大鼠骨髓胚胎样干细胞生物学特性和分化潜能

发布时间：2018-11-21 19:29

【摘要】：背景:大鼠骨髓胚胎样干细胞能表达胚胎干细胞标志物,能够分化形成三胚层来源细胞,并且植入机体后能改善心功能、组织心室重构,但是对于骨髓胚胎样干细胞的生物学特性及分化潜能尚不完全知晓。目的:分析大鼠骨髓胚胎样干细胞生物特性和分化潜能。方法:20只SD大鼠,采用阶段特异性胚胎抗原1(SSAE-1)标记骨髓单核细胞,利用流式细胞仪分选骨髓胚胎样干细胞。采用电镜扫描观察骨髓胚胎样干细胞形态、结构;免疫荧光化学染色测定胚胎干细胞转录因子(Oct4、Nanog、Sox2)在骨髓胚胎样干细胞中的表达,将骨髓胚胎样干细胞放入纤维细胞饲养层进行扩增,诱导心肌和内皮细胞,测定其特异蛋白表达。结果与结论:(1)采用SSEA-1标记大鼠骨髓胚胎样干细胞,对直径为2-6μm细胞设门,大鼠骨髓胚胎样干细胞呈现纺锤形,多边形以及圆形多细胞形态;体积较小;(2)透射电镜下可见细胞核增大,细胞质较少,含有少量的细胞器。(3)第2代大鼠骨髓胚胎样干细胞免疫细胞化学染色下能测定到多潜能抗原标志;随传代次数增加,细胞增殖能力出现下降趋势;(4)大鼠骨髓胚胎样干细胞分化1周后心肌细胞平行排列,诱导2周后细胞形态发生明显的变化,细胞相互连接。免疫荧光化学染色可见心肌特异蛋白cTnT以及Cx43表达;大鼠骨髓胚胎干细胞分化1周后内皮细胞呈现多角形、梭形。诱导2周后细胞相互连接,免疫细胞化学染色可见内皮细胞特异蛋白VWF以及CD31表达。(5)结果提示:大鼠骨髓胚胎样干细胞具备与骨髓间充质干细胞类似的生物学特性,能表达干细胞转录因子,能够在一定的条件下分化为心肌细胞和内皮细胞,具有较高的临床应用价值。
[Abstract]:Background: rat bone marrow embryonic stem cells (BMSCs) can express embryonic stem cell markers, differentiate into tridermal derived cells, and improve cardiac function and ventricular remodeling after implantation. However, the biological characteristics and differentiation potential of bone marrow embryonic stem cells are not fully known. Objective: to analyze the biological characteristics and differentiation potential of rat bone marrow embryonic stem cells (BMSCs). Methods: bone marrow monocytes were labeled with stage specific embryonic antigen 1 (SSAE-1) in 20 SD rats and bone marrow embryonic stem cells were sorted by flow cytometry. The morphology and structure of bone marrow embryonic stem cells were observed by scanning electron microscope. The expression of embryonic stem cell transcription factor (Oct4,Nanog,Sox2) in bone marrow embryonic like stem cells was determined by immunofluorescence staining. Bone marrow embryonic stem cells were added to fibroblast feeder layer to induce myocardial and endothelial cells. Its specific protein expression was determined. Results and conclusion: (1) SSEA-1 was used to label embryonic stem cells of rat bone marrow, and the diameter of 2-6 渭 m cells was set up. The embryonic stem cells of rat bone marrow showed fusiform, polygonal and round multicellular morphology, and the size was small. (2) under the transmission electron microscope, the nucleus was enlarged, the cytoplasm was less and there were a few organelles. (3) the multipotential antigen markers could be detected by immunocytochemical staining of the second generation of rat bone marrow embryonic stem cells. With the increase of passage times, the proliferation of rat bone marrow embryonic stem cells decreased. (4) after 1 week of differentiation, the cardiomyocytes were arranged in parallel, and after 2 weeks of induction, the morphology of the cells changed obviously and the cells connected with each other. The expression of myocardial specific protein cTnT and Cx43 was observed by immunofluorescence staining, and the endothelial cells of rat bone marrow embryonic stem cells showed polygonal and fusiform shape after 1 week of differentiation. After 2 weeks of induction, the cells were connected to each other, and the expression of endothelial cell specific protein VWF and CD31 was observed by immunocytochemistry. (5) the results suggested that the embryonic stem cells of rat bone marrow had similar biological characteristics to those of mesenchymal stem cells. It can express stem cell transcription factors and can differentiate into cardiomyocytes and endothelial cells under certain conditions.
【作者单位】：白城医学高等专科学校;吉林大学基础医学院;
【分类号】：R54

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