Hepatic stellate cell-derived delta-like homolog 1 (DLK1) protein in liver regeneration.
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ABSTRACT:
SUBMITTER: Zhu NL
PROVIDER: S-EPMC3322997 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:BACKGROUND:As a critical cellular component in the hepatic stem cell niche, hepatic stellate cells (HSCs) play critical roles in regulating the expansion of hepatic stem cells, liver regeneration, and fibrogenesis. However, the signaling of HSCs, particularly that involved in promoting hepatic stem cell expansion, remains unclear. While the overexpression of galectins has been identified in regenerating liver tissues, their involvement in cell-cell interactions between HSCs and hepatic stem cells remains to be elucidated. METHODS:To generate a liver regeneration rat model and establish a hepatic oval cell microenvironment as a stem cell niche, 2-acetylaminofluorene treatment plus partial hepatectomy was performed. Immunofluorescence staining was conducted to detect the emergence of hepatic stem cells and their niche. Liver parenchymal cells, non-parenchymal cells, and HSCs were isolated for gene and protein expression analysis by qPCR or western blotting. To evaluate the effect of galectins on the colony-forming efficiency of hepatic stem cells, c-Kit-CD29+CD49f+/lowCD45-Ter-119- cells were cultured with recombinant galectin protein, galectin antibody, galectin-producing HSCs, and galectin-knockdown HSCs. RESULTS:Following liver injury, the cytokeratin 19+ ductal cells were robustly induced together with the emergence of OV6+CD44+CD133+EpCAM+ hepatic stem cells. The activated desmin+ HSCs were recruited around the periportal area and markedly enriched in the galectin-positive domain compared to the other non-parenchymal cells. Notably, the HSC fraction isolated from regenerating liver was accompanied by dramatically elevated gene and protein expression of galectins. Hepatic stem cells co-cultured with HSCs significantly enhanced colony-forming efficiency. Conversely, single or double knockdown of galectin-1 and galectin-3 led into a significant function loss, impaired the co-cultured hepatic stem cells to attenuated colony size, inhibited colony frequency, and reduced total cell numbers in colonies. On the other hand, the promotive function of galectins was further confirmed by recombinant galectin protein supplementation and galectins blocking antibodies. CONCLUSIONS:Our findings, for the first time, demonstrated that galectins from activated HSCs contribute to hepatic stem cell expansion during liver regeneration, suggesting that galectins serve as important stem cell niche components.
| S-EPMC7526193 | biostudies-literature
Project description:The stem cell-like characteristics of tumor cells are not only essential for tumor development and malignant progression, but also significantly contribute to therapy resistance. However, it remains poorly understood how cancer cell differentiation or stemness is regulated in vivo. We investigated the role of the stem cell gene DLK1, or delta-like 1 homolog (Drosophila), in the regulation of cancer cell differentiation in vivo using neuroblastoma (NB) xenografts as a model. We found that loss-of-function mutants of DLK1 significantly enhanced NB cell differentiation in vivo likely by increasing the basal phosphorylation of MEK and ERK kinases, a mechanism that has been shown to facilitate neuronal differentiation. We also found that DLK1(+) cells are preferentially located in hypoxic regions. These results clearly demonstrate that DLK1 plays an important role in the maintenance of undifferentiated, stem cell-like phenotypes of NB cells in vivo.
| S-EPMC3243111 | biostudies-literature
Project description:Retinoid-storing hepatic stellate cells (HSCs) have recently been described as a liver-resident mesenchymal stem cell (MSC) population; however, it is not clear whether these cells contribute to liver regeneration or serve as a progenitor cell population with hepatobiliary characteristics. Here, we purified HSCs with retinoid-dependent fluorescence-activated cell sorting from eGFP-expressing rats and transplanted these GFP(+) HSCs into wild-type (WT) rats that had undergone partial hepatectomy in the presence of 2-acetylaminofluorene (2AAF) or retrorsine, both of which are injury models that favor stem cell-based liver repair. Transplanted HSCs contributed to liver regeneration in host animals by forming mesenchymal tissue, progenitor cells, hepatocytes, and cholangiocytes and elevated direct bilirubin levels in blood sera of GUNN rats, indicating recovery from the hepatic bilirubin-handling defect in these animals. Transplanted HSCs engrafted within the bone marrow (BM) of host animals, and HSC-derived cells were isolated from BM and successfully retransplanted into new hosts with injured liver. Cultured HSCs transiently adopted an expression profile similar to that of progenitor cells during differentiation into bile acid-synthesizing and -transporting hepatocytes, suggesting that stellate cells represent a source of liver progenitor cells. This concept connects seemingly contradictory studies that favor either progenitor cells or MSCs as important players in stem cell-based liver regeneration.
