中国组织工程研究

中国组织工程研究 ›› 2018, Vol. 22 ›› Issue (9): 1338-1343.doi: 10.3969/j.issn.2095-4344.0462

• 骨髓干细胞 bone marrow stem cells • 上一篇    下一篇

慢病毒介导人骨形态发生蛋白2/骨髓间充质干细胞/脱钙骨基质修复股骨大段缺损

陶 旋，李 强，李诗鹏，石正松，周桢杰   

  1. 桂林医学院附属医院急诊创伤外科，广西壮族自治区桂林市 541001
  • 修回日期:2017-12-20 出版日期:2018-03-28 发布日期:2018-04-03
  • 通讯作者: 李强，硕士，主任医师，教授，硕士生导师，桂林医学院附属医院急诊创伤外科，广西壮族自治区桂林市 541001
  • 作者简介:陶旋，男，1991年生，四川省乐山市人，汉族，桂林医学院在读硕士，主要从事骨组织工程研究。
  • 基金资助:

    国家自然科学基金资助项目(31160199)；广西壮族自治区自然科学基金资助项目(2014GXNSFAA118263)；桂林医学院科研项目(LX2014254)

Repair of large-segmental femoral defects using lentivirus-mediated bone morphogenetic protein 2/bone marrow mesenchymal stem cells/demineralized bone matrix

Tao Xuan, Li Qiang, Li Shi-peng, Shi Zheng-song, Zhou Zhen-jie   

  1. Department of Emergency Traumatic Surgery, the Affiliated Hospital of Guilin Medical University, Guilin 541001, Guangxi Zhuang Autonomous Region, China
  • Revised:2017-12-20 Online:2018-03-28 Published:2018-04-03
  • Contact: Li Qiang, Master, Chief physician, Professor, Master supervisor, Department of Emergency Traumatic Surgery, the Affiliated Hospital of Guilin Medical University, Guilin 541001, Guangxi Zhuang Autonomous Region, China
  • About author:Tao Xuan, Master candidate, Department of Emergency Traumatic Surgery, the Affiliated Hospital of Guilin Medical University, Guilin 541001, Guangxi Zhuang Autonomous Region, China
  • Supported by:

    the National Natural Science Foundation of China, No. 31160199; the Natural Science Foundation of Guangxi Zhuang Autonomous Region, No. 2014GXNSFAA118263; the Natural Science Foundation of Guilin Medical University, No. LX201425

PDF

301

摘要：

文章快速阅读：

文题释义：
慢病毒载体：是以人类免疫缺陷Ⅰ型病毒为基础发展起来的基因治疗载体。区别一般的逆转录病毒载体，它对分裂细胞和非分裂细胞均具有感染能力。其可以将外源基因有效地整合到宿主染色体上，从而达到持久性表达。在感染能力方面可有效地感染神经元细胞、肝细胞、心肌细胞、肿瘤细胞、内皮细胞、干细胞等多种类型的细胞，从而达到良好的的基因治疗效果。
脱钙骨基质：是一种是由胶原蛋白、非胶原蛋白以及较低浓度的生长因子等组成的复合物天然骨移植材料，主要来源于人或动物的颅骨、股骨干和胫骨干骺端。其具有良好的生物学特性、骨诱导性和骨传导性，可生物降解，促进新骨形成及骨组织矿化，加速骨愈合，可以单独或与自体骨、其它生物材料、生长因子联合有效修复骨损伤，是比较理想的骨组织工程支架材料。

 

