组织工程骺板的构建及其治疗骺板损伤的实验研究
|
摘要
一、骺板组织工程种子细胞——兔骺板软骨细胞体外培养及其细胞表型的研究
目的探索体外培养兔骺板软骨细胞的方法,并对其细胞表型进行初步研究,为骺板组织工程寻找良好的种子细胞。
方法取两周龄,体重250g左右幼兔胫骨上端骺板,经过机械剪切、胰酶、胶原酶的消化获得细胞后,1640培养基体外培养。应用倒置显微镜、透射电镜、Alcian blue染色、蕃红花“O”--亮绿染色,免疫组织化学染色、PCR、MTT等方法,对细胞进行定性、定量检测。
结果在体外观察到了骺板软骨细胞贴壁生长及传代的生长过程,多次传代后增殖能力降低。PCR检测有II型胶原的表达、蕃红花“O”--亮绿染色阳性、Alcian blue染色显示第三代细胞蛋白聚糖含量高,MTT显示第三代细胞生长曲线呈倒“S”型,于第12天出现生长抑制。
结论成功地建立了骺板软骨细胞的体外培养方法,并证实所培养出的细胞是骺板软骨细胞。
二、组织工程骺板的构建——兔骺板软骨细胞分别与多孔聚乳酸、多孔丝素构建两种组织工程骺板并比较其体外相容性
目的探索兔骺板软骨细胞与生物支架材料复合构建组织工程骺板的可行性。
方法将体外培养的第三代兔骺板软骨细胞分别与多孔聚乳酸材料、多孔丝素材料复合,构建两种组织工程骺板。应用扫描电镜、激光共聚焦显微镜、MTT等方法,对细胞与材料的相容性进行观察,并对两种生物支架材料进行比较。
结果兔骺板软骨细胞在支架材料上生长良好,随着培养时间的延长,分裂增殖的细胞逐渐增多。细胞在两种材料上生长情况类似,统计学显示无显著性差异。
结论多孔聚乳酸材料和多孔丝素材料均具有良好的生物相容性,可以作为骺板组织工程理想的载体材料。
三、组织工程骺板治疗骺板损伤的动物实验研究——两种组织工程骺板分别治疗骺板损伤并对其进行比较
目的探索组织工程骺板在治疗骺板损伤中的作用。
方法将两种组织工程骺板分别植入骺板损伤的模型中,同时设单纯植入材料组、空白对照组。应用组织学、BrdU细胞示踪、X线等方法,观察成角畸形和短缩畸形情况。同时对两种组织工程骺板、以及两种支架材料进行比较。
结果空白组,骨桥明显,短缩畸形和成角畸形显著;单纯植入支架材料组,短缩畸形和成角畸形明显;组织工程骺板组,存在较小的短缩畸形和成角畸形。统计学显示三种方法存在显著性差异,两种组织工程骺板之间,两种支架材料之间无显著性差异。
结论组织工程骺板治疗骺板损伤是一种有效的方法,其疗效优于单纯植入生物支架材料。
四、人骨骺软骨细胞体外培养及鉴定的初步研究
目的建立体外培养及鉴定人骨骺软骨细胞的方法。
方法取人多指切除手术之多指骨骺,先行机械剪切,后用胰蛋白酶及II型胶原酶消化,再接种于1640培养基中体外培养。应用蕃红花“O”--亮绿染色,Alcian blue染色,免疫组化II型胶原染色和MTT检测,对其生物学特性进行了观察和鉴定。
结果在体外观察到了人骨骺软骨细胞贴壁生长及传代的生长过程,多次传代后增殖能力降低。蕃红花“O”--亮绿染色阳性。Alcian blue染色阳性。免疫组化II型胶原染色阳性。MTT显示生长曲线呈倒“S”型,至第15天细胞出现生长抑制。
结论成功地建立了人骨骺软骨细胞的体外培养方法,并证实所培养出的细胞是人骨骺软骨细胞。
Part I: The seed cells of epiphyseal plate tissue engineering -- the methods of culture the epiphyseal plate chondrocyte in vitro and its cellular phenotype in rabbits.
Objective To establish a kind of method about culture the epiphyseal plate chondrocyte in vitro, so that we can obtain the seed cells for the useage of epiphyseal plate tissue engineering.
Methods The cells we used come from proximal epiphyseal plate of tibia of rabbits. Before we culture the cells, we used Trypsin and Collagenese II . Checking up biological characteristics of the cells by applying the ways of microscope and MTT test. Identifying the cells by applying the ways of safrarine“o”staining and PCR test.
Results We have observed the conditions of the growth of the epiphyseal plate chondrocyte. The test of safrarine“o”staining shows positive. The test of PCR shows the expreesion of II collagen. The test of MTT shows the biological properties of the cells decrease from the 12th day.
Conclusions We have established a method of culture the epiphyseal plate chondrocyte, at the same time we have identified that the cells were the epiphyseal plate chondrocyte.
Part II: The construction of engineering epiphyseal plate ----the study on the biocompatibility of epiphyseal plate chondrocyte with scaffold PLA and epiphyseal plate chondrocyte with scaffold SF.
Objective To explore the way of construction about engineering epiphyseal plate using epiphyseal plate chondrocyte and scaffold.
Methods To construct two engineering epiphyseal plates by culturing the epiphyseal plate chondrocyte on scaffold PLA and scaffold SF. Checking up biological characteristics of the cells by applying the ways of scanning electron- microscope, laser scan confocal microscope and MTT test.
Results The epiphyseal plate chondrocyte grew well on scaffold PLA and scaffold SF. The function of the tow scaffold is similar.
Conclusions Both scaffold PLA and scaffold SF have good biocompatibility and can be used as ideal scaffold in tissue engineering.
Part III: The animal experimental study on the application in epiphyseal plate injury of the two engineering epiphyseal plates.
Objective To observe the role of engineering epiphyseal plate in the treatment of epiphyseal plate injury.
Methods The two engineering epiphyseal plates were implanted into the model of epiphyseal plate injury. Evaluating the role by histology, BrdU and X-ray. At the same time, comparing engineering epiphyseal plate to scaffold .
