Expression of cellular prion protein in activated hepatic stellate cells.
Ontology highlight
ABSTRACT:
SUBMITTER: Ikeda K
PROVIDER: S-EPMC1866339 | biostudies-literature | 1998 Dec
REPOSITORIES: biostudies-literature
Publications
Expression of cellular prion protein in activated hepatic stellate cells.
Ikeda K K Kawada N N Wang Y Q YQ Kadoya H H Nakatani K K Sato M M Kaneda K K
The American journal of pathology 19981201 6
Suppression subtractive hybridization was used to clone genes associated with the activation of hepatic stellate cells and 13 genes were found to be dominantly expressed in activated stellate cells. Among them, one was identical to the 421-837th base pairs of cDNA sequence reported for rat prion-related protein (PrP). In cultured stellate cells, PrP mRNA expression increased in a time-dependent manner in parallel with smooth muscle (SM) alpha-actin mRNA expression. In situ hybridization demonstr ...[more]
Similar Datasets
Project description:Hepatic stellate cells (HSC) are the major cellular drivers of liver fibrosis. Upon liver inflammation caused by a broad range of insults including non-alcoholic fatty liver, HSC transform from a quiescent into a proliferating, fibrotic phenotype. Although much is known about the pathophysiology of this process, exact cellular processes which occur in HSC and enable this transformation remain yet to be elucidated. In order to investigate this HSC transformation, we employed a simple, yet reliable model of HSC activation via an increase in growth medium serum concentration (serum activation). For that purpose, immortalized human LX-2 HSC were exposed to either 1% or 10% fetal bovine serum (FBS). Resulting quiescent (1% FBS) and activated (10% FBS) LX-2 cells were then subjected to in-depth mass spectrometry-based proteomics analysis as well as comprehensive phenotyping. Protein network analysis of activated LX-2 cells revealed an increase in the production of ribosomal proteins and proteins related to cell cycle control and migration, resulting in higher proliferation and faster migration phenotypes. Interestingly, we also observed a decrease in the expression of cholesterol and fatty acid biosynthesis proteins in accordance with a concomitant loss of cytosolic lipid droplets during activation. Overall, this work provides an update on HSC activation characteristics using contemporary proteomic and bioinformatic analyses and presents an accessible model for HSC activation. Data are available via ProteomeXchange with identifier PXD029121.
| S-EPMC8657869 | biostudies-literature
Project description:Bone marrow-derived mesenchymal stem cells (MSCs) have been reported to prevent the development of liver fibrosis in a number of pre-clinical studies. Marked changes in liver histopathology and serological markers of liver function have been observed without a clear understanding of the therapeutic mechanism by which stem cells act. We sought to determine if MSCs could modulate the activity of resident liver cells, specifically hepatic stellate cells (SCs) by paracrine mechanisms using indirect cocultures. Indirect coculture of MSCs and activated SCs led to a significant decrease in collagen deposition and proliferation, while inducing apoptosis of activated SCs. The molecular mechanisms underlying the modulation of SC activity by MSCs were examined. IL-6 secretion from activated SCs induced IL-10 secretion from MSCs, suggesting a dynamic response of MSCs to the SCs in the microenvironment. Blockade of MSC-derived IL-10 and TNF-alpha abolished the inhibitory effects of MSCs on SC proliferation and collagen synthesis. In addition, release of HGF by MSCs was responsible for the marked induction of apoptosis in SCs as determined by antibody-neutralization studies. These findings demonstrate that MSCs can modulate the function of activated SCs via paracrine mechanisms provide a plausible explanation for the protective role of MSCs in liver inflammation and fibrosis, which may also be relevant to other models of tissue fibrosis.
