Chen Ye, Dong Xue-Jun, Zhang Guo-Rong, Shao Jian-Zhong, Xiang Li-Xin
College of Life Sciences, Zhejiang University, Hangzhou 310058, P.R. China.
J Cell Biochem. 2007 Sep 1;102(1):52-63. doi: 10.1002/jcb.21275.
The differentiation potential of adult stem cells has long been believed to be limited to the tissue or germ layer of their origin. However, recent studies have demonstrated that adult stem cells may encompass a greater potential than once thought. In the present study, we examined whether murine bone marrow derived stromal stem cells (BMSSCs) are able to differentiate into functional hepatocytes in vitro. BMSSCs were isolated from murine femora and tibiae, and the mesodermal multilineage differentiation potentials of these cells were functionally characterized. To effectively induce hepatic differentiation, we designed a novel protocol by using hepatocyte-conditioned medium. Hepatic differentiation from mouse BMSSCs was examined by a variety of assays at morphological and molecular levels. Morphologically, mouse BMSSCs became round and epithelioid, binucleated after induction. Differentiated cells were harvested on Days 0, 10, and 20 and subjected to examination of hepatocyte characteristics by reverse transcription-polymerase chain reaction (RT-PCR) and immunocytochemistry. We detected AFP, HNF-3beta, CK19, CK18, ALB, TAT, and G-6-Pase at the mRNA and/or protein levels, hepatocyte-like cells by culture in conditioned medium further demonstrated in vitro functions characteristic of liver cells, including glycogen storage, and urea secretion. Moreover, transplantation of the differentiated cells into liver-injured mice partially restored serum albumin level and significantly suppressed transaminase activity. Our findings indicated the transdifferentiation potential of mouse BMSSCs developing into the functional hepatocyte-like cells by conditioned culture medium and, hence, may serve as a model system for the study of mechanisms involved in the transdifferentiation, and a cell source for cell therapy of hepatic diseases.
长期以来，人们一直认为成体干细胞的分化潜能仅限于其起源的组织或胚层。然而，最近的研究表明，成体干细胞可能具有比以往认为的更大的潜能。在本研究中，我们检测了小鼠骨髓来源的基质干细胞（BMSSCs）在体外是否能够分化为功能性肝细胞。从鼠的股骨和胫骨中分离出BMSSCs，并对这些细胞的中胚层多谱系分化潜能进行了功能鉴定。为了有效诱导肝脏分化，我们通过使用肝细胞条件培养基设计了一种新的方案。通过多种形态学和分子水平的检测方法，研究了小鼠BMSSCs的肝脏分化情况。形态学上，诱导后小鼠BMSSCs变成圆形、上皮样，具有双核。在第0、10和20天收获分化细胞，并通过逆转录聚合酶链反应（RT-PCR）和免疫细胞化学检测肝细胞特征。我们在mRNA和/或蛋白质水平检测到甲胎蛋白（AFP）、肝细胞核因子-3β（HNF-3β）、细胞角蛋白19（CK19）、细胞角蛋白18（CK18）、白蛋白（ALB）、酪氨酸转氨酶（TAT）和葡萄糖-6-磷酸酶（G-6-Pase），通过在条件培养基中培养进一步证明了肝细胞样细胞具有肝细胞的体外功能特征，包括糖原储存和尿素分泌。此外，将分化细胞移植到肝损伤小鼠体内可部分恢复血清白蛋白水平，并显著抑制转氨酶活性。我们的研究结果表明，通过条件培养基培养，小鼠BMSSCs具有转分化为功能性肝细胞样细胞的潜能，因此，可作为研究转分化机制的模型系统，以及肝脏疾病细胞治疗的细胞来源。
