摘要
我们用建系的小鼠胚胎干细胞ES-5细胞为材料,研究了维生素A酸(RA)和双丁酰基环腺苷单磷酸(dBcAMP)对该胚胎干细胞的体外诱导分化。在ES-5细胞单层培养的条件下,RA单独作用或RA与dBcAMP共同作用,都产生神经元样和成纤维样两种细胞;在后一种作用情况下分化细胞可达90—95%,通过对细胞形态特征表型标志(GFAP,层粘蛋白等)的分析,初步证明绝大部分细胞为神经胶质细胞。除单层培养外,我们还研究了RA对ES-5细胞聚集体贴壁培养的诱导分化,在这种条件下,分化细胞类型较多,其是最突出的是有节律收缩的心肌样细胞。本文讨论了ES-5细胞单层培养与细胞聚集体贴壁培养两种条件下,RA诱导的差异及其可能原因。
In vitro induced differentiation of mouse embryonic stem cells (ES-5 cells), derived from 5-day 129 mouse blastocyst was studied with retinoic acid (RA) and dibutyryl cyclic adenosine monophosphate (dBcAMP). RA only or RA with dBcAMP together can both induce monolayer ES-5 cells to differentiate into cells of two types: neuron-like cells and fibroblast-like cells. After treated with 10^(-6)mol/L RA for 6 days, the differentiated cells were about 80% of all cells, among which most cells were fibroblast-like cells and others were neuron-like cells. While after 6 days of treatment with 10^(-8)mol/L RA and 1 mmol/ L dBcAMP, the ratio of differentiated cells can be up to 90—95%, and most cells (about 90—95% of differentiated cells) are neuron-like cells. Immunocytochemical analysis of phenotypic markers, especially GFAP and laminin, showed that the neuronlike cells were glia cells. DBcAMP affected the direction and efficiency of induction by RA. The induced differentiation by RA on attached aggregated ES-5 cells was studied as well. In this case, more cell types appeared, such as cpitheloid cells, fibroblast-like cells and spindle shaped cells and so on. The expect nature of these differentiated cells was not identified. After attacked culture for about 15 days, rhythmieally contracting cardiac-like muscle cell were most attractive among those several dilferentiated cell types.The change of phenotypic markers during induced differentiation of ES-5 cells in monolayer and aggregated state was summarized in table 1. Transforming growth factor-beta 1 (TGF-beta 1) was also examined in undifferentiated and differentiated cells. Untreated ES-5 cells showed positive immunofluorescent reaction to TGF-beta 1 and various differentiated cells showed different reactions. Glia cells and cardiac-like cells displayed a much stronger TGF-bcta 1 reaction. These results indicete that the exact role plqyed by TGF-bcta 1 during induced differcntiation necds further investigation.The different effect of RA on monolayer and aggregated ES cells and the possible significance of cell to cell interaction in the latter case are discussed.
出处
《实验生物学报》
CSCD
1991年第4期353-367,共15页
Acta Biologiae Experimentalis Sinica
关键词
胚胎
细胞
分化
维生素A酸
DBCAMP
Embryonic stem cells. Retinoic acid. Dibutyryi cAMP. Induced differentiation.