| S-EPMC2096777 | biostudies-literature
Project description:SerpinB3 is a hypoxia- and hypoxia-inducible factor-2?-dependent cystein protease inhibitor that is up-regulated in hepatocellular carcinoma and in parenchymal cells during chronic liver diseases (CLD). SerpinB3 up-regulation in CLD patients has been reported to correlate with the extent of liver fibrosis and the production of transforming growth factor-?1, but the actual role of SerpinB3 in hepatic fibrogenesis is still poorly characterized. In the present study we analyzed the pro-fibrogenic action of SerpinB3 in cell cultures and in two different murine models of liver fibrosis. "In vitro" experiments revealed that SerpinB3 addition to either primary cultures of human activated myofibroblast-like hepatic stellate cells (HSC/MFs) or human stellate cell line (LX2 cells) strongly up-regulated the expression of genes involved in fibrogenesis and promoted oriented migration, but not cell proliferation. Chronic liver injury by CCl4 administration or by feeding a methionine/choline deficient diet to transgenic mice over-expressing human SerpinB3 in hepatocytes confirmed that SerpinB3 over-expression significantly increased the mRNA levels of pro-fibrogenic genes, collagen deposition and ?SMA-positive HSC/MFs as compared to wild-type mice, without affecting parenchymal damage. The present study provides for the first time evidence that hepatocyte release of SerpinB3 during CLD can contribute to liver fibrogenesis by acting on HSC/MFs.
| S-EPMC5469760 | biostudies-literature
Project description:BackgroundLiver fibrosis is characterized by the excessive formation and accumulation of matrix proteins as a result of wound healing in the liver. A main event during fibrogenesis is the activation of the liver resident quiescent hepatic stellate cell (qHSC). Recent studies suggest that reversion of the activated HSC (aHSC) phenotype into a quiescent-like phenotype could be a major cellular mechanism underlying fibrosis regression in the liver, thereby offering new therapeutic perspectives for the treatment of liver fibrosis. Whether human HSCs have the ability to undergo a similar reversion in phenotype is currently unknown. The aim of the present study is to identify experimental conditions that can revert the in vitro activated phenotype of primary human HSCs and consequently to map the molecular events associated with this reversion process by gene expression profiling.ResultsWe find that epidermal growth factor (EGF) and fibroblast growth factor 2 (FGF2) synergistically downregulate the expression of ACTA2 and LOX in primary human aHSCs. Their combination with oleic acid, palmitic acid, and retinol further potentiates a more quiescent-like phenotype as demonstrated by the abundant presence of retinyl ester-positive intra-cytoplasmic lipid droplets, low expression levels of activation markers, and a reduced basal as well as cytokine-stimulated proliferation and matrix metalloproteinase activity. Gene expression profiling experiments reveal that these in vitro reverted primary human HSCs (rHSCs) display an intermediary phenotype that is distinct from qHSCs and aHSCs. Interestingly, this intermediary phenotype is characterized by the increased expression of several previously identified signature genes of in vivo inactivated mouse HSCs such as CXCL1, CXCL2, and CTSS, suggesting also a potential role for these genes in promoting a quiescent-like phenotype in human HSCs.ConclusionsWe provide evidence for the ability of human primary aHSCs to revert in vitro to a transitional state through synergistic action of EGF, FGF2, dietary fatty acids and retinol, and provide a first phenotypic and genomic characterization of human in vitro rHSCs.
| S-EPMC4527231 | biostudies-literature
Project description:Alcoholic liver disease (ALD) is a leading cause of cirrhosis in the United States, which is characterized by extensive deposition of extracellular matrix proteins and formation of a fibrous scar. Hepatic stellate cells (HSCs) are the major source of collagen type 1 producing myofibroblasts in ALD fibrosis. However, the mechanism of alcohol-induced activation of human and mouse HSCs is not fully understood. We compared the gene-expression profiles of primary cultured human HSCs (hHSCs) isolated from patients with ALD (n = 3) or without underlying liver disease (n = 4) using RNA-sequencing analysis. Furthermore, the gene-expression profile of ALD hHSCs was compared with that of alcohol-activated mHSCs (isolated from intragastric alcohol-fed mice) or CCl4-activated mouse HSCs (mHSCs). Comparative transcriptome analysis revealed that ALD hHSCs, in addition to alcohol-activated and CCl4-activated mHSCs, share the expression of common HSC activation (Col1a1 [collagen type I alpha 1 chain], Acta1 [actin alpha 1, skeletal muscle], PAI1 [plasminogen activator inhibitor-1], TIMP1 [tissue inhibitor of metalloproteinase 1], and LOXL2 [lysyl oxidase homolog 2]), indicating that a common mechanism underlies the activation of human and mouse HSCs. Furthermore, alcohol-activated mHSCs most closely recapitulate the gene-expression profile of ALD hHSCs. We identified the genes that are similarly and uniquely up-regulated in primary cultured alcohol-activated hHSCs and freshly isolated mHSCs, which include CSF1R (macrophage colony-stimulating factor 1 receptor), PLEK (pleckstrin), LAPTM5 (lysosmal-associated transmembrane protein 5), CD74 (class I transactivator, the invariant chain), CD53, MMP9 (matrix metallopeptidase 9), CD14, CTSS (cathepsin S), TYROBP (TYRO protein tyrosine kinase-binding protein), and ITGB2 (integrin beta-2), and other genes (compared with CCl4-activated mHSCs). Conclusion: We identified genes in alcohol-activated mHSCs from intragastric alcohol-fed mice that are largely consistent with the gene-expression profile of primary cultured hHSCs from patients with ALD. These genes are unique to alcohol-induced HSC activation in two species, and therefore may become targets or readout for antifibrotic therapy in experimental models of ALD.
