猪小肠与犬食管脱细胞基质(ECM)构建组织工程化人工食管的比较研究

摘要

Objective Background Bonc marrow mcscnchymal stem cell is a kind of non-diffcrcntiation cell which can scif-rcncwai and have potent of multiiincagc differentiation, wide variety of sources,convenient separation,proliferate rapidly,and is currently accepted as the best seed cells for tissue engineering. Extracellular matrix (ECM) has good biocompatibility,plenty of porosity,conducives to the seed cell adhesion,proliferation,and induces cell along the "mold" growth,differentiation,is the ideal scaffold material for tissue engineering. Objective As the carrier,extracellular matrix composite with bone marrow mcscnchymal stem cells in vitro, contrast with autologous csophagcal dcccllularizcd scaffolds and dacron graft respectively. Methods Use physical and chemical method for preparation of ECM,observed the surface structure by scanning electron microscopy. Isolation and culture of canine bone marrow mcscnchymal stem cell in vitro, to identified by flow cytomctry. The third generation of canine BMSCs were taken and seeded on porcine intestinal extracellular matrix (IECM) , canine esophagus extracellular matrix (EECM) and cPTFE, observed by HE staining and scanning electron microscopy. Results Extracellular matrix is a translucent white membrane, has a good stereo three-dimensional space structure,visible fiber mesh structure clearly and interconneetivity between each other. Flow cytomctry showed that: the expression rate of CD29,CD44 ,CD90 and CD105 almost 100% , while CD34.CD45 almost not expressed. BMSCs arc cultured on the carrier bracket for 15d,thcn fixed in formalin,stained with HE and observed by scanning electron microscopy showed that the cell seeded scaffolds of IECM and EECM scaffold's growth rate all well. Conclusion The cell seeded IECM scaffold's biological traits is more close to the EECM scaffold,at the same time,the source of the IECM is more widely,so it is a better choice of the scaffold.%背景骨髓间充质干细胞(BMSCs)具有多向分化潜能,来源广泛、分离方便、增殖迅速,是目前公认的组织工程最佳种子细胞;脱细胞基质膜(ECM)有良好的组织相容性,充足的孔隙率,利于种子细胞的附着、增殖,并诱导细胞沿"模具"的性状生长、分化,是组织工程的理想支架材料.目的 为比较研究骨髓间充质干细胞在体外在不同的支架材料上复合生长情况,分别与猪小肠脱细胞基质(IECM);犬自体食管脱细胞基质(EECM)和人工补片(ePTFE)做对比研究.方法 使用物理方法和化学方法制备ECM,扫描电镜观察其表面结构.体外提取分离培养犬骨髓间充质干细胞,做流式细胞学鉴定后,取第三代BMSCs分别与IECM、EECM、ePTFE复合培养,HE染色和扫描电镜观察.结果 ECM为白色菲薄,半透明的膜状物,扫描电镜可见其清晰的纤维网状结构,具有良好的立体三维空间结构,并且之间相互连通.BMSCs经流式细胞学鉴定CD29、CD44、CD90、CD105表达率几乎100%,而CD34、CD45几乎不表达.BMSCs与载体支架复合培养15d后,做HE染色切片观察,细胞与IECM、EECM复合生长率良好,与ePTFE较差.结论 复合培养后的IECM组与EECM组生物性状较为接近,同时IECM的来源更加广泛,是组织工程化人工食管支架材料的更好选择.