自体软骨细胞修复关节软骨缺损的机制探讨  

Mechanisms of autologous chodrocytes mass transplantation in the repair of cartilage defects of rabbits' knee

作　　者：余方圆[1] 卢世璧[2] 黄利虹[2] 许文静[2] 彭江[2] 赵斌[2] 袁玫[2] 黄靖香[2] 

机构地区：[1]解放军309医院骨科,北京100091 [2]解放军总医院骨科研究所,北京100853

出　　处：《中国骨伤》2010年第9期683-687,共5页China Journal of Orthopaedics and Traumatology

基　　金：国家863计划(编号:2002AA205021);国家自然基金青年项目(编号:30801159)

摘　　要：目的:观察团块样自体软骨细胞植入关节软骨缺损后的病理变化,探讨自体软骨细胞移植修复关节软骨缺损的病理生理机制。方法:24只3.0kg以上4～6月龄新西兰大白兔,雌雄不限,随机分为两组:实验组和对照组。实验组12只,20%的速眠新(1mg/kg)肌肉注射麻醉后取肩关节软骨组织,0.2%Ⅱ型胶原酶消化分离软骨细胞,体外单层培养,细胞长成肉眼可见的膜状后收集固体的组织样细胞团,动物再次麻醉制造双膝股骨滑车4.0mm×6.0mm方形缺损,植入细胞团块,骨膜覆盖,缝合骨膜于双股骨髁上。对照组12只,同实验组手术方法进行缺损单纯骨膜移植。1、3、12、24周两组各3只动物空气栓塞处死取材,观察细胞团块变化和缺损修复情况。结果:1周时软骨细胞朝向关节面部分细胞变大变圆,产生大量基质;3周时此种变化更加明显,但骨膜与细胞团块已然不能分开;12周时缺损为类透明软骨组织修复;24周时修复组织为透明软骨样组织,对照组为纤维软骨组织修复。结论:关节软骨细胞体外聚集培养形成的细胞团块内的细胞有迁移生长能力;细胞团块移植方法植入的细胞数量大,表型好,细胞在缺损内不会流失;关节软骨缺损修复是由植入的细胞团块生长分化而来;自体关节软骨细胞团块植入关节缺损内后,在关节应力的影响下,先从朝向关节面的一侧逐渐发生细胞成熟分化和软骨基质产生,逐渐完成缺损的修复。Objective：To trace the pathological changes of the cultured autologous chondrocytes mass after implanted in cartilage defects and investigate the pathophysiological mechanisms of the antologous chondrocytes mass transplantation in the repair of cartilage defects.Methods：Twenty four New Zealand white rabbits of 4 to 6 month old and weighing more than 3.0 kg （female and male was unrestricted） were randomly divided into experiment group and the control group.For 12 rabbits of experiment group,the cartilage defects were repaired with the autologous chondrocytes mass and sealed with one piece of periosteum.Firstly,cartilage tissue of 10 to 30 mg was obtained from the shoulder of the rabbits after anaesthetized by 1 mg/kg 20% sumianxin.Then,chondrocytes were isolated from the cartilage tissue with 0.2% type Ⅱ collagenase digestion and were cultured in DMEM/F-12 supplemented with 20% fetal bovine serum （FBS）,50 μg/ml ascorbic acid 2 phosphate,0.4 mM proline,5 μg/ml insulin and 1 mM non essential amino acids （NEAA） in flasks in vitro.The cells were harvested until a thin film of the cells covered the bottom of the flask could be seen with naked eyes.Then the film was collected with a curled glass stick and formed a solid mass.On this time,the animal was anaesthetized again and the full thickness cartilage square defect of 4.0 mm×6.0 mm was fab-ricated in the patellar grove of distal femur,and then the cellular mass was transplanted into the defect covered by one piece of periosteum which obtained from the upper anterior of tibia and sealed with the femoral condyles.For 12 rabbits of the control group, the defects were sealed with one piece of periosteum only.The animals were sacrificed in the 1st,3rd,6th and 12th weeks after the operation respectively.The histologic sections were stained with safranin O fast green,hematoxylin eosin （HE） and picric acid Sirius red and immunostained for type Ⅱ collagen and aggrecan.Results：In the 1st week,the transplanted cells oriented to articular surface dif

关 键 词：软骨 关节 移植 自体 软骨细胞 软组织损伤 

分 类 号：R684.705[医药卫生—骨科学]