Results The degree of deformity in engineering epiphyseal plate group was less that scaffold group ,whereas the degree of deformity in scaffold group less than control group .The two engineering epiphyseal plate groups are similar , and the two scaffold groups are also similar.
Conclusions It is feasible to treat epiphyseal plate injury by engineering epiphyseal plate, and it is better than by scaffold.
PartⅣ: The methods of culture and identification of the human epiphyseal chondrocyte.
Objective To establish a kind of method about culture and identification of the human epiphyseal chondrocyte.
Methods The cells we used come from epiphysis of polydactyly. Before we culture the cells, we used Trypsin and Collagenase II . Checking up biological characteristics of the cells by applying the ways of MTT test. Identifying the cells by applying the ways of safrarine“o”staining, Alcian blue test and immuno- histochemistry staining for type II collagen.
Results We have observed the conditions of the growth of the human epiphyseal chondrocyte. The test of safrarine“o”staining and Alcian blue shows positive. The test of immunohistochemistry staining for type II collagen shows the expression of II collagen. The test of MTT shows the biological properties of the cells decrease from the 15th day.
Conclusions We have established a method of culture the human epiphyseal chondrocyte , at the same time we have identified that the cells were the human epiphyseal chondrocyte.
目的探索体外培养兔骺板软骨细胞的方法,并对其细胞表型进行初步研究,为骺板组织工程寻找良好的种子细胞。
方法取两周龄,体重250g左右幼兔胫骨上端骺板,经过机械剪切、胰酶、胶原酶的消化获得细胞后,1640培养基体外培养。应用倒置显微镜、透射电镜、Alcian blue染色、蕃红花“O”--亮绿染色,免疫组织化学染色、PCR、MTT等方法,对细胞进行定性、定量检测。
结果在体外观察到了骺板软骨细胞贴壁生长及传代的生长过程,多次传代后增殖能力降低。PCR检测有II型胶原的表达、蕃红花“O”--亮绿染色阳性、Alcian blue染色显示第三代细胞蛋白聚糖含量高,MTT显示第三代细胞生长曲线呈倒“S”型,于第12天出现生长抑制。
结论成功地建立了骺板软骨细胞的体外培养方法,并证实所培养出的细胞是骺板软骨细胞。
二、组织工程骺板的构建——兔骺板软骨细胞分别与多孔聚乳酸、多孔丝素构建两种组织工程骺板并比较其体外相容性
目的探索兔骺板软骨细胞与生物支架材料复合构建组织工程骺板的可行性。
方法将体外培养的第三代兔骺板软骨细胞分别与多孔聚乳酸材料、多孔丝素材料复合,构建两种组织工程骺板。应用扫描电镜、激光共聚焦显微镜、MTT等方法,对细胞与材料的相容性进行观察,并对两种生物支架材料进行比较。
结果兔骺板软骨细胞在支架材料上生长良好,随着培养时间的延长,分裂增殖的细胞逐渐增多。细胞在两种材料上生长情况类似,统计学显示无显著性差异。
结论多孔聚乳酸材料和多孔丝素材料均具有良好的生物相容性,可以作为骺板组织工程理想的载体材料。
三、组织工程骺板治疗骺板损伤的动物实验研究——两种组织工程骺板分别治疗骺板损伤并对其进行比较
目的探索组织工程骺板在治疗骺板损伤中的作用。
方法将两种组织工程骺板分别植入骺板损伤的模型中,同时设单纯植入材料组、空白对照组。应用组织学、BrdU细胞示踪、X线等方法,观察成角畸形和短缩畸形情况。同时对两种组织工程骺板、以及两种支架材料进行比较。
结果空白组,骨桥明显,短缩畸形和成角畸形显著;单纯植入支架材料组,短缩畸形和成角畸形明显;组织工程骺板组,存在较小的短缩畸形和成角畸形。统计学显示三种方法存在显著性差异,两种组织工程骺板之间,两种支架材料之间无显著性差异。
结论组织工程骺板治疗骺板损伤是一种有效的方法,其疗效优于单纯植入生物支架材料。
四、人骨骺软骨细胞体外培养及鉴定的初步研究
目的建立体外培养及鉴定人骨骺软骨细胞的方法。
方法取人多指切除手术之多指骨骺,先行机械剪切,后用胰蛋白酶及II型胶原酶消化,再接种于1640培养基中体外培养。应用蕃红花“O”--亮绿染色,Alcian blue染色,免疫组化II型胶原染色和MTT检测,对其生物学特性进行了观察和鉴定。
结果在体外观察到了人骨骺软骨细胞贴壁生长及传代的生长过程,多次传代后增殖能力降低。蕃红花“O”--亮绿染色阳性。Alcian blue染色阳性。免疫组化II型胶原染色阳性。MTT显示生长曲线呈倒“S”型,至第15天细胞出现生长抑制。
结论成功地建立了人骨骺软骨细胞的体外培养方法,并证实所培养出的细胞是人骨骺软骨细胞。
Part I: The seed cells of epiphyseal plate tissue engineering -- the methods of culture the epiphyseal plate chondrocyte in vitro and its cellular phenotype in rabbits.
Objective To establish a kind of method about culture the epiphyseal plate chondrocyte in vitro, so that we can obtain the seed cells for the useage of epiphyseal plate tissue engineering.
Methods The cells we used come from proximal epiphyseal plate of tibia of rabbits. Before we culture the cells, we used Trypsin and Collagenese II . Checking up biological characteristics of the cells by applying the ways of microscope and MTT test. Identifying the cells by applying the ways of safrarine“o”staining and PCR test.
Results We have observed the conditions of the growth of the epiphyseal plate chondrocyte. The test of safrarine“o”staining shows positive. The test of PCR shows the expreesion of II collagen. The test of MTT shows the biological properties of the cells decrease from the 12th day.
Conclusions We have established a method of culture the epiphyseal plate chondrocyte, at the same time we have identified that the cells were the epiphyseal plate chondrocyte.
Part II: The construction of engineering epiphyseal plate ----the study on the biocompatibility of epiphyseal plate chondrocyte with scaffold PLA and epiphyseal plate chondrocyte with scaffold SF.