| S-EPMC7109347 | biostudies-literature
Project description:The matricellular protein connective tissue growth factor (CCN2) is considered a faithful marker of fibroblast activation in wound healing and in fibrosis. CCN2 is induced during activation of hepatic stellate cells (HSC). Here, we investigate the molecular basis of CCN2 gene expression in HSC. Fluoroscence activated cell sorting was used to investigate CCN2 expression in HSC in vivo in mice treated with CCl(4). CCN2 and TGF-beta mRNA expression were assessed by polymerase chain reaction as a function of culture-induced activation of HSC. CCN2 promoter/reporter constructs were used to map cis-acting elements required for basal and TGFbeta-induced CCN2 promoter activity. Real-time polymerase chain reaction analysis was used to further clarify signaling pathways required for CCN2 expression in HSC. CCl(4) administration in vivo increased CCN2 production by HSC. In vitro, expression of CCN2 and TGF-beta mRNA were concommitantly increased in mouse HSC between days 0 and 14 of culture. TGFbeta-induced CCN2 promoter activity required the Smad and Ets-1 elements in the CCN2 promoter and was reduced by TGFbeta type I receptor (ALK4/5/7) inhibition. CCN2 overexpression in activated HSC was ALK4/5/7-dependent. As CCN2 overexpression is a faithful marker of fibrogenesis, our data are consistent with the notion that signaling through TGFbeta type I receptors such as ALK5 contributes to the activation of HSC and hence ALK4/5/7 inhibition would be expected to be an appropriate treatment for liver fibrosis.
| S-EPMC2570012 | biostudies-other
Project description:To investigate serotonergic Ca²+ signaling and the expression of 5-hydroxytryptamine (5-HT) receptors, as well as Ca²+ transporting proteins, in hepatic stellate cells (HSCs).The intracellular Ca²+ concentration [Ca²+](i) of isolated rat HSCs was measured with a fluorescence microscopic imaging system. Quantitative PCR was performed to determine the transcriptional levels of 5-HT receptors and endoplasmic reticulum (ER) proteins involved in Ca²+ storage and release in cultured rat HSCs.Distinct from quiescent cells, activated HSCs exhibited [Ca²+](i) transients following treatment with 5-HT, which was abolished by U-73122, a phospholipase C inhibitor. Upregulation of 5-HT(2A) and 5-HT(2B) receptors, but not 5-HT?, was prominent during trans-differentiation of HSCs. Pretreatment with ritanserin, a 5-HT? antagonist, inhibited [Ca²+](i) changes upon application of 5-HT. Expression of type 1 inositol-5'-triphosphate receptor and type 2 sarcoplasmic/endoplasmic reticulum Ca²+ ATPase were also increased during activation of HSCs and serve as the major isotypes for ER Ca²+ storage and release in activated HSCs. Ca²+ binding chaperone proteins of the ER, including calreticulin, calnexin and calsequestrin, were up-regulated following activation of HSCs.The appearance of 5-HT-induced [Ca²+](i) response accompanied by upregulation of metabotropic 5-HT? receptors and Ca²+ transporting/chaperone ER proteins may participate in the activating process of HSCs.
