The Polyploid State Plays a Tumor-Suppressive Role in the Liver.
2018; 44 (4): 447?59.e5
Most cells in the liver are polyploid, but the functional role of polyploidy is unknown. Polyploidization occurs through cytokinesis failure and endoreduplication around the time of weaning. To interrogate polyploidy while avoiding irreversible manipulations of essential cell-cycle genes, we developed orthogonal mouse models to transiently and potently alter liver ploidy. Premature weaning, as well as knockdown of E2f8 or Anln, allowed us to toggle between diploid and polyploid states. While there was no detectable impact of ploidy alterations on liver function, metabolism, or regeneration, mice with more polyploid hepatocytes suppressed tumorigenesis and mice with more diploid hepatocytes accelerated tumorigenesis in mutagen- and high-fat-induced models. Mechanistically, the diploid state was more susceptible to Cas9-mediated tumor-suppressor loss but was similarly susceptible to MYC oncogene activation, indicating that polyploidy differentially protected the liver from distinct genomic aberrations. This suggests that polyploidy evolved in part to prevent malignant outcomes of liver injury.
View details for PubMedID 29429824
Bipotential Adult Liver Progenitors Are Derived from Chronically Injured Mature Hepatocytes
CELL STEM CELL
2014; 15 (5): 605-618
Adult liver progenitor cells are biliary-like epithelial cells that emerge only under injury conditions in the periportal region of the liver. They exhibit phenotypes of both hepatocytes and bile ducts. However, their origin and their significance to injury repair remain unclear. Here, we used a chimeric lineage tracing system to demonstrate that hepatocytes contribute to the progenitor pool. RNA-sequencing, ultrastructural analysis, and in vitro progenitor assays revealed that hepatocyte-derived progenitors were distinct from their biliary-derived counterparts. In vivo lineage tracing and serial transplantation assays showed that hepatocyte-derived proliferative ducts retained a memory of their origin and differentiated back into hepatocytes upon cessation of injury. Similarly, human hepatocytes in chimeric mice also gave rise to biliary progenitors in vivo. We conclude that human and mouse hepatocytes can undergo reversible ductal metaplasia in response to injury, expand as ducts, and subsequently contribute to restoration of the hepatocyte mass.
View details for DOI 10.1016/j.stem.2014.09.008
View details for Web of Science ID 000345012700013
View details for PubMedID 25312494
View details for PubMedCentralID PMC4254170
Human islets contain four distinct subtypes of beta cells
2016; 7: 11756
Human pancreatic islets of Langerhans contain five distinct endocrine cell types, each producing a characteristic hormone. The dysfunction or loss of the insulin-producing ? cells causes diabetes mellitus, a disease that harms millions. Until now, ? cells were generally regarded as a single, homogenous cell population. Here we identify four antigenically distinct subtypes of human ? cells, which we refer to as ?1-4, and which are distinguished by differential expression of ST8SIA1 and CD9. These subpopulations are always present in normal adult islets and have diverse gene expression profiles and distinct basal and glucose-stimulated insulin secretion. Importantly, the ? cell subtype distribution is profoundly altered in type 2 diabetes. These data suggest that this antigenically defined ? cell heterogeneity is functionally and likely medically relevant.
View details for PubMedID 27399229
Fibroblast Growth Factor Signaling Controls Liver Size in Mice With Humanized Livers
2015; 149 (3): 728-?
The ratio of liver size to body weight (hepatostat) is tightly controlled, but little is known about how the physiologic functions of the liver help determine its size. Livers of mice repopulated with human hepatocytes (humanized livers) grow to larger than normal; the human hepatocytes do not recognize the fibroblast growth factor (FGF)-15 produced by mouse intestine. This results in up-regulation of bile acid synthesis in the human hepatocytes and enlargement of the bile acid pool. We investigated whether abnormal bile acid signaling affects the hepatostat in mice.We crossed Fah(-/-), Rag2(-/-), Il2r(-/-) mice with nonobese diabetic mice to create FRGN mice, whose livers can be fully repopulated with human hepatocytes. We inserted the gene for human FGF19 (ortholog to mouse Fgf15), including regulatory sequences, into the FRGN mice to create FRGN19(+) mice. Livers of FRGN19(+) mice and their FRGN littermates were fully repopulated with human hepatocytes. Liver tissues were collected and bile acid pool sizes and RNA sequences were analyzed and compared with those of mice without humanized livers (controls).Livers were larger in FRGN mice with humanized livers (13% of body weight), compared with control FRGN mice; they also had much larger bile acid pools and aberrant bile acid signaling. Livers from FRGN19(+) normalized to 7.8% of body weight, and their bile acid pool and signaling more closely resembled that of control FRGN19(+) mice. RNA sequence analysis showed activation of the Hippo pathway, and immunohistochemical and transcription analyses revealed increased hepatocyte proliferation, but not apoptosis, in the enlarged humanized livers of FRGN mice. Cell sorting experiments showed that although healthy human liver does not produce FGF19, nonparenchymal cells from cholestatic livers produce FGF19.In mice with humanized livers, expression of an FGF19 transgene corrects bile acid signaling defects, resulting in normalization of bile acid synthesis, the bile acid pool, and liver size. These findings indicate that liver size is, in part, regulated by the size of the bile acid pool that the liver must circulate.
