scholarly journals Expression of P2Y nucleotide receptors and ectonucleotidases in quiescent and activated rat hepatic stellate cells

2004 ◽  
Vol 287 (2) ◽  
pp. G417-G424 ◽  
Author(s):  
Jonathan A. Dranoff ◽  
Mika Ogawa ◽  
Emma A. Kruglov ◽  
Marianna D. A. Gaça ◽  
Jean Sévigny ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Hepatic Stellate Cells ◽  
Stellate Cells ◽  
P2y Receptors ◽  
Extracellular Nucleotides ◽  
Nucleoside Triphosphate ◽  
P2y Receptor ◽  
Rt Pcr ◽  
Nucleoside Triphosphate Diphosphohydrolase ◽  
Receptor Inhibitor

Extracellular nucleotides regulate a variety of cellular activities, including proliferation of fibrogenic cells outside of the liver. However, the expression of receptors for extracellular nucleotides in hepatic stellate cells (HSC) is unknown. Thus our aims were to investigate the expression of mediators of nucleotide signaling in HSC and to determine whether extracellular nucleotides regulate HSC function. Confocal video microscopy was used to observe nucleotide-induced changes in cytosolic Ca2+ (Cai2+) in live HSC. P2Y receptor subtype expression and ectonucleotidase expression in quiescent and activated HSC were determined using RT-PCR, Northern blot, immunoblot, and confocal immunofluorescence. Functional ectonucleotidase activity was assessed using a colorimetric method. Nucleotide-sensitive procollagen-1 mRNA expression in activated HSC was assessed using real-time RT-PCR. Extracellular ATP increased Cai2+ in HSC; this was inhibited by the P2 receptor inhibitor suramin. Quiescent HSC expressed the P2Y subtypes P2Y2 and P2Y4 and were activated by ATP and UTP, whereas activated HSC expressed the P2Y subtype P2Y6 and were activated by UDP and ATP. Activated but not quiescent HSC expressed the ectonucleotidase nucleoside triphosphate diphosphohydrolase 2, extracellular UDP tripled procollagen-1 mRNA expression in activated HSC, and this was inhibited by the P2Y receptor inhibitor suramin. HSC express functional P2Y receptors and switch the expression of P2Y receptor subtypes on activation. Moreover, HSC differentially regulate nucleoside triphosphate diphosphohydrolase expression after activation. Because activation of P2Y receptors in activated HSC regulates procollagen-1 transcription, P2Y receptors may be an attractive target to prevent or treat liver fibrosis.

scholarly journals Role of Ca2+ in Mediating Plant Responses to Extracellular ATP and ADP

2018 ◽  
Vol 19 (11) ◽  
pp. 3590 ◽  
Author(s):  
Greg Clark ◽  
Stanley Roux
Keyword(s):  
Cytosolic Calcium ◽  
Defense Responses ◽  
Receptor Activation ◽  
Extracellular Atp ◽  
Extracellular Nucleotides ◽  
Nucleoside Triphosphate ◽  
Plant Responses ◽  
Downstream Signaling ◽  
Nucleoside Triphosphate Diphosphohydrolase

Among the most recently discovered chemical regulators of plant growth and development are extracellular nucleotides, especially extracellular ATP (eATP) and extracellular ADP (eADP). Plant cells release ATP into their extracellular matrix under a variety of different circumstances, and this eATP can then function as an agonist that binds to a specific receptor and induces signaling changes, the earliest of which is an increase in the concentration of cytosolic calcium ([Ca2+]cyt). This initial change is then amplified into downstream-signaling changes that include increased levels of reactive oxygen species and nitric oxide, which ultimately lead to major changes in the growth rate, defense responses, and leaf stomatal apertures of plants. This review presents and discusses the evidence that links receptor activation to increased [Ca2+]cyt and, ultimately, to growth and diverse adaptive changes in plant development. It also discusses the evidence that increased [Ca2+]cyt also enhances the activity of apyrase (nucleoside triphosphate diphosphohydrolase) enzymes that function in multiple subcellular locales to hydrolyze ATP and ADP, and thus limit or terminate the effects of these potent regulators.

scholarly journals Effect of Fibroblast Growth Factor 21 on the Expression of TLR4 and BAMBI Genes in Cholesterol-Treated Human Hepatic Stellate Cells

