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Experimental research for treatment of liver failure with hepatic tissue stem cell (SP cell)

肝组织干细胞（SP细胞）治疗肝衰竭的实验研究

基本信息

批准号：
14207026
负责人：
HORIE Yoshinori
金额：
$ 27.37万
依托单位：
Keio University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2004
项目状态：
已结题

项目摘要

1. Hepatic differentiation induction method in vitroIn an effort to reconstruct the hepatocyte function and cellular polarity normally found in the liver, adult rat hepatocytes were sandwiched between two layers of hydrated collagen matrix. Functionally, sandwiched hepatocytes maintained the secretion of albumin, the expression of liver specific proteins and the distribution of actin filaments, whereas the cells cultured on single layer of collagen gel decreased the albumin secretion, the liver specific proteins and showed abnormal formation of actin stress fibers and cell spreading. Overlaying a second layer of collagen gels on the hepatocytes that had been cultured on a single gel reversed the cell spreading, reduced actin stress fibers and recovered the liver specific protein expressions.Bone marrow cells could differentiate into hepatocytes when they co-cultured with primary cultured hepatocytes in collagen gel sandwich indicating that hepatocytes cultured in collagen gel sandwich was functionally good enough to induce differentiation of bone marrow cell into hepatocytes.2. SP cellsWe could successfully induce SP cells derived from liver and bone marrow transdifferentiation into hepatocytes in vitro and in vivo. Splenic SP cells have a remarkable capacity for hepatic differentiation, as they were co-cultured with primary cultured hepatocytes in collagen gel. It is our great surprise that the prevalence of SP cells in spleen is 5 to 10 times higher than in bone marrow. Our results indicated that splenic-SP cells might have the potential to transdifferentiate into hepatocytes. Since the prevalence of SP cells in spleen is much higher than in bone marrow, total amount of SP cells could be much larger. Thus the spleen could have the highest prevalence of SP cells and spleen could provide an alternative source of stem cell for stem cell transplantation into liver.

1.体外肝分化诱导方法为了重建肝细胞功能和细胞极性，将成年大鼠肝细胞夹在两层水合胶原基质之间。在功能上，夹心肝细胞维持了白蛋白的分泌、肝特异性蛋白的表达和肌动蛋白丝的分布，而单层胶原凝胶上培养的细胞减少了白蛋白、肝特异性蛋白的分泌，并显示出肌动蛋白应力纤维的异常形成和细胞伸展。将第二层胶原凝胶覆盖在已经在单一凝胶上培养的肝细胞上，逆转了细胞的扩散，骨髓细胞与肝细胞共孵育后，可分化为肝细胞。在胶原凝胶夹层中与原代培养的肝细胞一起培养，表明胶原凝胶夹层中培养的肝细胞功能良好，足以诱导骨分化骨髓细胞转化为肝细胞. SP细胞体外和体内实验均能成功诱导来源于肝脏和骨髓的SP细胞向肝细胞转分化。脾SP细胞与原代培养的肝细胞在胶原凝胶中共培养，具有显著的肝分化能力。令人惊讶的是，脾脏中SP细胞的流行率比骨髓中高5至10倍。我们的研究结果表明，脾SP细胞可能具有转分化为肝细胞的潜力。由于SP细胞在脾脏中的流行率远高于骨髓，因此SP细胞的总量可能更大。因此，脾脏可能具有最高的SP细胞患病率，脾脏可以为干细胞移植到肝脏中提供替代的干细胞来源。