端粒酶逆转录酶基因修饰人骨髓间充质干细胞的实验研究
发布时间：2018-10-13 19:34
【摘要】：人骨髓中除造血干细胞(hematopoietic stem cell,HSC)外还存在另外一类干细胞—骨髓间质干细胞(mesenchymal stem cell,MSC),是中胚层发育的早期细胞。新近研究发现MSC不仅具有向中胚层来源的细胞(如:成骨细胞、软骨细胞、脂肪细胞、成肌细胞和心肌细胞)发育的潜能,而且具有向中胚层外细胞(如神经细胞、肝细胞、内皮细胞等)分化的潜能。 尽管如此,但由于骨髓有核细胞中平均每10~6个细胞中只有1个MSC,且人MSCs和其它正常体细胞一样,具有有限的生命,在经过有限次的细胞传代后,就会停止增殖,发生衰老和死亡,这就限制了MSCs在科研与临床的进一步应用,所以如何实现MSCs在体外既长期扩增又保持多潜能分化状态是当前亟待解决的问题。 研究表明,端粒与细胞寿命的控制密切相关。随着细胞分裂次数增加,端粒进行性缩短,当缩短到一定限度即不能维持染色体的稳定时,细胞失去分裂能力,进而衰老死亡。而端粒长度的维持需要端粒酶的激活,端粒酶是一种核糖核蛋白,由RNA和蛋白质组成,具有逆转录的活性,其活性主要由端粒酶逆转录酶(human telomerase reverse transcriptase,hTERT)表达水平的高低决定。hTERT是端粒酶的催化亚单位,在端粒酶的激活中起关键作用。 鉴于上述,本实验旨在将携带有EGFP报告基因和hTERT目的基因的质粒pEGFP-hTERT通过阳离子脂质体转染法转入人骨髓MSCs中,设想通过外源性hTERT基因的异位表达,诱导人骨髓MSCs的端粒酶活性,维持端粒长度的稳定,从而延长人骨髓MSCs的生命周期并保持其多潜能分化特性。
[Abstract]:In addition to hematopoietic stem cell (hematopoietic stem cell,HSC), there is another kind of stem cell, bone marrow mesenchymal stem cell (mesenchymal stem cell,MSC), which is the early stage of mesoderm development. Recent studies have found that MSC not only has the potential to develop to mesodermal cells (such as osteoblasts, chondrocytes, adipocytes, myoblasts and cardiomyocytes), but also to mesodermal extracellular cells (such as nerve cells, hepatocytes). The potential for differentiation of endothelial cells, etc. Nevertheless, because of the fact that there is on average only one MSC, per 10 to 6 cells in nucleated bone marrow cells and that human MSCs, like other normal somatic cells, has limited life, it will cease to proliferate after a limited number of cell passages. Aging and death limit the further application of MSCs in scientific research and clinic. Therefore, how to realize the long-term expansion of MSCs in vitro and maintain the state of multipotential differentiation is an urgent problem to be solved. Studies have shown that telomere is closely related to the control of cell life. With the increase of cell division times, telomere gradually shortened. When the telomere was shortened to a certain limit, it could not maintain the stability of chromosome, the cell lost the ability of division and then died of senescence. The maintenance of telomere length requires the activation of telomerase, a ribonucleoprotein that is composed of RNA and proteins and has the activity of reverse transcription. Its activity is mainly determined by the level of telomerase reverse transcriptase (human telomerase reverse transcriptase,hTERT) expression. HTERT is the catalytic subunit of telomerase and plays a key role in the activation of telomerase. In view of the above, the aim of this study was to transfer the plasmid pEGFP-hTERT carrying EGFP reporter gene and hTERT target gene into human bone marrow MSCs by cationic liposome transfection. It was envisaged that the telomerase activity of human bone marrow MSCs could be induced by heterotopic expression of exogenous hTERT gene. To maintain the stability of telomere length, prolong the life cycle of human bone marrow MSCs and maintain its multipotential differentiation characteristics.
【学位授予单位】：郑州大学
【学位级别】：硕士
【学位授予年份】：2005
【分类号】：R329.2
本文编号：2269680
[Abstract]:In addition to hematopoietic stem cell (hematopoietic stem cell,HSC), there is another kind of stem cell, bone marrow mesenchymal stem cell (mesenchymal stem cell,MSC), which is the early stage of mesoderm development. Recent studies have found that MSC not only has the potential to develop to mesodermal cells (such as osteoblasts, chondrocytes, adipocytes, myoblasts and cardiomyocytes), but also to mesodermal extracellular cells (such as nerve cells, hepatocytes). The potential for differentiation of endothelial cells, etc. Nevertheless, because of the fact that there is on average only one MSC, per 10 to 6 cells in nucleated bone marrow cells and that human MSCs, like other normal somatic cells, has limited life, it will cease to proliferate after a limited number of cell passages. Aging and death limit the further application of MSCs in scientific research and clinic. Therefore, how to realize the long-term expansion of MSCs in vitro and maintain the state of multipotential differentiation is an urgent problem to be solved. Studies have shown that telomere is closely related to the control of cell life. With the increase of cell division times, telomere gradually shortened. When the telomere was shortened to a certain limit, it could not maintain the stability of chromosome, the cell lost the ability of division and then died of senescence. The maintenance of telomere length requires the activation of telomerase, a ribonucleoprotein that is composed of RNA and proteins and has the activity of reverse transcription. Its activity is mainly determined by the level of telomerase reverse transcriptase (human telomerase reverse transcriptase,hTERT) expression. HTERT is the catalytic subunit of telomerase and plays a key role in the activation of telomerase. In view of the above, the aim of this study was to transfer the plasmid pEGFP-hTERT carrying EGFP reporter gene and hTERT target gene into human bone marrow MSCs by cationic liposome transfection. It was envisaged that the telomerase activity of human bone marrow MSCs could be induced by heterotopic expression of exogenous hTERT gene. To maintain the stability of telomere length, prolong the life cycle of human bone marrow MSCs and maintain its multipotential differentiation characteristics.
【学位授予单位】：郑州大学
【学位级别】：硕士
【学位授予年份】：2005
【分类号】：R329.2
【参考文献】
相关期刊论文 前1条
1 冯凯,裴雪涛;间充质干细胞——现代组织工程的新资源[J];国外医学.生物医学工程分册;2000年06期
，本文编号：2269680
本文链接：https://www.wllwen.com/yixuelunwen/binglixuelunwen/2269680.html
最近更新
教材专著