摘要
背景：课题组前期研究已证实，慢病毒载体介导EGFP/骨形态发生蛋白2 基因转染兔骨髓间充质干细胞在表达持续时间上具有一定的优势。但慢病毒介导人骨形态发生蛋白基因转染兔骨髓间充质干细胞复合脱钙骨基质能否在体内促进骨缺损修复的问题有待探究。
目的：观察兔股骨大段缺损中植入慢病毒介导人骨形态发生蛋白2转染骨髓间充质干细胞复合脱钙骨基质后的修复效果，探寻治疗股骨大段缺损的新途径。
方法：选取48只新西兰大白兔，股骨中段截骨钢板内固定法制备兔左侧股骨大段缺损模型。造模成功后，随机分成A，B，C，D组，每组12只。A 组无修复材料植入，B，C，D组分别植入脱钙骨基质、脱钙骨基质/骨髓间充质干细胞、慢病毒载体介导人骨形态发生蛋白2/骨髓间充质干细胞/脱钙骨基质。造模后2，4，8 及12 周分别处死每组3只兔子，通过苏木精-伊红染色、生物力学分析，X射线检查评价股骨大段缺损修复的情况。
结果与结论： ①X 射线示A，B，C，D组均有不同的骨痂修复形成，且Lane-Sandhu X射线评分A<B<C<D组(P < 0.05)；②三点弯曲实验结果表明造模后8，12周时D组最大负荷显著高于C组，但显著低于正常股骨(P < 0.05)；③苏木精-伊红染色显示A组可见少量排列紊乱的骨小梁和大量的纤维组织；B组脱钙骨基质支架已降解，可见部分排列紊乱的骨小梁和大量的纤维组织；C组骨小梁已基本连接成线，开始塑形为皮质骨，髓腔再通不明显；D 组已基本形成新的骨皮质，有髓腔再通；④结果提示，骨缺损区域植入慢病毒载体介导人骨形态发生蛋白2/骨髓间充质干细胞/脱钙骨基质后有大量的骨痂形成，良好的生物力学性能，完整的骨皮质，说明支架促进了骨传导、骨诱导和骨生成，对兔股骨大段缺损有较好的修复效果。

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程
ORCID: 0000-0001-9327-3574(李强)

关键词: 慢病毒, 骨形态发生蛋白2, 骨髓间充质干细胞, 脱钙骨基质, 股骨大段缺损, 钢板内固定, 组织修复, 国家自然科学基金

Abstract:

BACKGROUND: Our research team has confirmed that compared to the adenoviral vector, transfection by lentiviral vector to rabbit bone marrow mesenchymal stem cells (BMSCs) is more effective that the expression of enhanced green fluorescent protein (EGFP)/bone morphogenetic protein 2 (BMP-2) can be persistent for a longer term. But further investigations are needed to explore whether BMSCs transfected with hBMP-2 through lentivirus combined with demineralized bone matrix (DBM) can promote bone defect repair.
OBJECTIVE: To evaluate the effect of lentivirus-mediated hBMP-2/BMSCs/DBM (LV-hBMP-2/BMSCs/DBM) on the repair of large-segmental femoral defects and to explore the new treatments for large-segmental femoral defects.
METHODS: Large-segmental bone defect models were made in the right femur of 48 New Zealand white rabbits by cutting the middle femoral bone and steel plate fixation. Then, animal models were randomly divided into four groups (n=12 per group) and implanted with nothing (control), DBM, hBMP-2/DBM, and LV-hBMP-2/BMSCs/DBM. Three rabbits from each group were sacrificed at 2, 4, 8 and 12 weeks after surgery to evaluate the repairing effect of femoral defects through hematoxylin-eosin staining, biomechanical analysis and radiological examination.
RESULTS AND CONCLUSION: X-ray results revealed that osteotylus formed in all the four groups to different extents, and Lane - Sandhu X-ray scores were ranked as follows: control group < DBM group < hBMP-2/DBM group < LV-hBMP-2/BMSCs/DBM group (P < 0.05). Findings from the three-point bending test showed that the maximum load of the LV-hBMP-2/BMSCs/DBM group was significantly higher than that of the hBMP-2/DBM group, but is still lower than that of the normal femur at 8 and 12 weeks after modeling (P < 0.05). Hematoxylin-eosin staining results revealed that a few trabecular bones arranged disorderedly and a large amount of fibrous tissues in the control group; the DBM scaffold was basically degraded in the DBM group presenting with partially disordered trabecular bones and a large amount of fibrous tissues; the trabecular bones in the bone defect area were basically connected into line to start the shaping of the cortical bone, and recanalization of the medullary cavity was insignificant in the hBMP-2/DBM group; new cortical bone formed in the bone defect area and the medullary cavity was recanalized in the LV-hBMP-2/BMSCs/DBM group. These findings indicate that LV-hBMP-2/BMSCs/DBM can produce a large amount of calluses, promote formation of new cortical bone, and promote bone conduction, bone induction and osteogenesis after implantation into the bone defect; and this material has good repairing effect on large-segmental femoral defects of rabbits.

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

Key words: Lentivirus, Bone Morphogenetic Proteins, Mesenchymal Stem Cells, Tissue Engineering

中图分类号: