Cheng Zanzan, Ou Ya, Zhang Lili, Zhang Pingshu, Yuan Xiaodong, Peng Wei, Wang Shujuan, Zhu Xuhong, Zhang Liping, Meng Yan
Department of Neurology, Affiliated Kailuan General Hospital of North China University of Science and Technology, Tangshan 063000, Hebei Province, China.
Key Laboratory of Neurological and Biological Function of Hebei Province, Tangshan 063000, Hebei Province, China; Key Laboratory of Neurology of Tangshan, Tangshan 063000, Hebei Province, China.
Neurosci Lett. 2018 Jun 11;677:94-102. doi: 10.1016/j.neulet.2018.04.048. Epub 2018 Apr 25.
ADSCs-derived astrocytes qualify the morphology, ultrastructure and membrane electrical potential, which are all unique to astrocytes. But whether they have the glutamate clearance function like mature astrocytes is under exploration. ADSCs were extracted, cultured and induced into astrocytes for 48 h, 7d, 14d and 21d in vitro. Inverted phase contrast microscope was used to observe the morphology of the cells in each group. Immunocytochemistry assay, immunofluorescence assay and Western blotting were used to detect the expression of GFAP, EAAT2 and GS of the cells in each group. The cells were cultured in glutamate solution for 1, 2, 3 and 4 h respectively before the solution collected. The glutamate concentration of the solution was detected using Glutamate Colorimetric Assay Hit. ADSCs-derived astrocytes expressed GFAP, EAAT2 and GS, all of which increased gradually and reached peak when induced for 14 days. In induction for 48 h, 7d and 14d groups, the extracellular glutamate concentration decreased gradually during the cells cultured in glutamate solution for 1, 2, 3 and 4 h, among which the decrease extent was most prominent in 14d group, while the extracellular glutamate concentration had no change in uninduction and induction for 21d group. ADSCs-derived astrocytes expressed EAAT2 and GS, meanwhile had the function of clearing glutamate, which was prominent when induced into astrocytes for 7-14 days.
脂肪来源干细胞分化的星形胶质细胞具备星形胶质细胞特有的形态、超微结构和膜电位。但它们是否具有像成熟星形胶质细胞那样的谷氨酸清除功能仍在探索中。体外提取、培养脂肪来源干细胞并将其诱导分化为星形胶质细胞，诱导时间分别为48小时、7天、14天和21天。采用倒置相差显微镜观察各组细胞形态。运用免疫细胞化学检测、免疫荧光检测和蛋白质免疫印迹法检测各组细胞中胶质纤维酸性蛋白（GFAP）、兴奋性氨基酸转运体2（EAAT2）和谷氨酰胺合成酶（GS）的表达。在收集培养液前，将细胞分别置于谷氨酸溶液中培养1、2、3和4小时。采用谷氨酸比色法试剂盒检测培养液中谷氨酸浓度。脂肪来源干细胞分化的星形胶质细胞表达GFAP、EAAT2和GS，三者表达量均逐渐增加，并在诱导14天时达到峰值。在诱导48小时、7天和14天的组中，细胞在谷氨酸溶液中培养1、2、3和4小时期间，细胞外谷氨酸浓度逐渐降低，其中14天组降低幅度最为显著，而未诱导组和诱导21天组的细胞外谷氨酸浓度无变化。脂肪来源干细胞分化的星形胶质细胞表达EAAT2和GS，同时具有清除谷氨酸的功能，在诱导分化为星形胶质细胞7至14天时该功能尤为突出。