| S-EPMC4348953 | biostudies-literature
Project description:Progression of fibrosis and the development of cirrhosis are responsible for the liver related morbidity and mortality associated with chronic liver diseases. There is currently a great unmet need for effective anti-fibrotic strategies. Stem cells play a central role in wound healing responses to restore liver homeostasis following injury. Here we tested the hypothesis that extracellular vesicles (EVs) isolated from induced pluripotent stem cells (iPSC) modulate hepatic stellate cell (HSCs) activation and may have anti-fibrotic effects. Human iPSCs were generated by reprogramming primary skin fibroblasts. EVs were isolated by differential centrifugation, quantified by flow cytometry (FACS) and characterized by dynamic light scattering (DLS) and electron microscopy (TEM). Primary human HSCs were activated with TGFβ (10 ng/mL) and exposed to iPSC-EVs. Efficacy of iPSC-EVs was tested on HSC in vitro and in two murine models of liver injury (CCl4 and bile duct ligation). Characterization of iPSC-derived EVs by flow cytometry identified a large population of EVs released by iPSC, primarily with a diameter of 300 nm and that could be visualized by TEM as round, cup-shaped objects. Fluorescent tracing assays detected iPSC-EVs in HSC cytosol after a short incubation and EV uptake by HSCs resulted in both decrease of pro-fibrogenic markers αSMA, CollagenIα1, Fibronectin and TIMP-1 and HSC pro-fibrogenic responses such as chemotaxis and proliferation. Genomics analyses of iPSC-EV miRNA cargo revealed 22 highly expressed miRNAs, among which miR-92a-3p resulted the most abundant. Transcriptome analysis identified 60 genes down-modulated and 235 up-regulated in TGF-β-primed HSC in presence or absence of iPSC-EVs. Intravenous injection of iPSC-EVs in CCl4 and bile duct ligation-induced liver fibrosis resulted in anti-fibrotic effects at protein and gene levels. Results of this study identify iPSC-EVs as a novel anti-fibrotic approach that may reduce or reverse liver fibrosis in patients with chronic liver disease.
| S-EPMC6675559 | biostudies-literature
Project description:Several studies indicate that adult stem cells may improve the recovery from acute tissue injury. It has been suggested that they may contribute to tissue regeneration by the release of paracrine factors promoting proliferation of tissue resident cells. However, the factors involved remain unknown. In the present study we found that microvesicles (MVs) derived from human liver stem cells (HLSC) induced in vitro proliferation and apoptosis resistance of human and rat hepatocytes. These effects required internalization of MVs in the hepatocytes by an alpha(4)-integrin-dependent mechanism. However, MVs pre-treated with RNase, even if internalized, were unable to induce hepatocyte proliferation and apoptosis resistance, suggesting an RNA-dependent effect. Microarray analysis and quantitative RT-PCR demonstrated that MVs were shuttling a specific subset of cellular mRNA, such as mRNA associated in the control of transcription, translation, proliferation and apoptosis. When administered in vivo, MVs accelerated the morphological and functional recovery of liver in a model of 70% hepatectomy in rats. This effect was associated with increase in hepatocyte proliferation and was abolished by RNase pre-treatment of MVs. Using human AGO2, as a reporter gene present in MVs, we found the expression of human AGO2 mRNA and protein in the liver of hepatectomized rats treated with MVs. These data suggested a translation of the MV shuttled mRNA into hepatocytes of treated rats. In conclusion, these results suggest that MVs derived from HLSC may activate a proliferative program in remnant hepatocytes after hepatectomy by a horizontal transfer of specific mRNA subsets.
