Research on differentiation of endodermal cells from ES cells by introducing genes of transcription factor

ES细胞被证明是多能的，被认为是再生医学的良好材料。我们的研究重点是内胚层细胞的分化。由于转录因子参与分化的基因很多，我们试图通过在ES细胞中导入基因来诱导分化。Sox17是内胚层早期发育和肝脏形成的关键基因之一。在ES细胞中强制表达Sox17基因会导致一组在内胚层表达的基因的诱导。此外，地塞米松还能诱导Sox17表达细胞中白蛋白基因的表达。为了研究表达Sox17的ES细胞在体内的分化，我们在表达Sox17的ES细胞中引入了SAP(血清淀粉样蛋白)下的EGFP表达单位。将这些ES细胞经脾静脉注入肝脏1个月后，在肝脏内可见表达EGFP的ES细胞。为了分析Sox17基因在ES细胞分化中的作用，我们建立了四环素可调控Sox17基因表达的ES细胞系。高密度培养结合Sox17的强制表达可诱导原始内胚层细胞在Wnt信号调控下分化。用同样的方法建立胰腺发育的关键基因PDX-1基因表达可调控的ES细胞，可以高效地获得胰岛素表达细胞，这些结果将有助于了解内胚层的早期发育以及再生医学的进一步发展。

ES cells are proved to be pluripotent and are postulated as good materials for regenerative medicine. Focus of our research is on the differentiation of endodermal cells. As the numerous genes for transcription factor are known to involve in the differentiation, we try to induce the differentiation by introducing the genes in ES cells.Sox17 is on of the crucial genes for development of early stage of endoderm and liver formation. Forced expression of sox17 gene in ES cells results the induction of a set of genes expressed in the endoderm. Moreover dexamethason induce the expression of albumin gene in the sox17-expressing cells. To investigate the in vivo differentiation of sox17-expressing ES cells, we have introduced the EGFP expression unit under SAP (serum amyloid protein) into sox17-expressing ES cells. After one month of injection of these ES cells into liver through splenic vein, EGFP expressing ES cells were found in the liver. Thus sox17-expressing ES cells have suggested to differentiate into mature hepatocyte in vivo.To analyze the effect of sox17 gene in the differentiation of ES cells, we have established the ES cell line in which expression of sox17 gene were regulatable by tetracycline. High density culture in combination with forced expression of sox17 leads to differentiate primitive endoderm cells under regulation of Wnt signals. Using the same strategy to establish the ES cells in which expression of pdx-1 gene, one of the crucial genes for development of pancreas, was regulatable, we can obtain insulin expression cells efficiently.These results will contribute to the understanding of early development of endoderm as well as further progress of regenerative medicine.