心脏瓣膜组织工程支架的制备及细胞种植  

Preparation of Heart Valve Scaffold and Cell Seeding

作　　者：龙莉[1] 吴春根[1] 潘銮凤[1] 齐晓岚[2] 洪涛[3] 

机构地区：[1]复旦大学上海医学院分子生物学实验室 [2]贵阳医学院分子生物教研室,贵阳550004 [3]复旦大学中山医院心外科,上海200032

出　　处：《生物医学工程学杂志》2004年第4期610-613,共4页Journal of Biomedical Engineering

摘　　要：为了探讨心脏瓣膜组织工程支架的制备及细胞种植 ,我们将新鲜猪心瓣膜用胰蛋白酶及 DNA酶消化去除细胞 ,光镜和电镜观察其结构 ,并将人脐静脉内皮细胞株、猪颈动脉内皮细胞和狗肌成纤维细胞滴种在脱细胞猪心瓣膜上 ,HE和免疫组化染色观察。结果显示胰蛋白酶联合 DNA酶消化去除了所有细胞 ,而瓣膜的三维结构保持完好。种植的人内皮细胞几乎完全覆盖了瓣膜的表面 ,猪内皮细胞在瓣膜上呈斑块状生长 ,狗肌成纤维细胞不但生长于瓣膜表面 ,且渗透到基质内部生长。这表明 ,胰蛋白酶联合 DNA酶消化是一种较为理想的猪瓣膜脱细胞方法 ;人和猪血管内皮细胞及狗肌成纤维细胞均能在猪瓣膜支架上较好地黏附和生长。To prepare scaffolds for heart valve tissue engineering, porcine heart valves were treated with varied concentrations of trypsin for 32,56,80 and 104 h or followed with DNase. And then the structure of acellular valves was observed under light microscope,scanning and transmission electron microscope. Porcine endothelial cells,human endothelial cells,and canine myofibroblasts were reseeded onto the acellularized porcine heart valve scaffolds once a day for 3 days. The valves were analyzed by immunohistochemical staining and electron microscopy. Results show that all endothelial cells and the majority of interstitial cells were removed from the heart valves after digestion with trypsin for 104 h,and the collagen fiber structure remains intact,but the space between collagen fibers increased slightly. Incubation with trypsin for 80 h and then with DNase almost removed all cells,and the collagen fiber structure and the space between the fibers remain intact. After reseeding,human endothelial cells almost fully cover the valve scaffold surface as shown by H-E staining and platelet endothelial cell adhesion molecules (PECAM-1) staining. Xenogeneic porcine endothelial cells also adhered to and grew on the scaffolds. As shown by H-E staining and actin staining,canine myofibroblasts not only adhered to the surface of valve scaffold but also migrated to the inner part of matrix after one week culture. These results suggest that the digestion of porcine heart valves with trypsin combining with DNase is a suitable method to remove cells. The acellular porcine heart valve scaffolds have a quite favorable biocompatibility with human and porcine endothelial cells as well as canine myofibroblasts.

关 键 词：心脏瓣膜 组织工程 支架 制备 细胞种植 血管内皮细胞 肌成纤维细胞 

分 类 号：R318.1[医药卫生—生物医学工程]