yang J, et al. Repairing defect and preventing collapse of canine femoral head using titanium implant enhanced by autogenous bone graft and rhBMP-2. Connect Tissue Res 2007;48(4):171-179
[1]Yoshikawa T,Ohgushi H,Nakajima H,et al.In vivo osteogenic durability of cultured bone in porous ceramics:a novel method for autogenous bone graft substitution [J].Trasplantation,2000.69(1):128-134.
[2]Tang TT,Lu B,Yue B,et al.Treatment of osteonecrosis of the femoral head with hBMP-2-gene-modified tissue-engineered bone in goats[J].J Bone Joint Surg Br.2007 Jan;89(1):127-129.
[3]Abukawa H,Terai H,Hannouche D,et al.Formation of a mandibular condyle in vitro by tissue engineering[J]. J Oral Maxillofac Surg.2003 Jan;61(1):94-100.
[4]Mont MA,Ragland PS,Biggins B,et al.Use of bone morphogenetic proteins for musculoskeletal applications.An overview[J].J Bone Joint Surg Am 2004;86-A Suppl 1(Pt 2):S151-158.
[5]Li X,Gong Y,Song Y,et al.Study on the effect of composite of basic fibroblast growth factor and partially deproteinized bone on the repair of femoral head defects[J].Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi.2005 Mar 15;19(3):183-186.Chinese.
[6]Yang C,Yang SH,Du JY,et al.Basic fibroblast growth factor gene transfection to enhance the repair of avascular necrosis of the femoral head[J].Chin Med Sci J.2004 Jun;19(2):111-115.
[7]Matsumoto T,Sasaki J,Alsberg E,et al.Three-dimensional cell and tissue patterning in a strained firin gel system [J].PLoS ONE.2007 Nov 21;2(11):e1211.
[8]Khan YM,Katti DS,Laurencin CT.Novel polymer-synthesized ceramic compositer-based system for bone repair:an in vitro evaluation [J]?J Biomed Mater Res A,2004,69(4):723-728.
[9]Ochi K,Chen G,Ushida T,et al.Use of isolated matureosteoblasts in abundance acts as desired-shaped bone regenerationin combination with a modified poly-DL-lacitc-co-glycolic acid(PLGA)-collagen sponge [J].J Cell Physiol,2003,194:45-53.
[10]Hollinger JO,Winn SR.Tissue engineering of bone in the craniofacial complex[J].Ann N Y Acad Sci,1999,875(1):379-385.
[11]Alam I,Asahina I.Evaluation of ceramics composed of different hydroxyapatite to tricalcium phosphate ratios as carriers for rhBMP-2.Biomaterials,2001,22:1643-1651.
[12]Tao H,Zhang C,Zeng B,et al.Experimental study on the treatment of femur head necrosis with tricalcium phosphate and platelet-rich plasma[J].Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi.2005 Mar 15;19(3):170-173.
[13]Crane GM,Lshaug SL,Mikos AQet al.Bone tissue engineering[J].Nat Med.1995 Dec;1(12):1322-1324.
[14]Ji WF,Tong PJ,Zheng WB,et al.Experimental study on treatment of femoral head necrosis with arterial perfusion of marrow stem cells [J].Zhongguo Xiufu Chongjian Waike Zazhi 2004;24(11):999-1002.
[15]Kawate K,Yajima H,Ohgushi H,et al.Tissue-engineered approach for the treatment of steroid-induced osteonecrosis of the femoral head:transplantation of autologous mesenchymal stem cells cultured with beta-tricalcium phosphate ceramics and free vascularized fibula[J].Artif Organs.2006 Dec;30(12):960-962.
[16]Qiu LY,Wang JF.Advances of studies on mesenchymal stem cells[J].Sheng Wu Gong Cheng Xue Bao.2003 Mar; 19(2):136-140.
[17]Geiger F,Lorenz H,Xu W,et al.VEGF producing bone marrow stromal cells(BMSC) enhance vascularization and resorption of a natural coral bone substitute[J].Bone.2007 Oct;41(4):516-522.Epub 2007 Jul 6.
[18]Xu XL,Lou J,Tang T,et al.Evaluation of different scaffolds for BMP-2 genetic orthopedic tissue engineering[J].J Biomed Mater Res B Appl Biomater.2005 Nov;75(2):289-303.
[19]Mont MA,Jones LC,Einhorn TA,et al.Osteonecrosis of the femoral head.Potential treatment with growth and differentiation factors[J].Clin Orthop,1998,(355) (Suppl):314-325.
[20]Schedel H,Schneller A,Vogl T,et al.Dynamic magnetic resonance tomography(MRI):a follow-up study after femur core decompression and instillation of recombination human bone morphogenetic protein-2(rhBMP-2) in avascular femur head necrosis[J].Rontgenpraxis.2000;53(1):16-24.
[21]Rosen DM,Stempien SA,Thompson AY,et al.Transforming growth factor-beta modulates the expression of osteoblast and chondroblast phenotypes in vitro[J].Cell Physiol,1988,134:337-346.
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