| S-EPMC3060338 | biostudies-literature
Project description:BACKGROUND:Recent results from single cell gene and protein regulation studies are starting to uncover the previously underappreciated fact that individual cells within a population exhibit high variability in the expression of mRNA and proteins (i.e., molecular variability). By combining cellular network modeling, and high-throughput gene expression measurements in single cells, we seek to reconcile the high molecular variability in single cells with the relatively low variability in tissue-scale gene and protein expression and the highly coordinated functional responses of tissues to physiological challenges. In this study, we focus on relating the dynamic changes in distributions of hepatic stellate cell (HSC) functional phenotypes to the tightly regulated physiological response of liver regeneration. RESULTS:We develop a mathematical model describing contributions of HSC functional phenotype populations to liver regeneration and test model predictions through isolation and transcriptional characterization of single HSCs. We identify and characterize four HSC transcriptional states contributing to liver regeneration, two of which are described for the first time in this work. We show that HSC state populations change in vivo in response to acute challenges (in this case, 70% partial hepatectomy) and chronic challenges (chronic ethanol consumption). Our results indicate that HSCs influence the dynamics of liver regeneration through steady-state tissue preconditioning prior to an acute insult and through dynamic control of cell state balances. Furthermore, our modeling approach provides a framework to understand how balances among cell states influence tissue dynamics. CONCLUSIONS:Taken together, our combined modeling and experimental studies reveal novel HSC transcriptional states and indicate that baseline differences in HSC phenotypes as well as a dynamic balance of transitions between these phenotypes control liver regeneration responses.
| S-EPMC6171157 | biostudies-literature
Project description:Senescent cells have long been associated with deleterious effects in aging-related pathologies, although recent studies have uncovered their beneficial roles in certain contexts, such as wound healing. We have found that hepatic stellate cells (HSCs) underwent senescence within 2 days after 2/3 partial hepatectomy (PHx) in young (2-3 months old) mice, and the elimination of these senescent cells by using the senolytic drug ABT263 or by using a genetic mouse model impaired liver regeneration. Senescent HSCs secrete IL-6 and CXCR2 ligands as part of the senescence-associated secretory phenotype, which induces multiple signaling pathways to stimulate liver regeneration. IL-6 activates STAT3, induces Yes-associated protein (YAP) activation through SRC family kinases, and synergizes with CXCL2 to activate ERK1/2 to stimulate hepatocyte proliferation. The administration of either IL-6 or CXCL2 partially restored liver regeneration in mice with senescent cell elimination, and the combination of both fully restored liver weight recovery. Furthermore, the matricellular protein central communication network factor 1 (CCN1, previously called CYR61) was rapidly elevated in response to PHx and induced HSC senescence. Knockin mice expressing a mutant CCN1 unable to bind integrin α6β1 were deficient in senescent cells and liver regeneration after PHx. Thus, HSC senescence, largely induced by CCN1, is a programmed response to PHx and plays a critical role in liver regeneration through signaling pathways activated by IL-6 and ligands of CXCR2.
| S-EPMC9431681 | biostudies-literature
Project description:Activation of hepatic stellate cells (HSCs) represents the primary driving force to promote the progression of chronic cholestatic liver diseases. We previously reported that cholangiocyte-derived exosomal long noncoding RNA-H19 (lncRNA-H19) plays a critical role in promoting cholestatic liver injury. However, it remains unclear whether cholangiocyte-derived lncRNA-H19 regulates HSC activation, which is the major focus of this study. Both bile duct ligation (BDL) and Mdr2 knockout (Mdr2-/- ) mouse models were used. Wild-type and H19maternalΔExon1/+ (H19KO) mice were subjected to BDL. Mdr2-/- H19maternalΔExon1/+ (DKO) mice were generated. Exosomes isolated from cultured mouse and human cholangiocytes or mouse serum were used for in vivo transplantation and in vitro studies. Fluorescence-labeled exosomes and flow cytometry were used to monitor exosome uptake by hepatic cells. Collagen gel contraction and bromodeoxyuridine assays were used to determine the effect of exosomal-H19 on HSC activation and proliferation. Mouse and human primary sclerosing cholangitis (PSC)/primary biliary cholangitis (PBC) liver samples were analyzed by real-time PCR, western blot analysis, histology, and immunohistochemistry. The results demonstrated that hepatic H19 level was closely correlated with the severity of liver fibrosis in both mouse models and human patients with PSC and PBC. H19 deficiency significantly protected mice from liver fibrosis in BDL and Mdr2-/- mice. Transplanted cholangiocyte-derived H19-enriched exosomes were rapidly and preferentially taken up by HSCs and HSC-derived fibroblasts, and promoted liver fibrosis in BDL-H19KO mice and DKO mice. H19-enriched exosomes enhanced transdifferentiation of cultured mouse primary HSCs and promoted proliferation and matrix formation in HSC-derived fibroblasts. Conclusion: Cholangiocyte-derived exosomal H19 plays a critical role in the progression of cholestatic liver fibrosis by promoting HSC differentiation and activation and represents a potential diagnostic biomarker and therapeutic target for cholangiopathies.
