Development of efficient process for isolation, propagation and differentiation induction of normal human hepatic stem cells and its application to artificial liver

1、正常人肝干细胞在增殖过程中转化及其预防的研究如果正常人肝干细胞（肝母细胞）在增殖过程中转化，这些细胞就不能应用于再生医学和人工肝。事实上，干细胞和转化的肿瘤细胞具有相似的特性和制造者。本研究以肝细胞的临床肿瘤生成因子PIVKA-II（异常凝血酶原）为制造因子，建立了肝干细胞（肝母细胞）转化的检测方法。我们通过PIVKA-II实验发现，人胎肝细胞包括肝母细胞在第8代繁殖时开始转化。结果表明，丁酸钠对人胎肝细胞成肝细胞的诱导作用与丁酸钠浓度有关。还发现维生素K对人胎儿肝细胞的转化有预防作用。2、肝母细胞高密度培养基质材料的研究。为了开发人成肝细胞人工肝模块，有必要对培养底物的材料特性进行高密度培养、功能表达和人成肝细胞转化的预防研究。在本研究中，我们检测了几种培养基质。大孔siran材料更适合高密度培养。多孔羟基磷灰石更有利于细胞色素P450活性的表达。

1, Studies on transformation of normal human hepatic stem cells in propagation process and its preventionIf the normal human hepatic stem cells (hepatoblast) transform in propagation process, these cells can not be applicable to regenerative medicine and artificial liver. In fact, the stem cells and transformed tumor cells have similar characteristics and makers. In this research, we developed the examination method for the transformation of hepatic stem cells (hepatoblast) using PIVKA-II (abnormal prothrombin) as a maker, which is clinical tumor maker of hepatocytes. We indicated that human fetal hepatocytes including hepatoblasts started to transform at the8th passage of propagation by using PIVKA-II assay. And we also indicated that induction of hepatoblast from human fetal hepatocytes teated with Sodium butyrate, caused the transformation depended on the concentration of sodium butyrate. It was also found that Vitamin K has prevention effect of the transformation of human fetal heptaocytes.2,Studies on the materials of culture substrate for high density culture of hepatoblast.For development of artificial liver module using human hepatoblast, it is necessary to study on material characteristics of culture substrate for high density culture, functional expression and prevention of transformation of human hepatoblast. In this research, we examined several culture substrates. Macroporous siran material was better for high density culture. And porous hydroxyapatite was better for cytochrome P450 activity expression of hepatoblast.