Autophagy suppresses the formation of hepatocyte-derived cancer-initiating ductular progenitor cells in the liver.
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ABSTRACT:
SUBMITTER: Barthet VJA
PROVIDER: S-EPMC8177709 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Globally, about two million people die from liver diseases every year. Liver transplantation is the only reliable therapy for severe end-stage liver disease, however, the shortage of organ donors is a huge limitation. Human hepatocytes derived liver progenitor-like cells (HepLPCs) have been reported as a novel source of liver cells for development of in vitro models, cell therapies, and tissue-engineering applications, but their functionality as transplantation donors is unclear. Here, a 3-dimensional (3D) co-culture system using HepLPCs and human umbilical vein endothelial cells (HUVECs) was developed. These HepLPC spheroids mimicked the cellular interactions and architecture of mature hepatocytes, as confirmed through ultrastructure morphology, gene expression profile and functional assays. HepLPCs encapsulated in alginate beads are able to mitigate liver injury in mice treated with carbon tetrachloride (CCL4), while alginate coating protects the cells from immune attack. We confirmed these phenomena due to HUVECs producing glial cell line-derived neurotrophic factor (GDNF) to promote HepLPCs maturation and enhance HepLPCs tight junction through MET phosphorylation. Our results display the efficacy and safety of the alginate microencapsulated spheroids in animal model with acute liver injury (ALF), which may suggest a new strategy for cell therapy.
| S-EPMC8630579 | biostudies-literature
Project description:Rationale: The idiosyncratic drug-induced liver injury (iDILI) is a major cause of acute liver injury and a key challenge in late-stage drug development. Individual heterogeneity is considered to be an essential factor of iDILI. However, few in vitro model can predict heterogeneity in iDILI. We have previously shown that mouse and human hepatocytes can be converted to expandable liver progenitor-like cells in vitro (HepLPCs). However, the limited proliferation potential of human HepLPCs confines its industrial application. Here, we reported the generation of a novel hepatocyte model not only to provide unlimited cell sources for human hepatocytes but also to establish a tool for studying iDILI in vitro. Methods: Human primary hepatocytes were isolated by modified two-step perfusion technique. The chemical reprogramming culture condition together with gene-transfer were then used to generate the immortalized HepLPC cell lines (iHepLPCs). Growth curve, doubling time, and karyotype were analyzed to evaluate the proliferation characteristics of iHepLPCs. Modified Hepatocyte Maturation Medium and 3D spheroid culture were applied to re-differentiate iHepLPCs. Results: iHepLPCs exhibited efficient expansion for at least 40 population doublings, with a stable proliferative ability. They could easily differentiate back into metabolically functional hepatocytes in vitro within 10 days. Furthermore, under three-dimensional culture conditions, the formed hepatic spheroids showed multiple liver functions and toxicity profiles close to those of primary human hepatocytes. Importantly, we established a hepatocyte bank by generating a specific number of such cell lines. Screening for population heterogeneity allowed us to analyze the in vitro heterogeneous responses to hepatotoxicity induced by molecular targeted drugs. Conclusions: In light of the proliferative capacity and the heterogeneity they represented, these iHepLPCs cell lines may offer assistance in studying xenobiotic metabolism as well as liver diseases in vitro.
| S-EPMC6771233 | biostudies-literature
Project description:ObjectiveThe ductular reaction (DR) involves mobilisation of reactive-appearing duct-like cells (RDC) along canals of Hering, and myofibroblastic (MF) differentiation of hepatic stellate cells (HSC) in the space of Disse. Perivascular cells in stem cell niches produce pleiotrophin (PTN) to inactivate the PTN receptor, protein tyrosine phosphatase receptor zeta-1 (PTPRZ1), thereby augmenting phosphoprotein-dependent signalling. We hypothesised that the DR is regulated by PTN/PTPRZ1 signalling.DesignPTN-GFP, PTN-knockout (KO), PTPRZ1-KO, and wild type (WT) mice were examined before and after bile duct ligation (BDL) for PTN, PTPRZ1 and the DR. RDC and HSC from WT, PTN-KO, and PTPRZ1-KO mice were also treated with PTN to determine effects on downstream signaling phosphoproteins, gene expression, growth, and migration. Liver biopsies from patients with DRs were also interrogated.ResultsAlthough quiescent HSC and RDC lines expressed PTN and PTPRZ1 mRNAs, neither PTN nor PTPRZ1 protein was demonstrated in healthy liver. BDL induced PTN in MF-HSC and increased PTPRZ1 in MF-HSC and RDC. In WT mice, BDL triggered a DR characterised by periportal accumulation of collagen, RDC and MF-HSC. All aspects of this DR were increased in PTN-KO mice and suppressed in PTPRZ1-KO mice. In vitro studies revealed PTN-dependent accumulation of phosphoproteins that control cell-cell adhesion and migration, with resultant inhibition of cell migration. PTPRZ1-positive cells were prominent in the DRs of patients with ductal plate defects and adult cholestatic diseases.ConclusionsPTN, and its receptor, PTPRZ1, regulate the DR to liver injury by controlling the migration of resident cells in adult liver progenitor niches.