View details for DOI 10.1053/j.gastro.2015.05.043
View details for Web of Science ID 000360269800041
View details for PubMedID 26028580
View details for PubMedCentralID PMC4550566
Extensive double humanization of both liver and hematopoiesis in FRGN mice
STEM CELL RESEARCH
2014; 13 (3): 404?12
Preclinical research in animals often fails to adequately predict the outcomes observed in human patients. Chimeric animals bearing individual human tissues have been developed to provide improved models of human-specific cellular processes. Mice transplanted with human hematopoietic stem cells can be used to study human immune responses, infections of blood cells and processes of hematopoiesis. Animals with humanized livers are useful for modeling hepatotropic infections as well as drug metabolism and hepatotoxicity. However, many pathophysiologic processes involve both the liver and the hematolymphoid system. Examples include hepatitis C/HIV co-infection, immune mediated liver diseases, liver injuries with inflammation such as steatohepatitis and alcoholic liver disease. We developed a robust protocol enabling the concurrent double-humanization of mice with mature hepatocytes and human blood. Immune-deficient, fumarylacetoacetate hydrolase (Fah(-/-)), Rag2(-/-) and Il2rg(-/-) deficient animals on the NOD-strain background (FRGN) were simultaneously co-transplanted with adult human hepatocytes and hematopoietic stem cells after busulfan and Ad:uPA pre-conditioning. Four months after transplantation the average human liver repopulation exceeded 80% and hematopoietic chimerism also was high (40-80% in bone marrow). Importantly, human macrophages (Kupffer cells) were present in the chimeric livers. Double-chimeric FRGN mice will serve as a new model for disease processes that involve interactions between hepatocytes and hematolymphoid cells.
View details for PubMedID 25310256
The organoid-initiating cells in mouse pancreas and liver are phenotypically and functionally similar
STEM CELL RESEARCH
2014; 13 (2): 275?83
Pancreatic Lgr5 expression has been associated with organoid-forming epithelial progenitor populations but the identity of the organoid-initiating epithelial cell subpopulation has remained elusive. Injury causes the emergence of an Lgr5(+) organoid-forming epithelial progenitor population in the adult mouse liver and pancreas. Here, we define the origin of organoid-initiating cells from mouse pancreas and liver prior to Lgr5 activation. This clonogenic population was defined as MIC1-1C3(+)/CD133(+)/CD26(-) in both tissues and the frequency of organoid initiation within this population was approximately 5% in each case. The transcriptomes of these populations overlapped extensively and showed enrichment of epithelial progenitor-associated regulatory genes such as Sox9 and FoxJ1. Surprisingly, pancreatic organoid cells also had the capacity to generate hepatocyte-like cells upon transplantation to Fah(-/-) mice, indicating a differentiation capacity similar to hepatic organoids. Although spontaneous endocrine differentiation of pancreatic progenitors was not observed in culture, adenoviral delivery of fate-specifying factors Pdx1, Neurog3 and MafA induced insulin expression without glucagon or somatostatin. Pancreatic organoid cultures therefore preserve many key attributes of progenitor cells while allowing unlimited expansion, facilitating the study of fate determination.
View details for PubMedID 25151611
View details for PubMedCentralID PMC4243830
Clonal Tracing of Sox9(+) Liver Progenitors in Mouse Oval Cell Injury
2014; 60 (1): 278-289
Proliferating ducts, termed "oval cells," have long been thought to be bipotential, that is, produce both biliary ducts and hepatocytes during chronic liver injury. The precursor to oval cells is considered to be a facultative liver stem cell (LSC). Recent lineage tracing experiments indicated that the LSC is SRY-related HMG box transcription factor 9 positive (Sox9(+) ) and can replace the bulk of hepatocyte mass in several settings. However, no clonal relationship between Sox9(+) cells and the two epithelial liver lineages was established. We labeled Sox9(+) mouse liver cells at low density with a multicolor fluorescent confetti reporter. Organoid formation validated the progenitor activity of the labeled population. Sox9(+) cells were traced in multiple oval cell injury models using both histology and fluorescence-activated cell sorting. Surprisingly, only rare clones containing both hepatocytes and oval cells were found in any experiment. Quantitative analysis showed that Sox9(+) cells contributed only minimally (<1%) to the hepatocyte pool, even in classic oval cell injury models. In contrast, clonally marked mature hepatocytes demonstrated the ability to self-renew in all classic mouse oval cell activation injuries. A hepatocyte chimera model to trace hepatocytes and nonparenchymal cells also demonstrated the prevalence of hepatocyte-driven regeneration in mouse oval cell injury models.Sox9(+) ductal progenitor cells give rise to clonal oval cell proliferation and bipotential organoids, but rarely produce hepatocytes in vivo. Hepatocytes themselves are the predominant source of new parenchyma cells in prototypical mouse models of oval cell activation.
View details for DOI 10.1002/hep.27084
View details for Web of Science ID 000337969000032
View details for PubMedID 24700457
View details for PubMedCentralID PMC4077948