Identification and expansion of a unique stem cell population from adult mouse gallbladder

Understanding the resident stem cell populations of the biliary system has great importance for basic biology and biliary diseases. The biliary tree is divided into the intrahepatic and extrahepatic biliary systems. The latter consists of the gallbladder, cystic duct, and the common bile duct (1). The biliary system is a conduit for bile to be transported from the liver to the intestine. The gallbladder in turn stores the bile and regulates its content and concentration, playing an important role in the digestive process (2, 3). While there has been a lot of recent interest in the liver stem cell field (4) there is still a paucity of data regarding gallbladder stem cells. The biliary system, hepatocytes and ventral pancreas develop from the ventral foregut endoderm (5, 6). Histological evidence suggesting that both intra- and extrahepatic systems originate from the hepatic diverticulum has led to the hypothesis that they descend from the same progenitor cell. However, the cell-intrinsic factors that result in their specification have heretofore been unclear. Recently, it has been shown that the progenitor cells that give rise to each system separate out during development (7). Using a Pdx1-Cre mouse Spence et al. (7) demonstrated that the hepatocytes and intrahepatic bile duct (IHBD) cells derive from Pdx1- cells while the extrahepatic bile duct (EHBD) cells and ventral pancreas derive from Pdx1+ cells. Sox17 controls the specification of the EHBD and pancreatic cells. Sox17 loss-of-function embryos exhibit gallbladder agenesis and the presence of ectopic pancreatic tissue in the extrahepatic bile duct. Conversely Sox17 gain-of-function results in ectopic ductal tissue in the developing pancreas. In both cases, the intrahepatic system is not affected. It appears that the IHBD and EHBD cells descend from separate progenitor cells governed by separate transcriptional cascades. It is therefore possible that adult IHBD and EHBD cells could be distinct as well. The aims of this study were to isolate and characterize stem cells from the adult mouse gallbladder and compare their phenotypic and expression profiles with IHBD cells. In addition to basic biology, an understanding of gallbladder stem cells would be vital to the study of gallbladder carcinoma, a rare but poorly understand malignancy (8) and congenital diseases involving biliary dysmorphogenesis, such as biliary atresia (9). It would also elucidate the ontogeny of cells in the biliary system. Stem cells are defined as undifferentiated cells that can self-renew at the single cell level and form lineage committed progeny (10). In this study, we use colony forming and single cell assays along with a morphogenesis assay to characterize an EpCAM+CD49fhi epithelial sub-population within primary mouse gallbladder that has stem cell characteristics. The gallbladder stem cells can be propagated in vitro through long-term passage (>passage 20) and can engraft in the subcutaneous space of recipient mice. Last the gallbladder stem cells and IHBD cells have distinct expression profiles. These data represent one of the first reports to isolate and characterize the resident stem cell population in the adult mouse gallbladder.