Research on Fabrication of Tissue Engineering Scaffold for Reconstruction of Internal Organs with High Metabolic Rate by Laser Sintering Solid Freeform Fabrication and Cell Culture

肝脏等内脏器官的重建需要三维支架，这些支架具有精细的流动通道网络。为了开发这样的支架，我们应用激光烧结自由成型制造方法，使用可生物降解塑料粉末。这种方法的优点是提供了大量的材料选择，但另一方面，商业上可用的机器的精度不足以满足我们的目的。为了避免这一缺点，我们开发了一种新的设备，以减少激光光斑尺寸的粉末床上的一个因素的70至50%。此外，我们研究了制造精度和水可浸出填料的各种性质之间的关系，水可浸出填料用于提供具有约90%的非常高的孔隙率的支架。通过参数优化和新仪器的使用，使测量精度提高了一倍。新开发的支架配备有厚度最小为500 μ m的流动通道网络。这种改善 ...更多信息 预期将获得的细胞密度增加约4倍。通常，由于光学原因，减小激光束的焦斑尺寸牺牲了激光扫描器的扫描范围。为了找回丢失的构建信封，我们开发了一种新的系统，可以在平行于粉末床的平面上移动激光扫描仪。结果表明，尽管采用振镜式激光扫描器扫描范围仅为50×50mm^2，但仍获得了150×150mm^2的宽扫描范围，可实现大尺寸支架的制作。利用该设备，我们成功地开发了一种具有流动通道的多孔体。为了避免支架放大后氧气输送不足的问题，研究了使用纳米封装血红蛋白（人造红细胞）的可行性。在使用肝祖细胞的培养试验中，发现氧供应有一定的改善。然而，观察到从胶囊中释放的游离血红蛋白分子在细胞中被摄入并破裂所引起的细胞毒性，尽管在使用成熟肝细胞的初步试验中，血红蛋白的胶囊化成功地防止了这一问题。使用开发的具有流动通道网络的支架对人肝癌细胞系进行了灌注培养测试，并且观察到细胞生长和肝功能表达的显著改善。在该试验中，细胞增殖被限制在距每个流道200μm的范围内。这表明，最佳支架应设计为支架中的任何位置距离流道在200μm的范围内。少

Reconstruction of internal organs such as liver requires three dimensional scaffolds that are equipped with network of fine flow channels. To develop such scaffolds, we applied laser sintering freeform fabrication method using biodegradable plastic powder. This method provides a large choice of material as its advantage, but on the other hand, precision of commercially available machines is not sufficient for our purpose. To avoid this drawback, we developed a new apparatus to reduce laser spot size on the powder bed by a factor of 70 to 50%. Additionally, we investigated relationship between precision of fabrication and various properties of water leachable filler, which is applied to provide the scaffolds with very high porosity around 90%. As a result of the parameter optimization and use of the new apparatus, we succeeded in doubling the precision. The newly developed scaffold was equipped with a network of flow channels of which thickness is 500um at the minimum. This improvement … More is expected to increase cell density to be obtained by a factor of around 4. Generally, reducing focal spot size of a laser beam sacrifices the scanning range of laser scanner by optical reason. To retrieve the lost build envelope, we develop a new system that can move the laser scanner in a plane parallel to the powder bed. Resultantly, wide envelope of 150×150mm^2 is obtained although employed galvanomirror type laser scanner's range is as narrow as 50×50mm^2 to realize fabrication of large scaled scaffolds. Using this equipment, we succeeded in developing a porous body with flow channels.To avoid the problem of insufficient oxygen delivery following scaling up of scaffold, feasibility of using nano-encapsulated hemoglobin (artificial red blood cells) was investigated. In culture test using hepatic progenitor cells, a certain improvement of oxygen supply was found. However, cytotoxicity caused by free hemoglobin molecules released from the capsules that were ingested and broken in the cells was observed, although encapsulation of hemoglobin successfully prevented this problem in preliminary test using mature hepatic cells. Perfusion culture of human liver cancer cell lines using a developed scaffold with flow channel network was tested, and significant improvement both in cell growth and expression of liver function was observed. In the test, cell propagation was limited in the range of 200μm from each flow channel. This demonstrates that optimal scaffold should be so designed that any position in the scaffold is in the range of 200μm from a flow channel. Less