| S-EPMC4504836 | biostudies-literature
Project description:Differentiated epithelial cells show substantial lineage plasticity upon severe tissue injuries. In chronically injured mouse livers, part of hepatocytes become Sry-HMG box containing 9 (Sox9) (+) epithelial cell adhesion molecule (-) hepatocyte nuclear factor 4 α (+) biphenotypic hepatocytes. However, it is not clear whether all Sox9+ hepatocytes uniformly possess cellular properties as hepatocyte progenitors. Here, we examined the microarray data comparing Sox9+ hepatocytes with mature hepatocytes and identified CD24 as a novel marker for biphenotypic hepatocytes. Immunohistochemical analyses showed that part of Sox9+ hepatocytes near expanded ductular structures expressed CD24 in the liver injured by 3,5-diethoxycarbonyl-1,4-dihydro-collidine (DDC) diet and by bile duct ligation. Indeed, Sox9+ hepatocytes could be separated into CD24- and CD24+ cells by fluorescence activated cell sorting. The ratio of CD24+ cells against CD24- ones in Sox9+ hepatocytes gradually increased while DDC-injury progressed and colony-forming capability mostly attributed to CD24+ cells. Although hepatocyte markers were remarkably downregulated in of Sox9+ CD24+ hepatocytes, they re-differentiated into mature hepatocytes in vitro and in vivo. Our current results demonstrate that the emergence of biphenotypic hepatocytes is a sequential event including the transition from CD24- and CD24+ status, which may be a crucial step for hepatocytes to acquire progenitor properties.
| S-EPMC5209740 | biostudies-literature
Project description:The identification of specific cell surface markers that can be used to isolate liver progenitor cells will greatly facilitate experimentation to determine the role of these cells in liver regeneration and their potential for therapeutic transplantation. Previously, the cell surface marker, CD24, was observed to be expressed on undifferentiated bipotential mouse embryonic liver stem cells and 3,5-diethoxycarbonyl-1,4-dihydrocollidine-induced oval cells. Here, we describe the isolation and characterization of a rare, primary, nonhematopoietic, CD24+ progenitor cell population from normal, untreated mouse liver. By immunohistochemistry, CD24-expressing cells in normal adult mouse liver were colocalized with CK19-positive cholangiocytes. This nonhematopoietic (CD45-, Ter119-) CD24+ cell population isolated by flow cytometry represented 0.04% of liver cells and expressed several markers of liver progenitor/oval cells. The immunophenotype of nonhematopoietic CD24+ cells was CD133, Dlk, and Sca-1 high, but c-Kit, Thy-1, and CD34 low. The CD24+ cells had increased expression of CK19, epithelial cell adhesion molecule, Sox 9, and FN14 compared with the unsorted cells. Upon transplantation of nonhematopoietic CD24+ cells under the sub-capsule of the livers of Fah knockout mice, cells differentiated into mature functional hepatocytes. Analysis of X and Y chromosome complements were used to determine whether or not fusion of the engrafted cells with the recipient hepatocytes occurred. No cells were found that contained XXXY or any other combination of donor and host sex chromosomes as would be expected if cell fusion had occurred. These results suggested that CD24 can be used as a cell surface marker for isolation of hepatocyte progenitor cells from normal adult liver that are able to differentiate into hepatocytes.
| S-EPMC3225069 | biostudies-literature
Project description:Our previous studies have shown that autophagy mediates the link between ductular reaction (DR) and liver cirrhosis. Whether the subsequent fibrogenic response is regulated by increased autophagy in DR remains unclear. Here, using both human liver specimens and a rat model of liver cirrhosis induced by 2-acetylaminofluorene (AAF) and carbon tetrachloride (CCL4), we explored the involvement of autophagy in regulating mesenchymal transition of ductular cells. Ductular cells from AAF/CCL4 livers exhibited increased autophagy compared to those of normal livers. These cells showed morphological and functional characteristics of mesenchymal cells. Blocking autophagy using bafilomycin A1 or siRNA targeting ATG7 reduced the expression of mesenchymal markers in these ductular cells from AAF/CCL4 livers, indicating a role for autophagy in regulating the mesenchymal phenotype of ductular cells. Furthermore, we show that the mesenchymal transition in DR requires the activation of transforming growth factor-? (TGF-?) signaling in an autophagy-dependent manner. Importantly, in cirrhotic human livers, ductular cells that are positive for LC3B also showed increased expression of TGF-? and fibroblast-specific protein-1. Our data suggest activation of autophagy in ductular cells, and also demonstrate that it is required for the mesenchymal transition during the DR, processes that are critically involved in the pathogenesis of cirrhosis.
