牛肌肉卫星细胞向胰岛素分泌细胞的诱导转分化研究

发布时间：2018-02-14 12:27

本文关键词： 肌肉卫星细胞定向分化胰腺细胞胰岛素分泌　出处：《农业生物技术学报》2016年02期 　论文类型：期刊论文

【摘要】：肌肉卫星细胞是源自中胚层的肌源干细胞,具有增殖分化融合成肌管并形成肌细胞的能力。在体内条件下,肌肉卫星细胞不可能跨胚层分化为内胚层来源的胰腺细胞。本研究从牛(Bos taurus)胎儿肌肉中分离培养得到肌肉卫星细胞,并在体外条件下将其诱导成胰岛素分泌细胞。在诱导过程中,分析了与胰腺发育相关的胰十二指肠同源基因盒1基因(pancreatic and duodenal homeobox 1,PDX1)、神经原素3基因(neurogenin 3,NGN3)、淀粉酶基因(Amylase)和胰岛素基因(Insulin,INS)的动态表达情况。结果表明,PDX1作为决定胰腺分化发育和胰岛功能的主要调节基因,在诱导分化的第2天能够检测到其m RNA的表达,在第3天表达量达到了最大,第4天以后维持在较低水平。NGN3是内分泌细胞形成非常重要的转录因子,是能够将胰岛素分泌到胞外的关键基因,其m RNA在分化诱导的第3天开始表达,第4天达到最高水平,之后逐渐下降。Amylase是胰腺发育中细胞外分泌相关的基因,Amylase m RNA在第6天才开始表达,8 d后达到最高。INS m RNA表达量在11~12 d最高,酶联免疫吸附分析(enzyme-linked immunosorbent assay,ELISA)发现,诱导形成的胰腺细胞能够正常分泌胰岛素,培养液中胰岛素的浓度在诱导的11~12 d达到最高值。向培养液中添加葡萄糖后,可使细胞的胰岛素分泌量显著提高,说明诱导得到的细胞类似于成体胰岛的功能,其胰岛素分泌受外界葡萄糖浓度的调节。上述结果表明,肌肉卫星细胞可以被诱导转分化为胰岛素分泌细胞,并对培养环境中的葡萄糖刺激做出应答反应。本研究结果为进一步的研究和临床应用提供基础资料。
[Abstract]:Muscle satellite cells are myogenic stem cells derived from the mesoderm, which have the ability to proliferate, differentiate, and fuse into muscle tubes and form muscle cells. It is impossible for muscle satellite cells to differentiate into pancreatic cells derived from endoderm. In this study, muscle satellite cells were isolated and cultured from Bos taurus fetal muscles. It was induced into insulin-secreting cells in vitro. The dynamic expression of pancreaticoduodenal homologous gene cassette 1, neurogenin 3 (NGN3), amylase gene (Amylase) and insulin gene insulinine (ins) related to pancreatic development were analyzed. The results showed that PDX1 was the determinant of pancreas. The major regulatory genes for differentiation and development and islet function, The expression of m RNA was detected on the second day of differentiation, and reached the maximum on the third day, and maintained at a low level after 4 days. NGN3 was a very important transcription factor in endocrine cell formation. It is the key gene that can secrete insulin to extracellular, and its m RNA expression begins on the 3rd day of differentiation induction and reaches the highest level on the 4th day. After that, the expression of Amylase m RNA, a gene related to exocrine secretion of pancreatic cells, began to express at the 6th day and reached the highest level at 1112 days after the expression of Amylase m. Enzyme-linked immunosorbent assayayer Elisa revealed that the highest expression level of Amylase m was found by enzyme-linked immunosorbent assayay assay (Elisa) on the 6th day, and then reached the highest level at 1112 days after the expression of Amylase m. The pancreatic cells were able to secrete insulin normally, and the concentration of insulin in the culture medium reached the highest level on the 12th day after induction. After glucose was added to the culture medium, the insulin secretion of the cells increased significantly. These results indicate that the induced cells are similar to the adult islets, and their insulin secretion is regulated by the concentration of external glucose. These results suggest that the muscle satellite cells can be induced to differentiate into insulin-secreting cells. The results provide basic data for further research and clinical application.
【作者单位】：内蒙古大学哺乳动物生殖生物学与生物技术教育部重点实验室;内蒙古医科大学基础医学院生理教研室;
【基金】：内蒙古自治区科技厅重大基础研究开放课题(No.20130902)
【分类号】：S852.2

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