Effectiveness of fructose-modified chitosan as a scaffold for hepatocyte attachment.
Effectiveness of fructose-modified chitosan as a scaffold for hepatocyte attachment.
复制标题
果糖修饰的壳聚糖作为肝细胞附着支架的有效性。
DOI：
10.1248/bpb.20.1290
复制
发表时间：
1997
影响因子：
2
通讯作者：
M. Kawase
中科院分区：
文献类型：
--
作者：
K. Yagi;N. Michibayashi;N. Kurikawa;Y. Nakashima;T. Mizoguchi;A. Harada;S. Higashiyama;H. Muranaka;M. Kawase
壳聚糖是一种先前被证明是肝细胞附着的良好支架的物质，其游离氨基被果糖共价修饰。该修饰显著增加了壳聚糖凝胶表面可附着的细胞数量。在果糖-壳聚糖上培养的大鼠肝细胞与在未修饰的壳聚糖上培养的大鼠肝细胞表现相似，即它们保持了体内的球形，并且释放的乳酸脱氢酶比附着在胶原涂层表面的细胞少得多。果糖修饰并未改变壳聚糖用于肝细胞培养的重要特性：在果糖-壳聚糖培养的肝细胞中，尿素合成和药物代谢等肝脏特异性功能可稳定维持5 d。与此形成鲜明对比的是，附着在胶原包被表面的肝细胞发生了严重的形态变化，从球形变为扁平，并在5d内几乎失去了所有的利多卡因去除活性。通过将壳聚糖和胶原中的游离氨基与戊二醛交联，将极薄的果糖-壳聚糖层固定在聚苯乙烯板表面和葡聚糖微载体上。在果糖壳聚糖包被表面的肝细胞保持其球形，掩盖了胶原层的细胞扁平效应。然后用空心纤维筒进行灌注培养，其中附着在果糖壳聚糖包被微载体上的肝细胞悬浮在毛细血管外间隙中，培养4d期间肝脏特异性功能稳定维持。因此，果糖-壳聚糖包被表面似乎是一种非常有前途的肝细胞附着支架，可用于肝脏功能的细胞生物学研究，特别是与细胞色素P450相关的研究，以及生物人工肝支持系统。
Free amino groups of chitosan, a substance which has previously been shown to be a good scaffold for hepatocyte attachment, were covalently modified with fructose. The modification significantly increased the number of cells that could be attached on the surface of chitosan gel. Rat hepatocytes cultivated on fructose-chitosan behaved similarly to those on unmodified chitosan, i.e., they retained the spherical shape they have in vivo, and released much less lactate dehydrogenase than cells attached on a collagen-coated surface. The modification with fructose did not alter the important characteristics of chitosan for hepatocyte culture: liver-specific functions such as urea synthesis and drug metabolism were stably maintained for 5 d in the hepatocytes cultured on fructose-chitosan. In sharp contrast, hepatocytes attached on a collagen-coated surface underwent a severe morphological change, from spherical to flat, and lost almost all their lidocaine-removal activity within 5d. A very thin fructose-chitosan layer was also applied onto the collagen-coated surfaces of polystyrene plates and a dextran microcarrier by crosslinking free amino groups in the chitosan and collagen with glutaraldehyde to fix the thin layer. Hepatocytes on the fructose-chitosan-coated surface retained their spherical shape, masking the cell-flattening effect of the collagen layer. Perfusion culture was then carried out using a hollow-fiber cartridge in which hepatocytes attached on fructose-chitosan-coated microcarriers were suspended in the extracapillary space: the liver-specific functions were stably maintained during 4d of the culture. A fructose-chitosan-coated surface thus appears to be a very promising scaffold for hepatocyte attachment which can be used in cellular biological studies of liver functions, especially in relation to cytochrome P450, as well as in bioartificial liver support systems.