Objective To explore the way of construction about engineering epiphyseal plate using epiphyseal plate chondrocyte and scaffold.
Methods To construct two engineering epiphyseal plates by culturing the epiphyseal plate chondrocyte on scaffold PLA and scaffold SF. Checking up biological characteristics of the cells by applying the ways of scanning electron- microscope, laser scan confocal microscope and MTT test.
Results The epiphyseal plate chondrocyte grew well on scaffold PLA and scaffold SF. The function of the tow scaffold is similar.
Conclusions Both scaffold PLA and scaffold SF have good biocompatibility and can be used as ideal scaffold in tissue engineering.
Part III: The animal experimental study on the application in epiphyseal plate injury of the two engineering epiphyseal plates.
Objective To observe the role of engineering epiphyseal plate in the treatment of epiphyseal plate injury.
Methods The two engineering epiphyseal plates were implanted into the model of epiphyseal plate injury. Evaluating the role by histology, BrdU and X-ray. At the same time, comparing engineering epiphyseal plate to scaffold .
Results The degree of deformity in engineering epiphyseal plate group was less that scaffold group ,whereas the degree of deformity in scaffold group less than control group .The two engineering epiphyseal plate groups are similar , and the two scaffold groups are also similar.
Conclusions It is feasible to treat epiphyseal plate injury by engineering epiphyseal plate, and it is better than by scaffold.
PartⅣ: The methods of culture and identification of the human epiphyseal chondrocyte.
Objective To establish a kind of method about culture and identification of the human epiphyseal chondrocyte.
Methods The cells we used come from epiphysis of polydactyly. Before we culture the cells, we used Trypsin and Collagenase II . Checking up biological characteristics of the cells by applying the ways of MTT test. Identifying the cells by applying the ways of safrarine“o”staining, Alcian blue test and immuno- histochemistry staining for type II collagen.
Results We have observed the conditions of the growth of the human epiphyseal chondrocyte. The test of safrarine“o”staining and Alcian blue shows positive. The test of immunohistochemistry staining for type II collagen shows the expression of II collagen. The test of MTT shows the biological properties of the cells decrease from the 15th day.
Conclusions We have established a method of culture the human epiphyseal chondrocyte , at the same time we have identified that the cells were the human epiphyseal chondrocyte.
引文
[1]杨万波、顾洁夫等.关节软骨组织工程学研究进展.中国矫形外科杂志,2003,11(19):1387-1388
[2]李卿、刘伟、曹谊林等.爱茜蓝法检测组织工程化软骨及软骨细胞中蛋白聚糖的含量.中华实验外科杂志. 2003,20(1):89
[3] A. Vats, N. S. Tolley, L. D. K. Buttery, J. M. Polak. The stem cell in orthopaedic surgery. J Bone Joint Surg [Br]. 2004;86-B:159-64
[4] Li Q, Lu JH, Liu Wz ,et al. The effect of aging on aggrecan catabolism in cultured chondrocytes . Zhongguo Linchuang Kangfu. 2003; 7(2):216-7
[5]戴刚、李起鸿、周强等.相关生长因子对体外培养关节软骨细胞促增殖作用的比较研究.中华创伤杂志,2002,18:235-238.
[6]何清义、李起鸿、许建中等.人关节软骨细胞的永生化及其表型诱导.中华创伤杂志. 2002 ,18:727-731.
[7] Zhang ZW, Yang CD,Jin JJ ,et al. Maintain the characteristic phenolype of the rabbits articular chondrocytes after culture in alginate bead. Zhongguo Linchuang Kangfu 2003;7(4):556-7
[8] He QY, Li QH, Xu Jz ,et al. Immortalization of human articular chondrocytes and induction of their phenotypes. Zhongguo Linchuang Kangfu Zazhi 2002;18(12):727-31
[9] He QY, Li QH, Xu Jz ,et al. Comparing biological characteristics betweentransformed human chondrocytes and normal chondrocytes. Zhongguo Linchuang Kangfu 2002;6(10):1414-5
[10] Kawada K ,Yonei T ,Veoka H ,et al. Comparison of the mosesitivity test :donogenic assay versus MTT assay[J ]. Acta Medokayama , 2002 , 56 (3) :1291
[11] Noguchi K, Iwahashi M, Tani M, et al . Evaluation of hemosensitivity testing with highly purified tumor cells in 435 patients with gastric caicinoma using an MTT assay[J ] . Anticancer Res , 2005 ,25 (2A) :931 - 937.
[12]邓丹、武晓莉、许锋等. MTT法检测国产PGA支架对人成纤维细胞增殖的影响.组织工程与重建外科杂志. 2006;2(1):18-20
[1]颜文龙、孙恩杰、郭海英等.组织工程支架材料.上海生物医学工程. 2004 ,25(1):51～54
[2] Tal D , Orna TG,et al.“Designer”Scaffolds for Tissue Engineering and Regeneration[J ] . Israel Journal of Chemistry ,2005 ,45 :487～494
[3]方佩斐、贾维敏.组织工程中天然支架材料的研究现状.医学文选.2006,25(4):901-903
[4] Burg KJ, Holder WD, Culberson CR, et al. Parameters affecting cellular adhesion to polylactide films. Journal of Biomaterials Science, 1999, 10(2):147-161
[5] Krylova E, Ivanov A, Orlovski V, et al. J Mater Med , 2002 ; 13 (1) : 87 - 90
[6] Santos C, Luklinska ZB, Clarke RL, et al. J Mater Med, 2001;12 (3) : 565-573
[7] Queiroz AC, Santos JD, Monteiro FJ, et al. Biomaterials, 2001; 22(11):1393-1400
[8]宁丽、薛淼、黄海宁等.皮肤再生膜的生物相容性系列研究.中国修复重建外科杂志. 2000 ,14(1):44.
[9]吴徵宇.丝素蛋白作为生物医用材料的研究.材料导报. 2000 ,14(9):63.
[10] Yongzhong Wang , Ung-J ,K et al. In vitro cartilage tissue engineering with 3D porous aqueous derived silk scaffolds and mesenchymal stem cells [J ] . Biomaterials ,2005, (26) :7082～7094
[11]吴海涛、钟翠平、顾云娣.蚕丝在软骨细胞立体培养中的应用.组织工程. 2000,14(5):301 - 304.
[12]张亚、王晓东、张希峰等.兔骨髓间充质干细胞在丝素蛋白上的体外培养.中华小儿外科杂志. 2006,27(1):34-37
[13] Altman G H ,Diaz F ,Jakuba C ,et al. Silk - based biomaterials [J ] . Biomaterials , 2003 ,24 (3) :401 - 406.
[14]张真、卢晓凤.生物材料有效性和安全性评价的现状与趋势.生物医学工程学杂志. 2002, 19(1): 117-121
[15] Kue R, Sohrabi A, Nagle D,et al. Enhanced proliferation and osteocalcin production by human osteoblast-like MG63 cells om silicon nitride ceramic discs. Biomaterials, 1999;20(13):1195-1199
[16] Metzler V, Bienert H, Lehmann T, et al. A novel method for quantifying shape deformation applied to biocompatibility testing. A SAIO J,1999;45(4):264-271
[1] Wattenbarger JM, Gruber He , Phieffer LS.Physeal fractures , part I : histologic features of bone , cartilage , and bar formation in a small ani- mal model [J ] . J Pediatr Orthop ,2002 , 22 (6) : 703 - 9
[2]王仁法、刘运强等.导致儿童骺早闭的病因探讨研究.放射学实践. 2001;16(2):127-129
[3] Kay RM , Matthys GA . Pediatric ankle fractures : evaluation and treatment [J ] . J Am Acad Orthop Surg , 2001 , 9 : 268 - 278
[4]石峰、陈秋、刘卫东等.不同直径骺板贯穿伤对骺板发育影响的实验研究.中华小儿外科杂志,2002,2(23):53-55
[5] Xian CJ , Zhou FH , McCarty RC , et al . Intramembranous ossifycation mechanism for bone bridge formation at the growth plate cartilage injury site [J ] . J Ort hop Res , 2004 , 22 (2) :417-426
[6] Bruce KF , John B , Halster C. Free fat interpositional graft in acute physeal injuries : the anticipatory langenskiǒld procedure. J Pedia Orthop , 2000 , 20 :282～285
[7] Cheon J E , Kim IO , Kim CJ , et al . Imaging findings after fat graft interposition in an injured growt h plate : an experimental study in rabbits [ J ] . Invest Radiol , 2003 , 38 ( 11 ) : 695-703
[8]许瑞江、吴战坡、顾章平等.自体髂嵴骨骺游离移植重建内踝缺损初步报告.临床小儿外科杂志,2006,5(3);175-176
[9]尹飞、王金成、郭丽等.自体髂骨骺板移植重建胫骨骺板.骨与关节损伤杂志. 2003,18(4);221-222
[10] Mayr JM, Pierer GR, Linhart WE. Reconstruction of part of the distal tibial growth plate with an autologous graft from the iliac crest [J]. J bone Jone Surg, 2000, 82B(4):558-560
[11]王树峰、吕占辉、王明山等.带血供腓骨小头骨骺移植的解剖与临床研究.中华骨科杂志.2001;21(2):99
[12]王恩波、赵群、马瑞雪等.不同自体移植物防止骺板损伤后骨桥形成的实验研究.中华小儿外科杂志. 2001; 22(4): 235-237
[13] Oostendorp R A,Audet J, Eaves C J. High-resolution tracking of cell division suggests similar cell cycle kinetics of hematopoietic stem cells stimulated in vitro and in vivo[ J ]. Blood, 2000, 95 (3) : 8552862.
[14]郭启仓、刘志勇、李占清等. 5 -溴脱氧尿嘧啶核苷标记骨髓间充质干细胞的技术探讨.华北煤炭医学院学报. 2004,6(1);4--6
[15] Doig SG. The management of femoral and tibial diaphyseal fractures. J R Coll Surg Edinb , 2000, 45:204
[16] Chen , Fen MBBS; Hui , James H. P. FRCS. Cultured Mesenchymal Stem Cell Transfers in the Treatment of Partial Growth Arrest . J Pedia Orthop , 2003 , 23 : 425～429
[1]曹谊林、周广东. 21世纪组织工程面临的机遇与挑战.中华医学杂志. 2005;85(36):2523-2525
[2] Kono T, Obata Y, W u Q ,et al. Birth of parthenogenetic mice that can develop to adulthood. Nature, 2004, 428 ( 6985) : 860-864.
[3]裴雪涛、刘大庆.干细胞:组织器官重建的理想种子细胞.中国修复重建外科杂志. 2006;20(4):344-348
[4] Kawada K ,Yonei T ,Veoka H ,et al. Comparison of the mosesitivity test :donogenic assay versus MTT assay[J ]. Acta Medokayama , 2002 , 56 (3) :1291
[5] Noguchi K, Iwahashi M, Tani M, et al . Evaluation of hemosensitivity testing with highly purified tumor cells in 435 patients with gastric caicinoma using an MTT assay[J ] . Anticancer Res , 2005 ,25 (2A) :931 - 937.
[6]邓丹、武晓莉、许锋等. MTT法检测国产PGA支架对人成纤维细胞增殖的影响.组织工程与重建外科杂志. 2006;2(1):18-20
[7] Li Q, Lu JH, Liu Wz ,et al. The effect of aging on aggrecan catabolism in cultured chondrocytes . Zhongguo Linchuang Kangfu 2003; 7(2):216-7
[8] Lavik E, Langer R.Tissue engineering: current state and perspectives. ApplMicrobiol Biotechnol, 2004, 65: 1-8.
[9] Shieh SJ, Vacanti JP. State-of-the-art tissue engineering: from tissue engineering to organ building. Surgery, 2005, 137: 127.
[10] Levenberg S, Langer R. Advances in tissue engineering. Curr Top Dev Biol, 2004, 61: 113-134.)
[1] OO i. The exp ression of the nuclear oncogenes c-myc and c-jun in the groove of Ranvier of the rabbit growth plate. A fr J Med MedSci ( African journal of medicine and medical sciences). 2002, 31(4) : 325～327
[2]王仁法、刘运强、导致儿童骺早闭的病因探讨研究.放射学实践. 2001;16(2):127-129
[3]屈辉.儿童骺软骨骨折影像学诊断的进展.中华放射学杂志. 2001, 35: 867～869
[4]詹新立、周江南.骺板损伤的诊断与治疗的新进展.临床小儿外科杂志. 2003;2(2):115-118
[5] Wattenbarger JM , Gruber HE, Ph ie. er L S. Physeal fractures,part I: histologic features of bone, cartilage, and bar formation in a small animal model. J Pediatr Orthop, 2002, 22: 703～709.
[6] Cory J. Xian, Fiona H. Zhou, Rosa C. McCarty, Bruce K. Foster Intramembranous ossication mechanism for bone bridge formation at the growth plate cartilage injury site. Journal of Orthopaedic Research, 2004, 22: 417～426
[7]张克勇、蔡林. Ranvier区切除后外骨桥形成的细胞组织学变化.数理医药学杂志. 2006;19(5):477-479
[8] Leung AG, Peterson HA . Fractures of the proximal radial head and neckin children with emphasis on those that involve the articular artilage (Review) . J Pediatr Orthop , 2000 ,20 :7-14.
[9] Jaramillo D ,Kammen BF ,Shapiro F.Cartilaginous path of physeal fracture-separations : evalution with MR imaging:an experimental study with histologic correlation in rabbits [J] .Radiology, 2000, 215 (2): 504-511
[10] Lohman M , Kivisaari A ,Vehmas T ,et al. MRI in the assessment of growth arrest [J ] . Pediatr Radiol ,2002 ,33 (1) :41-45.
[11] Breburda E,Wirth T ,Leiser R ,et al. The influence of intermittent ex- ternal dynamic pressure and tension forces on the healing of an epiphys- eal fracture [J]. Arch Orthop Trauma Surg, 2001 ,121(8) :443-449.
[12] Lalaji A ,Umans H , Schneider R ,et al. MRI features of confirmed“pre-slip”capital femoral epiphyses : a report of two cases [ J ] . Skeletal Radiol ,2002 ,31 (6) :362-365.
[13] Futmai T ,Suzuki S ,Seto Y,et al. Sequential magnetic resonance imaging in slipped capital femoral epiphysis : assessment of preslip in the concralateral hip[J ] .J Pediatr Orthop B ,2001 ,10(4) :298-303.
[14] Varich LJ , Laor T , Jaramillo D. Normal maturation of the distal femoral epiphyseal cartilage : age-related changes at MR imaging [J ] . Radiology ,2000 , 214 (3) : 705-709.
[15]马瑞雪、吉士俊、陈秋.骨桥切除、脂肪填塞、截骨矫正治疗小儿骨骺早闭的临床观察.中华小儿外科杂志. 2001;22(2):73-75
[16]李五洲、刘克斌、张记恩.骨桥切除、脂肪填塞与截骨矫形治疗小儿骨骺早闭15例.骨与关节损伤杂志. 2004;19(5):349-350
[17]许瑞江、吴战坡、顾章平.自体髂嵴骨骺游离移植重建内踝缺损初步报告.临床小儿外科杂志. 2006;5(3):175-176
[18]尹飞、王金成、郭丽等.自体髂骨骺板移植重建胫骨骺板.骨与关节损伤杂志. 2003;18(4):221-222
[19] Mayr JM, Pierer GR, Linhart WE. Reconstruction of part of the distal tibial growth plate with an autologous graft from the iliac crest [J]. J bone Jone Surg, 2000, 82B(4): 558- 560.
[20] Glickman AM, Yang JP, Stevens DG, et al. Epiphyseal plate transplantation between sites of different growth potential. Pediatr Orthop, 2000 ; 20(3): 289
[21]王恩波、赵群、马瑞雪等.不同自体移植物防止骺板损伤后骨桥形成的实验研究.中华小儿外科杂志. 2001;22(4): 235-237
[22]杜晓冰,勇华,区士欢等.骺板移植的实验形态学观察.中华临床解剖杂志. 1995;13(2):297
[23]王树峰,吕占辉,王明山等.带血供腓骨小头骨骺移植的解剖与临床研究.中华骨科杂志. 2001;21(2):99
[24]周强、李起鸿、戴刚等.高数量骺板软骨细胞聚集培养生成软骨组织的观察.第三军医大学学报. 2002,24(5):556-558.
[25] Ballock RT, Zhou X, Mink LM, et al. Both retinoic acid and 1,25 (OH)2 vitamin D3 inhibitthyroid hormone induced terminal differentiaton of growth plate chondrocytes [J]. J Orthop Res, 2001 ,19 (1): 43-49.
[26] Ballock RT ,Zhou X, Mink LM, et al.Expression of cyclin-dependent Kinase inhibitors in epiphyseal chondrocytes induced to terminally differentiate with thyroid hormone [J]. Endocrinology, 2000, 141 (12) :4552-4557.
[27] Krylova E, Ivanov A, Orlovski V, et al. J Mater Med, 2002; 13 (1) : 87 - 90
[28] Santos C, Luklinska ZB, Clarke RL, et al. J Mater Med,2001;12 (3) : 565-573
[29]张鲁华、杨飞、公衍道等.壳聚糖和明胶材料对血管平滑肌细胞的作用.生物物理学报. 2004 ,20 (1) :19～24
[30]陈光、周平、杨芳等.天然生物材料壳聚糖支架上人胚肺成纤维细胞的生长.化学学报.2004 ,62 (10) :992～997
[31]宁丽、薛淼、黄海宁等.皮肤再生膜的生物相容性系列研究.中国修复重建外科杂志,2000 ,14(1):44.
[32]吴徵宇.丝素蛋白作为生物医用材料的研究.材料导报. 2000, 14 (9) : 63.
