骨髓间充质干细胞复合富血小板纤维蛋白异位成软骨的实验研究  

Ectopic chondrogenesis of bone marrow mesenchymal stem cells combined with platelet-rich fibrin in vivo

作　　者：李轶杰[1] 陈慧[1] 刘鹏[1] 张旻 陈永进[1] 

机构地区：[1]第四军医大学口腔医学院,陕西西安710032

出　　处：《牙体牙髓牙周病学杂志》2013年第3期162-167,共6页Chinese Journal of Conservative Dentistry

基　　金：国家自然科学基金资助(31170888)

摘　　要：目的:探讨富血小板纤维蛋白(platelet-rich fibrin,PRF)复合兔骨髓间充质干细胞膜片在软骨微环境下异位成软骨的可行性。方法:取3～4月龄雄性新西兰兔骨髓,体外分离、培养骨髓间充质干细胞;经来源鉴定后,取第3代细胞诱导培养形成细胞膜片;抽取同一只兔动脉血制备PRF后与自体骨髓间充质干细胞膜片复合。取4周龄BALB/C裸鼠30只,随机分组;实验组(n=15)于背部两侧植入骨髓间充质干细胞膜片复合富血小板纤维蛋白与兔耳软骨碎块的双膜结构复合物;对照组(n=15)背部两侧仅植入骨髓间充质干细胞膜片与兔耳软骨碎块的单膜结构复合物。分别于植入后2、4、8周取材,HE、甲苯胺蓝染色,用IPP 6.0软件对图片进行着色强度分析。所得结果用SPSS 13.0软件进行统计处理。结果:HE染色显示,实验组2、4、8周均可见软骨细胞生成,对照组仅8周时有少量软骨细胞生成。IPP分析显示,实验组2、4、8周甲苯胺蓝着色强度呈逐渐增加趋势,各时间点间OD值相比差异均有统计学意义(P<0.05);而且实验组各时间点OD值均显著高于同时间点的对照组(P<0.05)。结论:骨髓间充质干细胞与PRF复合的双膜结构在软骨微环境中具成软骨能力。AIM： To explore the ectopic chondrogenesis effects of double membrane transplant constructed by homologous rabbit platelet-rich fibrin （PRF） and bone marrow mesenchymal stem cell （BMSC） sheet in cartilage microenvironment in vivo. METHODS： BMSCs was obtained from New Zealand rabbits by density gradient centrifuga-tion. The cells were identified by flow cytomety, osteogenic and adipogenic induction assays and then used to form BM-SC sheet cultured by standard medium with Vitamin C. PRF membrane was prepared by centrifuging the whole blood from the rabbit auricle arterial and then squeezing out the serum from the clot. The PRF membrane was co-cultured with BMSC sheet. Afterwards, the BMSCs/PRF compound membranes enwraping auricle cartilage fragments were im-planted into the dorsal subcutaneous area of 15 nude mice. Only BMSC sheets enwraping auricle cartilage fragments were implanted into another 15 nude mice as the controls. 4, 8 and 12 weeks after implantation, cartilage formation was observed by HE staining and toluidine blue staining. IPP 6.0 was used for image analysis and SPSS 13.0 was used for data analysis. RESULTS ： HE staining showed ehondrogenesis in ectopic sites 2, 4 and 8 weeks after implantation in experimental group. In contrast, only a few chondrocytes were observed in control group at 8 weeks. IPP data analy-sis showed the average optical density value of experimental group increased gradually and was significantly higher than that of the control at the same time points （ P 〈 0.05 ）. CONCLUSION： The new autotransplant constructed by the BMSC sheet and PRF can provide an effective way for chondrogenesis.

关 键 词：骨髓间充质干细胞 富血小板纤维蛋白 软骨分化 组织工程 裸鼠 

分 类 号：R780.2[医药卫生—口腔医学]