2021 ◽  
Vol 12 (3) ◽  
Author(s):  
Elham Shakerian ◽  
Narges Mohammad Taghvaei ◽  
Zohre Askari ◽  
Reza Afarin
Keyword(s):  
Growth Factor ◽  
Mrna Expression ◽  
Fibroblast Growth Factor ◽  
Hepatic Stellate Cells ◽  
Stellate Cells ◽  
Fibroblast Growth Factor 21 ◽  
Control Group ◽  
Type I ◽  
Fibroblast Growth ◽  
Human Hscs

Background: Activated hepatic stellate cells (HSCs) are the primary mediators in the progression of hepatic fibrosis. The activation of toll-like receptor 4 (TLR4) signaling leads to the downregulation of the transmembrane inhibitory transforming growth factor-beta (TGF-β) pseudoreceptor BMP and activin membrane-bound inhibitor (BAMBI) on HSCs. Fibroblast growth factor 21 (FGF21) is a natural secretory protein in the body with effects, such as the reduction of fat accumulation and oxidation of lipids; however; no study has investigated FGF21 ability to prevent the progression of liver fibrosis. Objectives: This study aimed to examine the beneficial effects of FGF21 to reduce cholesterol-activated human HSCs. Methods: The human HSCs were incubated in media containing different concentrations of cholesterol, including 25, 50, 75, 100, 125, and 150 μM, for 24 h and then incubated with FGF21 for 24 h. Total ribonucleic acids were extracted and reversely transcribed into complementary deoxyribonucleic acid. A quantitative real-time polymerase chain reaction was performed in this study. Results: The results showed that the messenger ribonucleic acid (mRNA) expression of TGF-β, collagen, type I, alpha 1 (collagen1α), and TLR4 genes increased significantly in the presence of cholesterol (75 and 100 μM), compared to that of the control group (* P < 0.05, ** P < 0.01, and *** P < 0.001); nevertheless, the mRNA expression of the BAMBI gene significantly reduced, compared to that of the control group (* P < 0.05). The FGF21 significantly reduced the mRNA expression of TGF-β, collagen1α, and TLR4 genes (# P < 0.05). The mRNA expression of the BAMBI gene significantly increased with FGF21 (# P < 0.05). Conclusions: It was concluded that the treatment with FGF21 reduces the cholesterol-activated HSCs by decreasing the mRNA expression of the TLR4, TGF-β, and collagen1α genes and increasing the mRNA expression of the BAMBI gene.

Molecular basis for calcium signaling in hepatic stellate cells

2007 ◽  
Vol 292 (4) ◽  
pp. G975-G982 ◽  
Author(s):  
Emma A. Kruglov ◽  
Paulo R. A. V. Correa ◽  
Gaurav Arora ◽  
Jin Yu ◽  
Michael H. Nathanson ◽  
...  
Keyword(s):  
Liver Fibrosis ◽  
Hepatic Stellate Cells ◽  
Stellate Cells ◽  
P2y Receptors ◽  
Type I ◽  
Inositol 1,4,5 Trisphosphate ◽  
Trisphosphate Receptor ◽  
Healthy Liver ◽  
Organ Fibrosis ◽  
Injured Liver

Progressive liver fibrosis (with the resultant cirrhosis) is the primary cause of chronic liver failure. Hepatic stellate cells (HSCs) are critically important mediators of liver fibrosis. In the healthy liver, HSCs are quiescent lipid-storing cells limited to the perisinusoidal endothelium. However, in the injured liver, HSCs undergo myofibroblastic transdifferentiation (activation), which is a critical step in the development of organ fibrosis. HSCs express P2Y receptors linking extracellular ATP to inositol (1,4,5)-trisphosphate-mediated cytosolic Ca2+ signals. Here, we report that HSCs express only the type I inositol (1,4,5)-trisphosphate receptor and that the receptor shifts into the nucleus and cell extensions upon activation. These cell extensions, furthermore, express sufficient machinery to enable local application of ATP to evoke highly localized Ca2+ signals that induce localized contractions. These autonomous units of subcellular signaling and response reveal a new level of subcellular organization, which, in turn, establishes a novel paradigm for the local control of fibrogenesis in the liver.

scholarly journals MiR-571 affects the development and progression of liver fibrosis by regulating the Notch3 pathway

2021 ◽  
Author(s):  
Shuo Cong ◽  
Yongmei Liu ◽  
Yi Li ◽  
Yu Chen ◽  
Rui Chen ◽  
...  
Keyword(s):  
Liver Fibrosis ◽  
Western Blot ◽  
Signaling Pathway ◽  
Hepatic Stellate Cells ◽  
Stellate Cells ◽  
Rt Pcr ◽  
Promote Cell Proliferation ◽  
Notch 3 ◽  
And Migration ◽  
Set Up

