DEVELOPMENT OF COATING MATERIAL FOR ARTIFICIAL TRACHER

使用生长培养法培养NHBEC。将正常人支气管切成3 mm的正方形，将这些支气管段放置在聚苯乙烯培养皿上，并将Marlex网用作人工气管，其涂有一种或多种细胞外基质，如下所述。然后使用无血清培养物LHC-9进行外植体培养。用相差显微镜观察NHBEC在支气管段周围的生长情况，测量实验结束时的表面积，作为生长指标。（I、II、III或IV型）将促进NHBEC的生长。基于实验1中获得的结果，使用IV型胶原和两种细胞粘附蛋白层粘连蛋白和纤连蛋白涂覆培养皿以进行比较。实验三：实验A组中，Marlex网片仅涂有I型胶原，B组中，Marlex网片涂有I型胶原、IV型胶原、层粘连蛋白和纤连蛋白，以检测NHBEC的生长。实验I中获得的结果表明，与其他类型的胶原相比，NHBEC在涂有IV型胶原的聚苯乙烯培养皿中的生长最稳定，在实验2中显示IV型胶原、层粘连蛋白和纤连蛋白的组合促进NHBEC的生长，在实验3中显示涂覆有IV型胶原、层粘连蛋白和纤连蛋白的网状物促进NHBEC的生长。从这些发现，提示IV型胶原、层粘连蛋白和纤连蛋白的组合，层粘连蛋白和纤连蛋白可作为支气管上皮细胞覆盖人工气管内表面的有效基质。

The outgrowth culture method was used to grow NHBEC.A normal human bronchus was dissected into 3mm squares and these bronchial segments were placed with the mucosal surface up on a polystirene dish and a Marlex mesh used as an artificial tracheal that was coated with one or more extracellular matrices that are described below.Then explant culture was performed using a serum free culture, LHC-9. A phase contrase microscope was used to examine the outgrowth of NHBEC in the periphery of the bronchial segments, and the surface area on completion of the experiment was masured and used as a growth indicator.Experiment 1 : It was examined as to which type of collagen (type I,II,III or IV) would promote the growth of NHBEC.Experiment 2 : Based on the result obtained in experiment 1, type IV collagen and two cellular adhesive proteins, laminin and fibronectin, were used to coat a culture dish for comparison. Experiment 3 : In group A,a Marlex mesh was coated with type I collagen only, and in group B,a Marlex mesh was coated with type I collagen, type IV collagen, laminin and fibronectin to examine the growth of NHBEC.The results obtained in experiment I showed that the growth of NHBEC in the polystirene dish coated with type IV collagen was most stable when compared to the other types of collagen, in experiment 2 showed that the combination of type IV collagen, laminin and fibronectin promoted the growth of NHBEC,in experiment 3 showed that the mesh coated with type IV collagen, laminin and fibronectin promoted the growth of NHBEC.From these findings, it was suggested that the combination of type IV collagen, laminin and fibronectin could beused as an effective base to cover the interior surfase of artificial trachea with bronchial epithelial cells.