同种生物心脏组织工程瓣体外构建的初步研究  

Preliminary study of fabricating homologous tissue engineering heart valve in vitro

作　　者：刘迎龙[1] 冯滨[1] 谢宁[1] 冯凯[2] 宋来凤[3] 

机构地区：[1]中国医学科学院中国协和医科大学心血管疾病研究所阜外心血管病医院外科,100037 [2]中国军事医学科学院附属医院全军造血干细胞移植中心 [3]中国医学科学院中国协和医科大学心血管疾病研究所阜外心血管病医院病理科,100037

出　　处：《中华胸心血管外科杂志》2004年第5期282-285,共4页Chinese Journal of Thoracic and Cardiovascular Surgery

基　　金：国家高技术研究发展计划 (863 ) (2 0 0 1AA2 160 61) ;国家自然科学基金 (3 0 2 712 89) ;中国博士后科学基金 (2 0 0 3 0 3 3 2 2 4)资助

摘　　要：目的　在生物瓣支架上种植并静态培养自体内皮细胞 (ECs) ,形成完整的内皮细胞单层 ,为下一步脉动流培养并最终体外构建同种生物组织工程瓣 (TEHV)提供材料基础。方法　生物瓣支架选择经液氮保存的成人主动脉带瓣管道 ,用 0 1 %SDS脱去表面的ECs ;成人骨髓间充质干细胞 (MSCs)体外定向诱导分化的ECs作为种子细胞 ,高密度 (>1 0 5cell/cm2 )种植于瓣膜支架上静态培养 2 0d ,扫描电镜观察、摄片 ,以确定再内皮化程度。结果　同种生物瓣支架表面的ECs完全脱去 ,而细胞外基质成分保存良好 ;MSCs体外诱导分化的ECs与生物瓣支架复合体静态培养第 7、1 4和 2 0d ,再内皮化程度分别为 73%、85 %和 92 %。结论　静态培养条件下构建的TEHV基本上实现了体外再内皮化的预期目标。Objective: The endothelial cells (ECs) was implanted on decellularized scaffold of homologous bioprosthetic valve (HBV) in vitro, and the complex structure was statically cultured in order to form a confluent ECs monolayer, which provides basic material for pulsatile flow cultivation and fabrication of tissue engineering heart valve (TEHV). Methods: The superficial ECs of homologous valved aortic tube, conserved by liquid nitrogen, were removed by low osmotic Tris buffer, containing 0.1% sodium dodecylsulphate (SDS). Human bone marrow mesenchymal stem cells (MSCs) were separated by gradient centrifugation on Percoll (density 1.073 g/ml) from human bone marrow (HBM) and incubated for orientated differentiation into ECs in DMEM (high glucose) with 20% FBS and VEGF (10ng/ml) for about 14～21 days. As the seed cells, the ECs were implanted on the decellularized scaffold of HBV with high density seeding (>105 cells/cm 2), and statically cultured in vitro for 20 days. The morphologic structure was observed and photographed by scanning electron microscope (SEM). Results: The ECs of the HBV were not only removed completely, but also reserved collagen and elastic fibers integrally, which are two of main components of extracellular matrix. The ECs from MSCs differentiated were successful implanted on the HBV scaffold, which the recellularization level on the seventh, fourteenth and twentieth day was 73%, 85% and 92% respectively. Conclusion: Application of differentiating ECs from MSCs as seed cells, fabricating TEHV by us was primary realized prospective recellularization goal under static culture in vitro.

关 键 词：体外 生物瓣 支架 内皮化 心脏组织工程 EC 内皮细胞 MSC 高密度 单层 

分 类 号：R318.1[医药卫生—生物医学工程]