| S-EPMC3020369 | biostudies-literature
Project description:Previous evidence indicates that adiponectin possesses antifibrogenic activity in inhibiting liver fibrosis. Therapeutic strategies, however, are limited by adiponectin quaternary structure and effective concentrations in circulation. Here we postulate a novel molecular mechanism, whereby adiponectin targets focal adhesion kinase (FAK) activity and disrupts key features of the fibrogenic response. Adiponectin-null (Ad(-/-)) mice and wild-type littermates were exposed to either saline or carbon tetrachloride (CCl4) for 6 wk. CCl4-gavaged mice were also injected with attenuated adenoviral adiponectin (Ad-Adn) or Ad-LacZ for 2 wk. Hepatic stellate cells (HSCs) were treated with or without adiponectin to elucidate signal transduction mechanisms. In vivo delivery of Ad-Adn markedly attenuates CCl4-induced expression of key integrin proteins and markers of HSC activation: αv, β3, β1, α2(I) collagen, and α-smooth muscle actin. Confocal experiments of liver tissues demonstrated that adiponectin delivery also suppressed vinculin and p-FAK activity in activated HSCs. In vitro, adiponectin induced dephosphorylation of FAK, mediated by a physical association with activated tyrosine phosphatase, Shp2. Conversely, Shp2 knockdown by siRNA significantly attenuated adiponectin-induced FAK deactivation, and expression of TIMP1 and α2(I) collagen was abolished in the presence of adiponectin and si-FAK. Finally, we documented that either adiponectin or the synthetic peptide with adiponectin properties, ADP355, suppressed p-FAK in synthetic matrices with stiffness measurements of 9 and 15 kPa, assessed by immunofluorescent imaging and quantitation. The in vivo and in vitro data presented indicate that disassembly of focal adhesion complexes in HSCs is pivotal for hepatic fibrosis therapy, now that small adiponectin-like peptides are available.
| S-EPMC6190967 | biostudies-literature
Project description:AimTo understand which and how different miRNAs are implicated in the process of hepatic stellate cell (HSC) activation.MethodsWe used microarrays to examine the differential expression of miRNAs during in vitro activation of primary HSCs (pHSCs). The transcriptome changes upon stable transfection of rno-miR-146a into an HSC cell line were studied using cDNA microarrays. Selected differentially regulated miRNAs were investigated by quantitative real-time polymerase chain reaction during in vivo HSC activation. The effect of miRNA mimics and inhibitor on the in vitro activation of pHSCs was also evaluated.ResultsWe found that 16 miRNAs were upregulated and 26 were downregulated significantly in 10-d in vitro activated pHSCs in comparison to quiescent pHSCs. Overexpression of rno-miR-146a was characterized by marked upregulation of tissue inhibitor of metalloproteinase-3, which is implicated in the regulation of tumor necrosis factor-α activity. Differences in the regulation of selected miRNAs were observed comparing in vitro and in vivo HSC activation. Treatment with miR-26a and 29a mimics, and miR-214 inhibitor during in vitro activation of pHSCs induced significant downregulation of collagen type I transcription.ConclusionOur results emphasize the different regulation of miRNAs in in vitro and in vivo activated pHSCs. We also showed that miR-26a, 29a and 214 are involved in the regulation of collagen type I mRNA.
| S-EPMC3122263 | biostudies-literature
Project description:The activation of hepatic stellate cells (HSCs) plays a vital role in the progression of liver fibrosis, and the induction of HSCs apoptosis may attenuate or reverse fibrogenesis. The therapeutic effects of etoposide(VP-16), a widely used anticancer agent, on HSCs apoptosis and liver fibrosis resolution are still unclear. Here, we report that VP-16 reduced the proliferation of LX-2 cells and led to significantly high levels of apoptosis, as indicated by Annexin V staining and the proteolytic cleavage of the executioner caspase-3 and PARP. Additionally, the unfolded protein response regulators CHOP, BIP, caspase-12, p-eIF2α and IRE1α, which are considered endoplasmic reticulum (ER) stress markers, were upregulated by VP-16. The strong inhibitory effect of VP-16 on LX-2 cells was mainly dependent on ER stress, which activated JNK signaling pathway. Remarkably, VP-16 treatment decreased the expression of α-SMA and type I collagen and simultaneously increased the ratio of matrix metalloproteinases (MMPs) to tissue inhibitor of matrix metalloproteinases (TIMPs). In contrast, VP-16 induced significantly more apoptosis in HSCs than in normal hepatocytes. Taken together, our findings demonstrate that VP-16 exerts a proapoptotic effect on LX-2 cells and has an antifibrogenic effect on collagen deposition, suggesting a new strategy for the treatment of liver fibrosis.
| S-EPMC5041150 | biostudies-literature