Research for the formation of hepatic organoids by using human hepatic progenitor cells and the model of artificial liver device incorporated with human hepatic organoids

利用人肝祖细胞形成肝类器官及结合人肝类器官的人工肝装置模型研究

基本信息

批准号：
17390353
负责人：
MITAKA Toshihiro
金额：
$ 10.82万
依托单位：
Sapporo Medical University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2007
项目状态：
已结题

项目摘要

We have reported that CD44 is specifically expressed in rat small hepatocytes and that CD44^+ hepatocytes transiently appear in injured rat livers. Using a specific antibody, we sorted CD44^+ cells from galactosamine-treated rat livers and cultured them. The phenotypes and gene expression of the cells were similar to those of cultured SHs and the ^+ cells could differentiate into hepatocytes (J Hepatol, 2006). In addition, some Thy1^+ cells that appeared in the galactosamine-treated livers could differentiate into + SHs and mature into hepatocytes. When the Thyl+ and CD44^+ cells were transplanted into livers treated with retrorsine/PH, they could survive and proliferate to form hepatic foci in the recipient livers. Furthermore, when cultured SHs were transplanted into congenic rat livers treated with radiation/PH, the cells migrated into hepatic trabecules and the liver was partially repopulated by the donor cells (Liver Transplant, 2006). Rat SHs cultured in hyaluronic acid (HA ; ligand for CD44)-coated dishes and serum-free medium could selectively proliferate with hepatic functions (Nature Protocols, 2007) and this method could be applied for the isolation of human SHs from normal liver tissues, which were surgically resected from patients with informed consent. The number of human SHs increased about 100-fold within 3 weeks and the cells showed hepatic characteristics such as albumin secretion (Cell Transplant, 2008). Furthermore, we found that the proliferation of rat SHs was enhanced by follistatin produced by SHs to inhibit the action of activin A, which is secreted by mature hepatocytes. On the other hand, we succeeded in reconstructing functional hepatic tissues by stacking up two-dimensional tissues composed of rat SHs (FASEB J, 2006). In addition, we could culture normal rat biliary epithelial cells for a long term and reconstruct functional bile ductules between collagen gels (Am J Pathol, 2008).

我们报道CD44在大鼠小肝细胞中特异性表达，并且CD44^+肝细胞短暂地出现在受损的大鼠肝脏中。使用特定的抗体，我们从半乳糖胺处理的大鼠肝脏中分选 CD44^+ 细胞并培养它们。细胞的表型和基因表达与培养的SH相似，并且^+细胞可以分化为肝细胞（J Hepatol，2006）。此外，半乳糖胺处理的肝脏中出现的一些Thy1^+细胞可以分化为+SH并成熟为肝细胞。当Thyl+和CD44^+细胞被移植到用retrorsine/PH处理的肝脏中时，它们可以存活并增殖，在受体肝脏中形成肝病灶。此外，当培养的 SH 被移植到经过放射/PH 处理的同源大鼠肝脏中时，细胞迁移到肝小梁中，并且肝脏部分地被供体细胞重新填充（肝脏移植，2006）。在透明质酸（HA；CD44 配体）包被的培养皿和无血清培养基中培养的大鼠 SH 可以选择性地增殖并具有肝功能（Nature Protocols，2007），并且该方法可用于从正常肝组织中分离人 SH。这些是在知情同意的情况下通过手术从患者身上切除的。人类 SH 的数量在 3 周内增加了约 100 倍，并且细胞显示出肝脏特征，例如白蛋白分泌（Cell Transplant，2008）。此外，我们发现SH产生的卵泡抑素抑制成熟肝细胞分泌的激活素A的作用，从而增强大鼠SH的增殖。另一方面，我们通过堆叠由大鼠SH组成的二维组织成功地重建了功能性肝组织（FASEB J，2006）。此外，我们可以长期培养正常大鼠胆管上皮细胞，并在胶原凝胶之间重建功能性胆管（Am J Pathol，2008）。