tgf-β1及ctgf在骨不连接区表达的实验研究-experimental study on expression of tgf - β 1 and ctgf in nonunion region.docx

目录中文摘要····················································································2英文摘要····················································································4前言························································································7材料和仪器··············································································10实验方法···················································································12实验结果···················································································22讨论·······················································································32结论·······················································································35参考文献···················································································36致谢·······················································································39附录A中英文术语及缩略词对照表·····················································40附录B常用试剂配制·······································································42附录C个人简历及发表论文·······························································44附录D综述···················································································45TGF-β1及CTGF在骨不连接区表达的实验研究中文摘要目的：骨折局部常常因为纤维组织及软骨缺乏足够的骨化而形成骨不连，而转化生长因子β1（TGF-β1）及结缔组织生长因子（CTGF）的协同具有明显促进软骨及骨增殖和分化的特性，从而促进骨愈合。本实验通过观察骨缺损不愈合和愈合过程中TGF-β1及CTGF的局部分布和表达水平差异，研究骨缺损不愈合的分子机理，进一步探讨TGF-β1及CTGF对骨折愈合的影响。方法：1、制作骨折愈合及骨缺损不愈合动物模型。选成年新西兰大白兔30只，雌雄不限，随机分为6组，每组5只，制造一侧桡骨干骨缺损不愈合模型为实验组，制造另一侧骨折愈合模型为对照侧。术后1、2、4、6、8、12周取材。2、X线及组织学观察骨愈合及组织学成骨情况。3、免疫组化检测TGF-β1及CTGF的表达。4、逆转录聚合酶链反应（RT-PCR）检测以上两因子的基因表达。结果：1、X线及组织学检查显示实验组骨缺损区未形成骨性连接，主要为纤维结缔组织所充填，骨缺损两端有少量的膜内成骨及软骨内成骨存在，大部分肉芽组织及肥大软骨细胞转化为纤维结缔组织，断端硬化，髓腔闭塞。对照组膜内成骨及软骨内成骨均活跃，6周时骨折愈合。2、免疫组化显示实验组和对照组TGF-β1及CTGF表达在2周时均明显增强，但实验组表达量明显低于对照组（P0.05），4-6周实验组TGF-β1及CTGF在各种细胞中表达均明显减少至微弱表达，而对照组在成骨细胞和周围的骨基质中仍有较强表达，差异有显著性（P0.05）,8周后两组TGF-β1及CTGF表达均微弱，无显著性差异(P0.05).3、RT-PCR检测显示：实验组TGF-β1mRNA在1、2、4、6周的相对表达量分别为（0.347±0.013、0.785±0.025、0.378±0.029、0.257±0.030），均明显低于对照组