[33]刘广鹏、崔磊、曹谊林.软骨生长板基因调控机制的研究进展.组织工程与重建外科杂志. 2006;2(2):110-112
[34] Jurgen W, Silvia R, Martina S. Four distinct chondrocyte populations in the fetal bovine growth plate: highest exp ression levels of PTH / PTHrp receptor, Indian hedgehog, and MMP-13 in hypertrophic chondro- cytes and their suppression by PTH (1～34) and PTHrp ( 1～40) [ J ]. Exp Cell Res, 2002, 279 (1) : 1-13.
[35] Colin F, David J, Elaine S, et al. Regulation of chondrocyte terminal differentiation in the postembryonic growth p late: the role of the PTHrp- Indian hedgehog axis[ J ]. Endocrinology, 2001, 142 (9) : 4131-4140.
[36] OrnitzDMand Marie PJ. FGF signaling pathways in endochondral and intramembranous bone development and human genetic disease. Genes Dev. 2002, 16:1446- 1465.
[37] Minina E, Ornitz DM and Vortkamp A. BMP and Ihh/PTHrP signal- ing interact to coordinate chondrocytes proliferation and differentiation. Development, 2001, 128: 4523- 4534.
[38]黄仕龙、陈安民、郭风劲等.鼠骨骺增殖区细胞的分离鉴定和克隆构建PTHrp基因真核表达载体.中国矫形外科杂志. 2005;13(19):1489-1491
[2]李卿、刘伟、曹谊林等.爱茜蓝法检测组织工程化软骨及软骨细胞中蛋白聚糖的含量.中华实验外科杂志. 2003,20(1):89
[3] A. Vats, N. S. Tolley, L. D. K. Buttery, J. M. Polak. The stem cell in orthopaedic surgery. J Bone Joint Surg [Br]. 2004;86-B:159-64
[4] Li Q, Lu JH, Liu Wz ,et al. The effect of aging on aggrecan catabolism in cultured chondrocytes . Zhongguo Linchuang Kangfu. 2003; 7(2):216-7
[5]戴刚、李起鸿、周强等.相关生长因子对体外培养关节软骨细胞促增殖作用的比较研究.中华创伤杂志,2002,18:235-238.
[6]何清义、李起鸿、许建中等.人关节软骨细胞的永生化及其表型诱导.中华创伤杂志. 2002 ,18:727-731.
[7] Zhang ZW, Yang CD,Jin JJ ,et al. Maintain the characteristic phenolype of the rabbits articular chondrocytes after culture in alginate bead. Zhongguo Linchuang Kangfu 2003;7(4):556-7
[8] He QY, Li QH, Xu Jz ,et al. Immortalization of human articular chondrocytes and induction of their phenotypes. Zhongguo Linchuang Kangfu Zazhi 2002;18(12):727-31
[9] He QY, Li QH, Xu Jz ,et al. Comparing biological characteristics betweentransformed human chondrocytes and normal chondrocytes. Zhongguo Linchuang Kangfu 2002;6(10):1414-5
[10] Kawada K ,Yonei T ,Veoka H ,et al. Comparison of the mosesitivity test :donogenic assay versus MTT assay[J ]. Acta Medokayama , 2002 , 56 (3) :1291
[11] Noguchi K, Iwahashi M, Tani M, et al . Evaluation of hemosensitivity testing with highly purified tumor cells in 435 patients with gastric caicinoma using an MTT assay[J ] . Anticancer Res , 2005 ,25 (2A) :931 - 937.
[12]邓丹、武晓莉、许锋等. MTT法检测国产PGA支架对人成纤维细胞增殖的影响.组织工程与重建外科杂志. 2006;2(1):18-20
[1]颜文龙、孙恩杰、郭海英等.组织工程支架材料.上海生物医学工程. 2004 ,25(1):51～54
[2] Tal D , Orna TG,et al.“Designer”Scaffolds for Tissue Engineering and Regeneration[J ] . Israel Journal of Chemistry ,2005 ,45 :487～494
[3]方佩斐、贾维敏.组织工程中天然支架材料的研究现状.医学文选.2006,25(4):901-903
[4] Burg KJ, Holder WD, Culberson CR, et al. Parameters affecting cellular adhesion to polylactide films. Journal of Biomaterials Science, 1999, 10(2):147-161
[5] Krylova E, Ivanov A, Orlovski V, et al. J Mater Med , 2002 ; 13 (1) : 87 - 90
[6] Santos C, Luklinska ZB, Clarke RL, et al. J Mater Med, 2001;12 (3) : 565-573
[7] Queiroz AC, Santos JD, Monteiro FJ, et al. Biomaterials, 2001; 22(11):1393-1400
[8]宁丽、薛淼、黄海宁等.皮肤再生膜的生物相容性系列研究.中国修复重建外科杂志. 2000 ,14(1):44.
[9]吴徵宇.丝素蛋白作为生物医用材料的研究.材料导报. 2000 ,14(9):63.
[10] Yongzhong Wang , Ung-J ,K et al. In vitro cartilage tissue engineering with 3D porous aqueous derived silk scaffolds and mesenchymal stem cells [J ] . Biomaterials ,2005, (26) :7082～7094
[11]吴海涛、钟翠平、顾云娣.蚕丝在软骨细胞立体培养中的应用.组织工程. 2000,14(5):301 - 304.
[12]张亚、王晓东、张希峰等.兔骨髓间充质干细胞在丝素蛋白上的体外培养.中华小儿外科杂志. 2006,27(1):34-37
[13] Altman G H ,Diaz F ,Jakuba C ,et al. Silk - based biomaterials [J ] . Biomaterials , 2003 ,24 (3) :401 - 406.
