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同种异体组织工程化软骨局部免疫豁免诱导的实验 预览 被引量:6

Tissue engineered allogeneic cartilage induces local immune privilege in rabbits
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摘要 背景:同种异体软骨移植存在局部免疫排斥反应,有必要进一步减少其排斥反应,以促进软骨修复的临床应用,为此进行一系列局部免疫豁免诱导的实验很有意义。 目的:观察FasL基因重组逆转录病毒载体转染软骨细胞形成同种异体组织工程化软骨在体内生长情况。 设计:随机对照观察。 单位:上海交通大学。 材料:选用36只同种异体新西兰兔为受者,45只新生1周龄青紫蓝兔为供者,雌雄均不限。购自中国科学院实验动物研究中心。双嗜性逆转录病毒包装细胞系PT67购自Clontech公司;细胞培养基DMEM、胎牛血清、G418、Polybrene购自GIBCO BRL。 方法:实验于2000-01/2005-07在原上海第二医科大学完成。摸球法随机将新西兰兔分为3组:同种异体软骨细胞-FasL组、同种异体软骨细胞组及空白组。每组各12只。利用重组逆转录病毒载体转染的青紫蓝兔软骨细胞和组织工程材料pluronic F-127复合物构建兔同种异体组织工程软骨。①移植物观察其大体观察与质量变化:各组均将相应材料注入新西兰白兔的皮下,移植接种后1,2,3个月取出移植物观察其大体观察与质量变化。②染色观察:移植接种后1,2,3月取出移植物,切片,行苏木精-伊红、Safranin'O、Masson's trichrome染色观察。③抗体检测:移植接种后1,2月抽血1mL,将青紫蓝兔的软骨细胞用裂解液冻融裂解作为混合抗原,于琼脂糖介质中电泳分离,与受者新西兰兔血清作用,观察有无相应抗体产生。④补体依赖细胞毒实验:将青紫蓝兔的软骨细胞制备成2×10^9L^-1的细胞悬液,接种于96孔板上,贴壁生长24h,加受者新西兰兔血清进行细胞毒试验,在显微镜下计数凋亡细胞的百分率。 主要观察指标:①两组移植物的大体观察与质量变化。②组织学变化。③抗体检测结果。④凋亡细胞的百分率。 结果:纳入实验兔81只均进入结果分析 BACKGROUND: The transplantation of aIIogeneic cartilage has local immunological rejection, and it is necessary to further reduce the rejection to promote its application in clinic, thus it is significant to perform a series of experiments to induce local immune privilege. OBJECTIVE: To observe the in vivo growth of tissue engineered allogeneic cartilage reconstructed by chondrocytes transfected with recombinant retroviral vector pLNCX2-FasL. DESIGN : A randomized controlled observation. SETTING : Shanghai Jiao Tong University. MATERIALS : Thirty-six allogeneic New Zealand rabbits as recipients and 45 1-week-old chinchillas as donors, either sex, were purchased by the experimental animal center of Chinese Academy of Sciences. Amphotropic recombinant retrovirus coated cell line PT67 was purchased from Clontech Company; Dulbecco's modified eagle medium (DMEM), fetal bovine serum (FBS), G418 and Polybrene were bought from GIBCO BRL. METHODS: The experiment was carried out in original Shanghai Second Medical University from January 2000 to July 2005. The New Zealand rabbits were randomly divided into three groups: FasL-transfected group (n =12), untransfected group (n =12) and blank control group (n =12). The rabbit allogeneic cartilages were constructed by the compound of pLNCX2-FasL transfected chondrocytes and tissue engineered material of pluronic F-127. ① Gross observation and mass changes of the grafts: Corresponding materials were infused subcutaneously, the grafts were removed at 1, 2 and 3 months after transplantation for gross observation and the mass changes. ② Staining observation: The grafts were removed at 1, 2 and 3 months after transplantation, then prepared into sections, and observed by hematoxylin and eosin (HE), Safranin'O and Masson's trichrome stainings. ③ Antibody detection: Blood samples (1 mL) were collected at 1 and 2 months after transplantation, the chondrocytes of the chinchillas were lysed freezingly with lysis antigen as
作者 胡洪亮 曹谊林 陈婷婷 樊绮诗 胡翊群 Hu Hong-liang, Cao Yi-lin, Chen Ting-ting, Fan Qi-shi, Hu Yi-qun(1.Department of Clinical Laboratory, Ruijin Hospital, Medical College of Shanghai Jiao Tong University, Shanghai 200025, China; 2.Key Laboratory of Tissue Engineering, Affiliated Ninth People's Hospital, Medical College of Shanghai Jiao Tong University, Shanghai 200011, China; 3.Department of Internal Medicine, Renji Hospital, Medical College of Shanghai Jiao Tong University, Shanghai 200127. China)
出处 《中国组织工程研究与临床康复》 CAS CSCD 北大核心 2007年第14期 2757-2760,共4页 Journal of Clinical Rehabilitative Tissue Engineering Research
基金 国家重点基础研究发展规划(九七三)资助(G1999054304) 上海交通大学瑞金医院资助项目.
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