| S-EPMC6783323 | biostudies-literature
Project description:BackgroundHepatic stellate cell (HSC) differentiation/activation is central to liver fibrosis and is innately linked to the immune response to liver injury. Exosomes (EXOs) are important means of communication between cell populations. This study sought to characterize EXO release from HSCs and the effect of HSC-EXOs on macrophage cytokine release/function.MethodsLiver from a rat fibrosis model was analyzed for EXO expression and localization. Quiescent and culture-activated rat and mouse HSCs and activated human HSCs were analyzed for microRNA expression. Mouse, rat, and human HSCs were culture-activated and EXOs purified from culture medium prior to addition to macrophages, and interleukin-6 (IL-6) and tumor necrosis factor-α (TNFα) mRNA and protein measured. The effect of activated HSC-EXOs on macrophage migration was assayed.ResultsActivation of rat HSCs led to increased EXO production in vivo, an effect mirrored by in vitro rat HSC culture-activation. Culture activation of mouse and rat HSCs led to altered EXO microRNA profiles, with a similar microRNA profile detected in activated human HSCs. Addition of activated HSC-EXOs to macrophages stimulated IL-6 and TNFα mRNA expression and protein secretion in mouse and human macrophages, but not for rat HSC-EXO-macrophages. Addition of human EXOs to macrophages stimulated migration, effects mirrored by the direct addition of rhIL-6 and rhTNFα.ConclusionsHSC-EXOs associate with macrophages and stimulate cytokine synthesis-release and macrophage migration. Constructing a comprehensive understanding of EXO interactions between liver cell populations in the setting of inflammation/fibrosis increases the potential for developing new diagnostic/therapeutic approaches.
| S-EPMC8206040 | biostudies-literature
Project description:Hepatic fibrosis is a major consequence of chronic liver disease such as non-alcoholic steatohepatitis which is undergoing a dramatic evolution given the obesity progression worldwide, and has no treatment to date. Hepatic stellate cells (HSCs) play a key role in the fibrosis process, because in chronic liver damage, they transdifferentiate from a "quiescent" to an "activated" phenotype responsible for most the collagen deposition in liver tissue. Here, using a diet-induced liver fibrosis murine model (choline-deficient amino acid-defined, high fat diet), we characterized a specific population of HSCs organized as clusters presenting simultaneously hypertrophy of retinoid droplets, quiescent and activated HSC markers. We showed that hypertrophied HSCs co-localized with fibrosis areas in space and time. Importantly, we reported the existence of this phenotype and its association with collagen deposition in three other mouse fibrosis models, including CCl4-induced fibrosis model. Moreover, we have also shown its relevance in human liver fibrosis associated with different etiologies (obesity, non-alcoholic steatohepatitis, viral hepatitis C and alcoholism). In particular, we have demonstrated a significant positive correlation between the stage of liver fibrosis and HSC hypertrophy in a cohort of obese patients with hepatic fibrosis. These results lead us to conclude that hypertrophied HSCs are closely associated with hepatic fibrosis in a metabolic disease context and may represent a new marker of metabolic liver disease progression.
| S-EPMC7052210 | biostudies-literature