[14]张真、卢晓凤.生物材料有效性和安全性评价的现状与趋势.生物医学工程学杂志. 2002, 19(1): 117-121
[15] Kue R, Sohrabi A, Nagle D,et al. Enhanced proliferation and osteocalcin production by human osteoblast-like MG63 cells om silicon nitride ceramic discs. Biomaterials, 1999;20(13):1195-1199
[16] Metzler V, Bienert H, Lehmann T, et al. A novel method for quantifying shape deformation applied to biocompatibility testing. A SAIO J,1999;45(4):264-271
[1] Wattenbarger JM, Gruber He , Phieffer LS.Physeal fractures , part I : histologic features of bone , cartilage , and bar formation in a small ani- mal model [J ] . J Pediatr Orthop ,2002 , 22 (6) : 703 - 9
[2]王仁法、刘运强等.导致儿童骺早闭的病因探讨研究.放射学实践. 2001;16(2):127-129
[3] Kay RM , Matthys GA . Pediatric ankle fractures : evaluation and treatment [J ] . J Am Acad Orthop Surg , 2001 , 9 : 268 - 278
[4]石峰、陈秋、刘卫东等.不同直径骺板贯穿伤对骺板发育影响的实验研究.中华小儿外科杂志,2002,2(23):53-55
[5] Xian CJ , Zhou FH , McCarty RC , et al . Intramembranous ossifycation mechanism for bone bridge formation at the growth plate cartilage injury site [J ] . J Ort hop Res , 2004 , 22 (2) :417-426
[6] Bruce KF , John B , Halster C. Free fat interpositional graft in acute physeal injuries : the anticipatory langenskiǒld procedure. J Pedia Orthop , 2000 , 20 :282～285
[7] Cheon J E , Kim IO , Kim CJ , et al . Imaging findings after fat graft interposition in an injured growt h plate : an experimental study in rabbits [ J ] . Invest Radiol , 2003 , 38 ( 11 ) : 695-703
[8]许瑞江、吴战坡、顾章平等.自体髂嵴骨骺游离移植重建内踝缺损初步报告.临床小儿外科杂志,2006,5(3);175-176
[9]尹飞、王金成、郭丽等.自体髂骨骺板移植重建胫骨骺板.骨与关节损伤杂志. 2003,18(4);221-222
[10] Mayr JM, Pierer GR, Linhart WE. Reconstruction of part of the distal tibial growth plate with an autologous graft from the iliac crest [J]. J bone Jone Surg, 2000, 82B(4):558-560
[11]王树峰、吕占辉、王明山等.带血供腓骨小头骨骺移植的解剖与临床研究.中华骨科杂志.2001;21(2):99
[12]王恩波、赵群、马瑞雪等.不同自体移植物防止骺板损伤后骨桥形成的实验研究.中华小儿外科杂志. 2001; 22(4): 235-237
[13] Oostendorp R A,Audet J, Eaves C J. High-resolution tracking of cell division suggests similar cell cycle kinetics of hematopoietic stem cells stimulated in vitro and in vivo[ J ]. Blood, 2000, 95 (3) : 8552862.
[14]郭启仓、刘志勇、李占清等. 5 -溴脱氧尿嘧啶核苷标记骨髓间充质干细胞的技术探讨.华北煤炭医学院学报. 2004,6(1);4--6
[15] Doig SG. The management of femoral and tibial diaphyseal fractures. J R Coll Surg Edinb , 2000, 45:204
[16] Chen , Fen MBBS; Hui , James H. P. FRCS. Cultured Mesenchymal Stem Cell Transfers in the Treatment of Partial Growth Arrest . J Pedia Orthop , 2003 , 23 : 425～429
[1]曹谊林、周广东. 21世纪组织工程面临的机遇与挑战.中华医学杂志. 2005;85(36):2523-2525
[2] Kono T, Obata Y, W u Q ,et al. Birth of parthenogenetic mice that can develop to adulthood. Nature, 2004, 428 ( 6985) : 860-864.
[3]裴雪涛、刘大庆.干细胞:组织器官重建的理想种子细胞.中国修复重建外科杂志. 2006;20(4):344-348
[4] Kawada K ,Yonei T ,Veoka H ,et al. Comparison of the mosesitivity test :donogenic assay versus MTT assay[J ]. Acta Medokayama , 2002 , 56 (3) :1291
[5] Noguchi K, Iwahashi M, Tani M, et al . Evaluation of hemosensitivity testing with highly purified tumor cells in 435 patients with gastric caicinoma using an MTT assay[J ] . Anticancer Res , 2005 ,25 (2A) :931 - 937.
[6]邓丹、武晓莉、许锋等. MTT法检测国产PGA支架对人成纤维细胞增殖的影响.组织工程与重建外科杂志. 2006;2(1):18-20
[7] Li Q, Lu JH, Liu Wz ,et al. The effect of aging on aggrecan catabolism in cultured chondrocytes . Zhongguo Linchuang Kangfu 2003; 7(2):216-7
[8] Lavik E, Langer R.Tissue engineering: current state and perspectives. ApplMicrobiol Biotechnol, 2004, 65: 1-8.
[9] Shieh SJ, Vacanti JP. State-of-the-art tissue engineering: from tissue engineering to organ building. Surgery, 2005, 137: 127.
[10] Levenberg S, Langer R. Advances in tissue engineering. Curr Top Dev Biol, 2004, 61: 113-134.)
[1] OO i. The exp ression of the nuclear oncogenes c-myc and c-jun in the groove of Ranvier of the rabbit growth plate. A fr J Med MedSci ( African journal of medicine and medical sciences). 2002, 31(4) : 325～327
[2]王仁法、刘运强、导致儿童骺早闭的病因探讨研究.放射学实践. 2001;16(2):127-129
[3]屈辉.儿童骺软骨骨折影像学诊断的进展.中华放射学杂志. 2001, 35: 867～869
[4]詹新立、周江南.骺板损伤的诊断与治疗的新进展.临床小儿外科杂志. 2003;2(2):115-118
[5] Wattenbarger JM , Gruber HE, Ph ie. er L S. Physeal fractures,part I: histologic features of bone, cartilage, and bar formation in a small animal model. J Pediatr Orthop, 2002, 22: 703～709.
[6] Cory J. Xian, Fiona H. Zhou, Rosa C. McCarty, Bruce K. Foster Intramembranous ossication mechanism for bone bridge formation at the growth plate cartilage injury site. Journal of Orthopaedic Research, 2004, 22: 417～426
[7]张克勇、蔡林. Ranvier区切除后外骨桥形成的细胞组织学变化.数理医药学杂志. 2006;19(5):477-479
[8] Leung AG, Peterson HA . Fractures of the proximal radial head and neckin children with emphasis on those that involve the articular artilage (Review) . J Pediatr Orthop , 2000 ,20 :7-14.
