In vitro induction of pancreatic β-cell development from mouse embryonic stem cells

胚胎干细胞（ES细胞）是一种多能细胞，来源于受精囊胚的内部细胞群。胚胎干细胞显示出在体外分化成多种细胞系的能力。研究表明，胚胎干细胞可以分化为造血细胞、神经元细胞和心肌细胞。我们研究胚胎干细胞向分泌胰岛素的胰腺β细胞分化的可能性。为了促进胚胎干细胞向胰岛素生成细胞的分化，我们操纵胚胎干细胞稳定表达在胰腺和胰腺β细胞发育中起关键作用的转录因子IPF-1。在胚状体产生后，将产生IPF-1的胚胎干细胞在Lumelsky等人（Science 292: 1389）的无血清培养基中进行修饰，使其分化。产生IPF-1的胚胎干细胞来源的细胞产生的免疫反应性胰岛素比天然胚胎干细胞来源的分化细胞多5倍。我们通过免疫组织化学分析证实，在最终分化阶段，大约30%的细胞表达免疫反应性胰岛素。我们目前正在努力优化条件，以实现更有效的分化。同时，我们详细阐述了EGFP标记的ES细胞，其中EGFP可以在胰岛素产生细胞中特异性表达。分化后，胰岛素分泌细胞将能够被荧光辅助细胞分选使用这些细胞。

Embryonic stem cells (ES cells) are pluripotent cells derived from the inner cell mass of fertilized blastocysts. ES cells display the ability to differentiate in vitro into a variety of cell lineages. It has been shown that ES cells can be differentiated to hematopoietic cells, neurons and cardiomyocytes. We investigate the possibility to differentiate the ES cells to insulin secreting pancreatic β-cells. In order to facilitate the differentiation to the insulin producing cells, we manipulated ES cells to stably express IPF-1, a transcription factor which plays critical roles in the development of pancreas and pancreatic β-cells. After production of embryoid bodies, IPF-1 producing ES cells were rendered to differentiate in the serum free medium of Lumelsky et al (Science 292: 1389) with modification. IPF-1 producing ES cell-derived cells produced up to 5 times more immunoreactive insulin when compared with the differentiated cells derived from native ES cells. We confirmed the expression of immunoreactive insulin in approximately 30 % of the cells at final differentiation stage by immunohistochemical analysis. We are currently trying to optimize the condition for more efficient differentiation. At the same time, we have elaborated the ES cells marked by EGFP in which EGFP can be expressed specifically in the insulin-producing cells. Upon the differentiation, insulin secreting cells will be able to be selected by fluorescence-assisted cell sorting using these cells.