| S-EPMC6650611 | biostudies-literature
Project description:BackgroundCD24+CK19+/CD24+SOX9+ resident liver cells are activated and expanded after chronic liver injury in a ductular reaction. However, the sources and functions of these cells in liver damage remain disputed.ResultsThe current study combined genetic lineage tracing with in vitro small-molecule-based reprogramming to define liver progenitor cells (LPCs) derived from hepatic parenchymal and non-parenchymal tissues. tdTom+ hepatocytes were isolated from ROSA26tdTomato mice following AAV8-Tbg-Cre-mediated recombination, EpCAM+ biliary epithelial cells (BECs) from wild-type intrahepatic bile ducts and ALB/GFP-EpCAM- cells were isolated from AlbCreERT/R26GFP mice. A cocktail of small molecules was used to convert the isolated cells into LPCs. These in vitro cultured LPCs with CD24 and SOX9 expression regained the ability to proliferate. Transcriptional profiling showed that the in-vitro cultured LPCs derived from the resident LPCs in non-parenchymal tissues expressed Lipocalin-2 (Lcn2) at high levels. Accordingly, endogenous Cd24a+Lcn2+ LPCs were identified by integration of sc-RNA-sequencing and pathological datasets of liver dysfunction which indicates that LPCs produced by ductular reactions might also originate from the resident LPCs. Transplantation of in-vitro cultured Cd24a+Lcn2+ LPCs into CCl4-induced fibrotic livers exacerbated liver damage and dysfunction, possibly due to LCN2-dependent macrophage inflammatory response.ConclusionsCD24+LCN2+ LPCs constituted the expanding ductular reaction and contributed to macrophage-mediated inflammation in chronic liver damage. The current findings highlight the roles of LPCs from distinct origins and expose the possibility of targeting LPCs in the treatment of chronic hepatic diseases.
| S-EPMC10546777 | biostudies-literature
Project description:Hepatocellular carcinoma (HCC) generally arises in chronically inflamed liver and is thought to originate from regenerating liver cells that acquired genetic alterations. Both hepatocytes and ductular progenitor cells (DPCs) contribute to liver regeneration and are candidate cellular origins of HCCs. In the current study, we used lineage-tracing analysis to show that Epcam-expressing DPCs give rise to HCCs in inflamed liver. Under conditions in which proliferating DPCs differentiate into cholangiocytes but not into hepatocytes, the accumulation of genetic alterations in Epcam-expressing DPCs resulted in the development of HCCs with histologic features of concomitant ductule-like structures resembling human cholangiolocellular carcinoma (CLC). To elucidate the characteristics of DPC-derived HCC, gene expression of HCCs derived from Epcam-expressing DPCs were evaluated using Agilent SurePrint G3 Mouse Gene Expression 8x60K v2. Two DPC-derived HCCs with CLC components, two DPC-derived HCCs without CLC, and two HCCs derived from albumin-expressing cells were analyzed.
2020-02-19 | GSE95099 | GEO
Project description:The study of pathophysiological mechanisms in human liver disease has been constrained by the inability to expand primary hepatocytes in vitro while maintaining proliferative capacity and metabolic function. We and others have previously shown that mouse mature hepatocytes can be converted to liver progenitor-like cells in vitro with defined chemical factors. Here we describe a protocol achieving efficient conversion of human primary hepatocytes into liver progenitor-like cells (HepLPCs) through delivery of developmentally relevant cues, including NAD + -dependent deacetylase SIRT1 signaling. These HepLPCs could be expanded significantly during in vitro passage. The expanded cells can readily be converted back into metabolically functional hepatocytes in vitro and upon transplantation in vivo. Under three-dimensional culture conditions, differentiated cells generated from HepLPCs regained the ability to support infection or reactivation of hepatitis B virus (HBV). Our work demonstrates the utility of the conversion between hepatocyte and liver progenitor-like cells for studying HBV biology and antiviral therapies. These findings will facilitate the study of liver diseases and regenerative medicine.
| S-EPMC6318298 | biostudies-literature
Project description:Autophagy is a vital process that involves degradation of long-lived proteins and dysfunctional organelles and contributes to cellular metabolism. Glioma-initiating cells (GICs) have the ability to self-renew, differentiate into heterogeneous types of tumor cells, and sustain tumorigenicity; thus, GICs lead to tumor recurrence. Accumulating evidence indicates that autophagy can induce stem cell differentiation and increase the lethality of temozolomide against GICs. However, the mechanism underlying the regulation of GIC self-renewal by autophagy remains uncharacterized. In the present study, autophagy induced by AZD8055 and rapamycin treatment suppressed GIC self-renewal in vitro. We found that autophagy inhibited Notch1 pathway activation. Moreover, autophagy activated Notch1 degradation, which is associated with maintenance of the self-renewal ability of GICs. Furthermore, autophagy abolished the tumorigenicity of CD133 + U87-MG neurosphere cells in an intracranial model. These findings suggest that autophagy regulating GICs self-renewal and tumorigenicity is probably bound up with Notch1 degradation. The results of this study could aid in the design of autophagy-based clinical trials for glioma treatments, which may be of great value.
| S-EPMC6194143 | biostudies-literature