Abstract Exploring the expression of miR-571 in patients with liver fibrosis and its role in the progression of liver fibrosis. A total of 74 patients with chronic hepatitis and cirrhosis accompanied by liver fibrosis in our institution from September to December 2018 were collected for study, and the expression of miR-571 in patients with different progressions of liver fibrosis was determined by RT-PCR and Western blot analysis. Set up Notch3 up group and Notch3 down regulated group, RT-PCR and Western blot were used to determine the effect of Notch signaling on the expression of fibrogenic α-SMA, collagen I. CCK-8, cell scratch assays, Transwell assays, flow cytometry were used to determine the effect of miR-571 on LX-2 proliferation, migration, apoptosis in human stem stellate cells, and RT-PCR, Western blot assays were performed to determine the effect of miR-571 on the Notch3 signaling pathway and the expression of profibrogenic factors. miR-571 is up-regulated in patients with liver fibrosis and is associated with the progression of liver fibrosis. Notch3 signaling pathway can promote the expression of fibroblast in human hepatic stellate cells; miR-571 can inhibit the apoptosis of human hepatic stellate cells, promote cell proliferation and migration; up regulation of miR-571 can promote the expression of Notch3 and Jagged 1; up regulation of miR-571 can also promote the expression of fibroblast. miR-571 can promote the activation of human stem stellate cells and the expression of fibroblasts through Notch 3 signaling pathway.

Exposure to bacterial cell wall products triggers an inflammatory phenotype in hepatic stellate cells

2005 ◽  
Vol 289 (3) ◽  
pp. G571-G578 ◽  
Author(s):  
Paola Brun ◽  
Ignazio Castagliuolo ◽  
Massimo Pinzani ◽  
Giorgio Palù ◽  
Diego Martines
Keyword(s):  
Cell Wall ◽  
Liver Damage ◽  
Hepatic Stellate Cells ◽  
Bacterial Cell ◽  
Stellate Cells ◽  
Bacterial Cell Wall ◽  
Rt Pcr ◽  
Systemic Complications ◽  
Bacterial Endotoxins ◽  
Extracellular Matrix Components

Activated hepatic stellate cells (HSCs) secrete extracellular matrix components during hepatic fibrosis, but recent studies have shown that HSCs can also release a variety of proinflammatory cytokines. Moreover, bacterial endotoxemia is not only associated with systemic complications in the late stages of liver failure but is also a direct cause of liver damage, activating resident inflammatory cells. In this study, we investigated whether HSCs can respond directly to bacterial cell wall products acquiring a new phenotype. RT-PCR and immunocytochemistry assays were used to show that murine HSCs expressed specific mRNA transcripts and proteins for LPS and lipoteichoic acid (LTA) receptor systems and peptidoglycan recognition proteins. Exposing HSCs to bacterial endotoxins led to phosphorylation of mitogen-activated protein kinase ERK1 and the development of a proinflammatory phenotype. After exposure to LPS, LTA, or N-acetyl muramyl peptide, transforming growth factor-β1, IL-6, and monocyte chemoattractant protein-1 (MCP-1) mRNA specific transcripts and proteins increased significantly in HSCs, as assayed by quantitative real-time RT-PCR and ELISA. These LPS-mediated effects in HSCs were receptor dependent, because LPS-induced ERK1 phosphorylation, IL-6, and MCP-1 mRNA and protein level upregulation were significantly less pronounced in HSCs isolated from C3H/HeJ mice lacking Toll-like receptor 4. In conclusion, our results show that murine HSCs express functional receptors for bacterial endotoxins, and HSCs exposed to bacterial products develop a strong proinflammatory phenotype. We speculate that high levels of bacterial endotoxins in the portal vein may directly induce a proinflammatory phenotype in HSCs that contributes to liver damage.

scholarly journals Effect of Exogenous Fetuin-A on TGF-β/Smad Signaling in Hepatic Stellate Cells

2016 ◽  
Vol 2016 ◽  
pp. 1-6 ◽  
Author(s):  
Yulai Zhou ◽  
Shuang Yang ◽  
Pan Zhang
Keyword(s):  
Western Blot ◽  
Hepatic Stellate Cells ◽  
Stellate Cells ◽  
Relative Content ◽  
Negative Regulation ◽  
Rt Pcr ◽  
Fetuin A ◽  
Smad3 Protein ◽  
Smad7 Protein ◽  
Excessive Activation