[9] Jaramillo D ,Kammen BF ,Shapiro F.Cartilaginous path of physeal fracture-separations : evalution with MR imaging:an experimental study with histologic correlation in rabbits [J] .Radiology, 2000, 215 (2): 504-511
[10] Lohman M , Kivisaari A ,Vehmas T ,et al. MRI in the assessment of growth arrest [J ] . Pediatr Radiol ,2002 ,33 (1) :41-45.
[11] Breburda E,Wirth T ,Leiser R ,et al. The influence of intermittent ex- ternal dynamic pressure and tension forces on the healing of an epiphys- eal fracture [J]. Arch Orthop Trauma Surg, 2001 ,121(8) :443-449.
[12] Lalaji A ,Umans H , Schneider R ,et al. MRI features of confirmed“pre-slip”capital femoral epiphyses : a report of two cases [ J ] . Skeletal Radiol ,2002 ,31 (6) :362-365.
[13] Futmai T ,Suzuki S ,Seto Y,et al. Sequential magnetic resonance imaging in slipped capital femoral epiphysis : assessment of preslip in the concralateral hip[J ] .J Pediatr Orthop B ,2001 ,10(4) :298-303.
[14] Varich LJ , Laor T , Jaramillo D. Normal maturation of the distal femoral epiphyseal cartilage : age-related changes at MR imaging [J ] . Radiology ,2000 , 214 (3) : 705-709.
[15]马瑞雪、吉士俊、陈秋.骨桥切除、脂肪填塞、截骨矫正治疗小儿骨骺早闭的临床观察.中华小儿外科杂志. 2001;22(2):73-75
[16]李五洲、刘克斌、张记恩.骨桥切除、脂肪填塞与截骨矫形治疗小儿骨骺早闭15例.骨与关节损伤杂志. 2004;19(5):349-350
[17]许瑞江、吴战坡、顾章平.自体髂嵴骨骺游离移植重建内踝缺损初步报告.临床小儿外科杂志. 2006;5(3):175-176
[18]尹飞、王金成、郭丽等.自体髂骨骺板移植重建胫骨骺板.骨与关节损伤杂志. 2003;18(4):221-222
[19] Mayr JM, Pierer GR, Linhart WE. Reconstruction of part of the distal tibial growth plate with an autologous graft from the iliac crest [J]. J bone Jone Surg, 2000, 82B(4): 558- 560.
[20] Glickman AM, Yang JP, Stevens DG, et al. Epiphyseal plate transplantation between sites of different growth potential. Pediatr Orthop, 2000 ; 20(3): 289
[21]王恩波、赵群、马瑞雪等.不同自体移植物防止骺板损伤后骨桥形成的实验研究.中华小儿外科杂志. 2001;22(4): 235-237
[22]杜晓冰,勇华,区士欢等.骺板移植的实验形态学观察.中华临床解剖杂志. 1995;13(2):297
[23]王树峰,吕占辉,王明山等.带血供腓骨小头骨骺移植的解剖与临床研究.中华骨科杂志. 2001;21(2):99
[24]周强、李起鸿、戴刚等.高数量骺板软骨细胞聚集培养生成软骨组织的观察.第三军医大学学报. 2002,24(5):556-558.
[25] Ballock RT, Zhou X, Mink LM, et al. Both retinoic acid and 1,25 (OH)2 vitamin D3 inhibitthyroid hormone induced terminal differentiaton of growth plate chondrocytes [J]. J Orthop Res, 2001 ,19 (1): 43-49.
[26] Ballock RT ,Zhou X, Mink LM, et al.Expression of cyclin-dependent Kinase inhibitors in epiphyseal chondrocytes induced to terminally differentiate with thyroid hormone [J]. Endocrinology, 2000, 141 (12) :4552-4557.
[27] Krylova E, Ivanov A, Orlovski V, et al. J Mater Med, 2002; 13 (1) : 87 - 90
[28] Santos C, Luklinska ZB, Clarke RL, et al. J Mater Med,2001;12 (3) : 565-573
[29]张鲁华、杨飞、公衍道等.壳聚糖和明胶材料对血管平滑肌细胞的作用.生物物理学报. 2004 ,20 (1) :19～24
[30]陈光、周平、杨芳等.天然生物材料壳聚糖支架上人胚肺成纤维细胞的生长.化学学报.2004 ,62 (10) :992～997
[31]宁丽、薛淼、黄海宁等.皮肤再生膜的生物相容性系列研究.中国修复重建外科杂志,2000 ,14(1):44.
[32]吴徵宇.丝素蛋白作为生物医用材料的研究.材料导报. 2000, 14 (9) : 63.
[33]刘广鹏、崔磊、曹谊林.软骨生长板基因调控机制的研究进展.组织工程与重建外科杂志. 2006;2(2):110-112
[34] Jurgen W, Silvia R, Martina S. Four distinct chondrocyte populations in the fetal bovine growth plate: highest exp ression levels of PTH / PTHrp receptor, Indian hedgehog, and MMP-13 in hypertrophic chondro- cytes and their suppression by PTH (1～34) and PTHrp ( 1～40) [ J ]. Exp Cell Res, 2002, 279 (1) : 1-13.
[35] Colin F, David J, Elaine S, et al. Regulation of chondrocyte terminal differentiation in the postembryonic growth p late: the role of the PTHrp- Indian hedgehog axis[ J ]. Endocrinology, 2001, 142 (9) : 4131-4140.
[36] OrnitzDMand Marie PJ. FGF signaling pathways in endochondral and intramembranous bone development and human genetic disease. Genes Dev. 2002, 16:1446- 1465.
[37] Minina E, Ornitz DM and Vortkamp A. BMP and Ihh/PTHrP signal- ing interact to coordinate chondrocytes proliferation and differentiation. Development, 2001, 128: 4523- 4534.
[38]黄仕龙、陈安民、郭风劲等.鼠骨骺增殖区细胞的分离鉴定和克隆构建PTHrp基因真核表达载体.中国矫形外科杂志. 2005;13(19):1489-1491