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体外诱导小鼠脂肪来源干细胞向内皮细胞分化的实验研究 被引量：1

Experimental Research on the Differentiation of Induced Mice Adipose Stem Cells to Endothelial Cells In Vitro
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摘要 目的:探索体外小鼠脂肪来源干细胞(Adipose-derived stem cells,ASCs)诱导分化为内皮细胞(endothelial progenitor cells,EPCs)的可行性。方法:利用I型胶原酶消化法和传代培养纯化法从小鼠脂肪组织中分离、培养及扩增ASCs,通过流式细胞仪检测ASCs特异性表面抗原CD29和CD44、CD105的表达;取第2代ASCs进行内皮诱导:即BD基质胶包被+内皮细胞生长诱导培养基(M199+10%血清+10 ng/mL血管内皮细胞生长因子VEGF+10 ng/ml碱性生长因子bFGF)。诱导两周左右,倒置显微镜观察诱导前后细胞的一般形态学特征;同时通过成血管实验观察成管腔能力;通过免疫荧光鉴定CD31表型分子的表达。结果:成功培养小鼠ASCs;CD29、CD44、CD90、呈阳性表达,而CD31、CD34、CD45呈阴性表达;诱导后的细胞形态呈三角形或多边形;HE染色观察可见明显的管腔样结构;免疫荧光法CD31表达阳性;MTT法成功记录EPCs增殖生长曲线。结论:成功的诱导小鼠脂肪来源干细胞分化为血管内皮细胞,为后续体内移植实验提供理论基础,同时也为临床受损组织或器官的重建再生提供实验基础。 Objective： To explore the feasibility of the induced mice adipose stem cells（ASCs） differentiating into endothelial pro- genitor cells （EPCs） in vitro. Methods： We separated and digested the adipose stem cells from the adipose tissue of mice by means of the type I collagen enzyme, then we cultured and amplified the adipose stem cells. The stem cell phenotype CD29/CD44/CD105 were detected by the Flow Cytometer. The second generation of mice ASCs was induced to endothelial cells： BD matrigel ＋ growth＇induction medium of endothelial cells （M 199 ＋ 10% serum ＋ 10 ng/mL VEGF ＋ 10 ng/mL bFGF）. After two weeks, the cell morphology of induction was com- pared before and after inverting microscope. At the same time, the vascular experiment was carried on to observe wether the en- dothelial cells could form tube-like structure and to estimate the expression of CD31 by the immunofluorescence. Results： The mice＇ ASCs were cultured successfully which including the CD29（＋）, CD44（＋）, CD90（＋）, CD31 （-）, CD34（-） and CD45（-）. After the inductio, the cell mor- phology was changed into triangular or polygon. HE dyed observation showed that the cells were formed tube-like structure obviously. Immunofluorescence reflected that the CD31 was positive. The proliferation growth curve of EPCs was recorded by MTT method. Conclusion： It is successful that induced the mice ASCs to differentiate into EPCs, which could provide theoretical foundation for the subsequent transplantation experiment in vitro and lay the foundation for clinical regeneration and reconstruction of tissue or organ.
作者 胡翰青 邢红艳 马行健 刘蔚 刘彦普 王浈 刘斌
机构地区 解放军第九四医院口腔科 第四军医大学口腔医院颌面外科 第四军医大学口腔医院生物教研室及实验动物中心
出处 《现代生物医学进展》 CAS 2013年第25期4857-4860,共4页 Progress in Modern Biomedicine
基金 国家自然科学基金项目(31170942) 南京军区医药卫生科研项目(12MA048)
关键词 脂肪来源干细胞 体外诱导 内皮细胞 分化 鉴定 修复重建 Adipose stem cells Induced in vitro Endothelial cells Differentiation Identification Restoration
分类号 R782.7 [医药卫生—口腔医学]
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参考文献20

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同被引文献46

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现代生物医学进展
2013年 第25期

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