Objective. To explore the effects of low concentration of exogenous fetuin-A intervention on TGF-β1 induced LX2 cells through detection of the expression of mRNA and protein of Smad2, Smad3, and Smad7. Methods. MTT assay was used to detect the LX2 cells proliferation and the regression equation calculating software was applied to determine IC50 of fetuin-A. RT-PCR was used to determine the relative content of Smad2, Smad3, and Smad7 mRNA in LX2 cells. Western blot was used to detect the LX2 cells relative content of Smad2, Smad3, Smad7 protein expression, respectively. Results. The analysis from RT-PCR and western blot showed that when compared with the other groups TGF-β1 + fetuin-A group increased the expression of Smad2 and Smad3 while decreased the expression of Smad7 (P<0.05). Conclusion. Fetuin-A may improve the excessive activation of hepatic stellate cells which is caused by an enhanced positive regulation of Smad2 and Smad3 protein and the deficiency in negative regulation of Smad7 protein. This is through inhibiting the expression of Smad2 and Smad3 gene and promoting the expression of Smad7 gene. As a result, the development of liver fibrosis will be reduced.

scholarly journals Hepatic stellate cells (HSC) cell line (LX-2) and Toll-like receptors – phenotypic/mRNA expression and in vitro functional studies

2013 ◽  
Vol 4 ◽  
pp. 486-493
Author(s):  
Anna Mania ◽  
Mariusz Kaczmarek ◽  
Iwona Mozer-Lisewska ◽  
Arleta Kowala-Piaskowska ◽  
Agata Kolecka-Bednarczyk ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Cell Line ◽  
Hepatic Stellate Cells ◽  
Stellate Cells ◽  
Toll Like Receptors ◽  
Functional Studies

Differential catalytic properties and vascular topography of murine nucleoside triphosphate diphosphohydrolase 1 (NTPDase1) and NTPDase2 have implications for thromboregulation

Blood ◽  
2002 ◽  
Vol 99 (8) ◽  
pp. 2801-2809 ◽  
Author(s):  
Jean Sévigny ◽  
Christian Sundberg ◽  
Norbert Braun ◽  
Olaf Guckelberger ◽  
Eva Csizmadia ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Adenosine Monophosphate ◽  
Adenosine Diphosphate ◽  
Receptor Activation ◽  
Extracellular Nucleotides ◽  
Nucleoside Triphosphate ◽  
Uridine Diphosphate ◽  
P2 Receptor ◽  
Nucleoside Triphosphate Diphosphohydrolase ◽  
Vessel Injury

Abstract Nucleoside triphosphate diphosphohydrolases (NTPDases) are a recently described family of ectonucleotidases that differentially hydrolyze the γ and β phosphate residues of extracellular nucleotides. Expression of this enzymatic activity has the potential to influence nucleotide P2 receptor signaling within the vasculature. We and others have documented that NTPDase1 (CD39, 78 kd) hydrolyzes both triphosphonucleosides and diphosphonucleosides and thereby terminates platelet aggregation responses to adenosine diphosphate (ADP). In contrast, we now show that NTPDase2 (CD39L1, 75 kd), a preferential nucleoside triphosphatase, activates platelet aggregation by converting adenosine triphosphate (ATP) to ADP, the specific agonist of P2Y1 and P2Y12 receptors. We developed specific antibodies to murine NTPDase1 and NTPDase2 and observed that both enzymes are present in the cardiac vasculature; NTPDase1 is expressed by endothelium, endocardium, and to a lesser extent by vascular smooth muscle, while NTPDase2 is associated with the adventitia of muscularized vessels, microvascular pericytes, and other cell populations in the subendocardial space. Moreover, NTPDase2 represents a novel marker for microvascular pericytes. Differential expression of NTPDases in the vasculature suggests spatial regulation of nucleotide-mediated signaling. In this context, NTPDase1 should abrogate platelet aggregation and recruitment in intact vessels by the conversion of ADP to adenosine monophosphate, while NTPDase2 expression would promote platelet microthrombus formation at sites of extravasation following vessel injury. Our data suggest that specific NTPDases, in tandem with ecto-5′-nucleotidase, not only terminate P2 receptor activation and trigger adenosine receptors but may also allow preferential activation of specific subsets of P2 receptors sensitive to ADP (eg, P2Y1, P2Y3, P2Y12) and uridine diphosphate (P2Y6).

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