Mechanisms of differentiation of mesenchymal stem cells and their functional roles in diseases

为了阐明间充质细胞的分化机制，采用未分化间充质干细胞系(MT-9，TSD-B9)进行了以下实验。MT-9是从大鼠恶性纤维组织细胞瘤中分离出来的具有多向分化潜能的细胞系，TSD-B9是从大鼠恶性畸胎瘤中分离出来的细胞系，其来源是性腺生殖细胞，具有向内胚层、中胚层和外胚层组织分化的能力。1.成骨分化：在这些细胞系中加入包括BMP-2在内的成骨分化补充剂，使钙化面积增加，von Kossa染色显示，培养上清液中碱性磷酸酶升高；这些是成骨细胞分化的指标。2.脂肪母细胞分化：在这些细胞系中加入包括吲哚美辛、地塞米松和胰岛素在内的脂肪母细胞分化补充剂，增加了细胞质中含有油红O染色脂质的细胞数量。3.肌成纤维细胞分化：纤维化疾病中的纤维化因子转化生长因子-β1处理后，对α-平滑肌肌动蛋白(α-SMA)免疫反应的细胞数增加，Western blotting显示这种增加。5.这些研究部分阐明了间充质分化的机制，这些细胞系可用于研究间充质分化的机制。6.此外，从大鼠皮肤下恶性纤维组织细胞瘤中建立了一个新的细胞系，该细胞系具有组织细胞性质，另一个细胞系从大鼠宫颈恶性神经鞘瘤中分离出来。这些细胞系也将成为间充质分化研究的有利工具。

To clarify the differential mechanisms of mesenchymal cells, the following experiments were performed using undifferentiated mesenchymal stem cell lines (MT-9, TSD-B9). MT-9 is a cell line isolated from a rat malignant fibrous histiocytoma having pluripotential differentiation, and TSD-B9 is a cell line derived from a rat malignant teratoma, of which the origin is gonadal germ cells capable of differentiating towards endodermal, mesodermal and ectodermal tissues.1.Osteoblastic differentiation : The addition of a supplement for osteoblastic differentiation, including BMP-2, to these cell lines increased calcified areas, demonstrable by von Kossa stain, and showed an elevated alkaline phosphatase in supernatants ; these are indicators of osteoblastic differentiation.2.Lipoblastic differentiation : The addition of a supplement for lipoblastic differentiation, including indomethacin, dexamethasone and insulin, to these cell lines increased the number of cells having oil red O-stainable lipids in the cytoplasm. The stain is useful for detection of lipoblastic cells.3.Myofibroblastic differentiation : The treatment with TGF-β1, a fibrogenic factor in fibrotic diseases, increased the number of cells immuno-reacting to α-smooth muscle actin (α-SMA) ; the increase was demonstrated by the western blotting. α-SMA-expressing cells are regarded as myofibroblasts.5.These studies partly clarified the mechanisms of mesenchymal differentiation, and these cell lines would prove useful for investigating the mechanisms.6.Furthermore, a new cell line was established from a rat malignant fibrous histiocytoma arising in the subcutis, showing histiocytic nature, and another cell line was isolated from a rat malignant schwannoma generating from the uterine cervix. These cell lines would become also beneficial tools for studies on mesenchymal differentiation.