DESCRIPTION (provided by applicant): Adult stem cell niches have been identified in rapidly dividing tissues, such as the small intestine, skin and esophagus. Identification of adult stem cells in quiescent organs, such as the liver, has been more challenging. The notion that the liver contains an anatomically anchored stem cell zone near the portal tracts, giving rise to arrays of hepatocytes which progressively mature toward the central vein, is known as the "streaming liver" hypothesis. This model of liver homeostasis remains controversial, but may be operative during regeneration after severe liver injury exceeding the proliferative potential of mature hepatocytes. The goal of the work proposed here is to investigate the potential stem cell niche near the portal tracts, in the smallest branches of the bile ducts. This cell population expresses cytokeratin 19 (CK19), while mature hepatocytes do not. However, CK19 is not a unique marker for liver stem cells, as it is also expressed in mature bile ducts. Following severe liver injury, immature hepatocytes weakly expressing CK19 can be seen directly adjacent to clusters of strongly CK19-positive biliary cells, suggesting that biliary CK19+ cells can act as hepatocyte progenitors under conditions of severe liver injury. With time, the weakly CK19-positive periductal hepatocytes disappear, presumably through maturation to fully functional hepatocytes. These experiments will test the hypothesis that mature hepatocytes arise from CK19- expressing progenitors, using a state-of-the-art transgenic lineage tracing approach. CK19-expressing cells and their descendants will express yellow fluorescent protein (YFP). Aim 1: Determine the role of intrahepatic CK19+ hepatic progenitor cells during normal liver growth and homestasis. During normal development, CK19+ hepatoblasts give rise to normal bile ducts and hepatocytes. In the adult, CK19 is expressed by cholangiocytes, but not hepatocytes. I will use the tightly regulated, Tamoxifen-inducible CK19-CreERT transgenic mouse model to mark CK19-expressing cells at specific times during development, using a YFP reporter. Marking of CK19+ populations during embryonic development is expected to mark all descendants of hepatoblasts, including hepatocytes and cholangiocytes. Marking of CK19+ populations in the adult mouse is expected to mark mature bile ducts and peribiliary oval cells, but likely will not mark hepatocytes due to their inherently slow turnover and replacement by existing mature hepatocytes. Aim 2: Determine the role of intrahepatic CK19+ hepatic progenitor cells during liver regeneration after injury by 1) partial hepatectomy and 2) acute acetaminophen injury. Partial hepatectomy provides the classic example of an acute liver injury which triggers liver regeneration exclusively through proliferation of residual mature hepatocytes, rather than progenitors. In contrast, acute acetaminophen intoxication induces severe, diffuse hepatocyte necrosis and a marked increase of CK19+ oval cells in the periportal region. I predict that hepatocytes arise from CK19+ progenitors and will be lineage marked with YFP after acetaminophen injury, but not partial hepatectomy. PUBLIC HEALTH RELEVANCE: The liver has a remarkable capacity to regenerate in response to injury induced by toxins or viruses, driven in some cases by immature liver cells called "progenitor" cells. It is critical to understand the contribution of liver progenitor cells to normal and regenerating liver in order to develop treatment strategies to support regeneration and suppress tumor formation. The goal of the current proposal is to investigate the contribution of liver progenitor cells in the regenerating mouse liver, using a genetic labeling approach to permanently mark liver progenitor cells and their descendants.

NARRATIVE The liver has a remarkable capacity to regenerate in response to injury induced by toxins or viruses, driven in some cases by immature liver cells called "progenitor" cells. It is critical to understand the contribution of liver progenitor cells to normal and regenerating liver in order to develop treatment strategies to support regeneration and suppress tumor formation. The goal of the current proposal is to investigate the contribution of liver progenitor cells in the regenerating mouse liver, using a genetic labeling approach to permanently mark liver progenitor cells and their descendants.