Isolation of Murine Pancreatic Liver Stem Cells

描述(申请人提供)：许多器官在损伤后或正常周转期间依靠高度再生的干细胞进行组织更新。在胚胎发育过程中，共同的内胚层前体细胞/干细胞同时产生肝脏和胰腺实质，包括胰腺的导管、外分泌和内分泌成分。这种肝胰腺干细胞的分离和纯化将极大地促进对胰腺生物学的了解，并可能产生一种治疗糖尿病的药物。 造血干细胞是通过生物测试来鉴定的，即在致命辐射后重建宿主血液和免疫系统的能力。不幸的是，基于移植的体内胰腺干细胞再繁殖试验还没有开发出来。然而，几条独立的证据表明，成年小鼠胰腺中继续含有在某些实验条件下可以形成肝细胞的细胞。富马酸乙酰乙酸酯水解酶基因敲除小鼠代表了一种强大的肝脏再繁殖实验，在该实验中可以识别具有再生肝脏能力的细胞。初步数据显示，成年小鼠胰腺含有细胞，这些细胞具有很高的肝脏再生能力(胰腺肝干细胞)。我们推测，再生活性是一种原始的内胚层前体，能够产生多种类型的细胞，包括肝细胞、胆管、胰管、腺泡细胞和内分泌细胞。 本申请涉及使用肝脏再繁殖作为原始小鼠胰腺干细胞的替代试验。该检测方法将被用于开发它们的最佳分离方法。将产生可用于FACS胰腺细胞分选的单抗。我们将应用细胞分选方法来丰富胰腺肝干细胞。

DESCRIPTION (provided by applicant): Many organs depend on highly regenerative stem cells for tissue renewal after injury or during normal turnover. During embryonic development a common endodermal pre-cursor/stem cell gives rise to both the hepatic and pancreatic parenchyma, including the ductular, exocrine and endocrine components of the pancreas. The isolation and purification of this hepato-pancreatic stem cell would greatly advance the understanding of the biology of the pancreas and possibly generate a therapeutic agent for the treatment of diabetes. The hematopoietic stem cell was identified by a biological assay, the ability to reconstitute the blood and immune system of a host after lethal irradiation. Unfortunately, a transplantation based in vivo repopulation assay for pancreatic stem cells has not been developed. However, several independent lines of evidence suggest that adult mouse pancreas continues to contain cells which can give rise to hepatocytes under certain experimental conditions. The fumarylacetoacetate hydrolase knockout mouse represents a robust liver repopulation assay in which permits the identification of cells that have the capacity to regenerate the liver. Preliminary data have shown that adult murine pancreas contains cells, which have high liver repopulating capacity (pancreatic liver stem cells). We hypothesize that the repopulating activity is a primitive endodermal precursor capable of giving rise to multiple cell types including hepatocytes, bile ducts, pancreatic ducts, acinar cells and endocrine cells. This application pertains to the use of liver repopulation as a surrogate assay for primitive murine pancreatic stem cells. The assay will be used to develop methods for their optimal isolation. Monoclonal antibodies useful for FACS sorting of pancreatic cells will be generated. We will apply cell-sorting methods to enrich pancreatic liver stem cells.