重组人成纤维细胞生长因子10间充质干细胞诱导人脂肪干细胞分化的研究

发布时间：2018-04-10 17:01

本文选题：脂肪干细胞 + 人纤维细胞生长因子10　；参考：《第二军医大学》2007年硕士论文

【摘要】： 目的:研究人成纤维细胞生长因子10(FGF10)诱导脂肪干细胞分化的机制,并为临床以增脂为主要手段的组织工程和临床整形美容及创伤修复提供思路和方法。方法:1、分离,培养从外科手术术中物理分离得到的脂肪干细胞,并通过定向分化诱导等方法鉴定其干细胞特性。2、从人肺组织中抽提总RNA,并利用RT-PCR扩增FGF10,构建pcDNA3.0-FGF10重组质粒,并将其转染至MSC中,利用IHC, ELISA, RT-PCR和western-blotting的方法鉴定pcDNA3.0-FGF10-MSC表达目的蛋白FGF10的情况。3、利用pcDNA3.0-FGF10-MSC与ADSC体外共培养及裸鼠体内注射细胞的方法证明FGF10的促成脂作用,通过显微镜观察细胞改变及RT-PCR和real-time PCR监测ADSC分化过程中转录因子的变化。通过取裸鼠皮下组织切片染色及免疫组化证实FGF10的体内作用。结果:1、我们培养的ADSC具有干细胞特性,并能多向诱导分化。2、构建的pcDNA3.0-FGF10-MSC表达载体能持续表达目的蛋白FGF10。3、体外共培养实验发现ADSC细胞内脂滴增多,分化转录因子表达增高。裸鼠体内实验发现皮下脂肪层明显增厚。结论:FGF10能在体外及体内促进脂肪干细胞的成熟分化。
[Abstract]:Objective: to study the mechanism of adipose stem cell differentiation induced by human fibroblast growth factor (FGF10), and to provide ideas and methods for clinical tissue engineering, cosmetic surgery and wound repair.Methods the adipose stem cells were isolated and cultured during surgery. The stem cell characteristics of the stem cells were identified by directional differentiation induction. The total RNAs were extracted from human lung tissue. FGF10 was amplified by RT-PCR, and the recombinant plasmid of pcDNA3.0-FGF10 was constructed.The expression of pcDNA3.0-FGF10-MSC protein FGF10 was identified by IHC, Elsa, RT-PCR and western-blotting. The effect of FGF10 on lipid production was proved by co-culture of pcDNA3.0-FGF10-MSC and ADSC in vitro and injection of cells in nude mice.The changes of transcription factors in the differentiation of ADSC were observed by microscope and the changes of transcription factors during ADSC differentiation were monitored by RT-PCR and real-time PCR.The effect of FGF10 in vivo was confirmed by immunohistochemical staining and staining of subcutaneous tissue sections of nude mice.Results: the ADSC cultured in vitro had stem cell characteristics and was able to induce differentiation. The constructed pcDNA3.0-FGF10-MSC expression vector could continuously express the target protein FGF10.3. In vitro co-culture experiments showed that the lipid droplets in ADSC cells increased and the expression of differentiation transcription factors increased.The subcutaneous adipose layer was obviously thickened in nude mice.Conclusion: FGF10 can promote the maturation and differentiation of adipose stem cells in vitro and in vivo.
【学位授予单位】：第二军医大学
【学位级别】：硕士
【学位授予年份】：2007
【分类号】：R329

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