小鼠胚芽干细胞的分离培养和向肝细胞定向诱导分化的实验研究

发布时间：2019-04-29 07:18

【摘要】：自从1998年Thomson和Gearhart分别从胚泡内细胞群和原始生殖细胞成功地建立了人类胚胎干细胞系以来,干细胞研究已成为当今生命科学领域中的前沿和热点。研究内容主要涉及胚胎干细胞的培养建系、诱导干细胞向特定组织细胞的分化及在生物和医学中的应用。来源于原始生殖细胞(Primordial Germ Cell,PGC)的胚胎干细胞(embryonic stem cell,ESC),又称为胚芽干细胞(Embryonic Germ Cell,EGC),具有与内细胞群来源的胚胎干细胞相同的生物学特性。本文系统地研究了小鼠胚芽干细胞的提取、体外扩增;胚芽干细胞向肝细胞的定向诱导分化;和胚芽干细胞肝内移植的实验研究。一、胚芽干细胞的提取和体外扩增: 胚芽干细胞的提取和培养是进行深入研究的前提,我们首先进行了培养体系的筛选(见系列论文的第一部分)。胚芽干细胞与来源于内细胞群的胚胎干细胞一样,在体外培养过程中,极易发生分化。为了在体外维持胚胎干细胞的增殖和未分化状态,建立适宜的培养体系是关键。胚胎干细胞对体外培养条件要求非常严格,不同种属动物的胚胎干细胞要求的培养条件也不同。近年来对来源于内细胞群的胚胎干细胞的培养条件和方法进行了广泛的研究,积累了丰富的经验。但是有关胚芽干细胞的培养方法报道较少。本研究对适用于昆明种小鼠胚芽干细胞的体外培养体系进行了筛选。观察了基础培养液、添加白血病抑制因子、同种胎鼠成纤维细胞条件培养液、丝裂霉素处理的饲养层培养和与少量腹壁组织成纤维细胞共培养等
[Abstract]:Since Thomson and Gearhart successfully established human embryonic stem cell lines from blastocyst cell population and primordial germ cells in 1998, stem cell research has become the frontier and hot spot in the field of life sciences. The research mainly involves the establishment of embryonic stem cells, inducing stem cells to differentiate into specific tissues and cells, and its application in biology and medicine. Embryonic stem cells (embryonic stem cell,ESC), also known as embryonic stem cells (Embryonic Germ Cell,EGC), derived from primordial germ cells (Primordial Germ Cell,PGC), have the same biological characteristics as embryonic stem cells derived from inner cell group. In this paper, we systematically studied the extraction and expansion of mouse embryonic stem cells, the directional differentiation of embryonic stem cells into hepatocytes, and the experimental study of intrahepatic transplantation of embryonic stem cells. First, the extraction and in vitro expansion of germ stem cells: the extraction and culture of germ stem cells is the prerequisite for in-depth research. We first screened the culture system (see the first part of a series of papers). Like embryonic stem cells derived from inner cell group, embryonic stem cells are easy to differentiate in vitro. In order to maintain the proliferation and undifferentiation of embryonic stem cells in vitro, it is the key to establish an appropriate culture system. Embryonic stem cells require very strict culture conditions in vitro, and different kinds of animal embryonic stem cells require different culture conditions. In recent years, the culture conditions and methods of embryonic stem cells derived from inner cell group have been studied extensively, and abundant experience has been accumulated. However, there are few reports about the culture methods of germ stem cells. In this study, the in vitro culture system suitable for Kunming mouse germ stem cells was screened. The basic culture medium, adding leukemia inhibitory factor, conditioned medium of the same fetal rat fibroblast, feeder layer culture treated with mitomycin and co-culture with a small number of tissue fibroblasts of abdominal wall were observed.
【学位授予单位】：山东大学
【学位级别】：博士
【学位授予年份】：2005
【分类号】：R329

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