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Rat HSCs (rat + hsc)
Selected AbstractsMultidrug resistance,associated proteins are crucial for the viability of activated rat hepatic stellate cells,,HEPATOLOGY, Issue 2 2008Rebekka A. Hannivoort Hepatic stellate cells (HSCs) survive and proliferate in the chronically injured liver. ATP-binding cassette (ABC) transporters play a crucial role in cell viability by transporting toxic metabolites or xenobiotics out of the cell. ABC transporter expression in HSCs and its relevance to cell viability and/or activation have not been reported so far. The aim of this study was to investigate the expression, regulation, and function of multidrug resistance,associated protein (Mrp)-type and multidrug resistance protein (Mdr),type ABC transporters in activated rat HSCs. Rat HSCs were exposed to cytokines or oxidative stress. ABC transporter expression was determined by quantitative polymerase chain reaction and immunohistochemistry. HSCs were exposed to the Mdr inhibitors verapamil and PSC-833 and the Mrp inhibitor MK571. Mdr and Mrp transporter function was evaluated with flow cytometry. Apoptosis was determined by activated caspase-3 and acridine orange staining, and necrosis was determined by Sytox green nuclear staining. An in vivo model of carbon tetrachloride (CCl4),induced liver fibrosis was used. With respect to hepatocytes, activated HSCs expressed high levels of Mrp1 and comparable levels of Mrp3, Mrp4, Mdr1a, and Mdr1b but not the hepatocyte-specific transporters bile salt export pump, Mrp2, and Mrp6. Mrp1 protein staining correlated with desmin staining in livers from CCl4 -treated rats. Mrp1 expression increased upon activation of HSCs. Cytokines induced Mdr1b expression only. Oxidative stress was not a major regulator of Mdr and Mrp transporter expression. Activated HSCs became necrotic when exposed to the Mrp inhibitors. Conclusion: Activated HSCs contain relatively high levels of Mrp1. Mrp-type transporters are required for the viability of activated HSCs. Mrp-dependent export of endogenous metabolites is important for the survival of activated HSCs in chronic liver diseases. (HEPATOLOGY 2008.) [source] Differential regulation of TGF-, signal in hepatic stellate cells between acute and chronic rat liver injuryHEPATOLOGY, Issue 1 2002Yoshiya Tahashi During chronic liver injury, transforming growth factor , (TGF-,) plays a prominent role in stimulating liver fibrogenesis by myofibroblast-like cells derived from hepatic stellate cells (HSCs). On the other hand, Smad 7 was recently shown to antagonize the TGF-,,induced activation of signal-transducing Smads (2 and 3). In this study, we investigated the regulatory mechanisms of the TGF-, signals in rat HSCs during acute liver injury and myofibroblasts (MFBs) during chronic liver injury, focusing on the roles of Smad 2 and antagonistic Smad 7. In acute liver injury, HSC-derived TGF-, increased plasminogen activator inhibitor type 1 (PAI-1) and ,2(I) procollagen (COL1A2) transcripts. Smad 2 in HSCs during liver injury and primary cultured HSCs were activated by an autocrine mechanism, because high levels of Smad 2 phosphorylation and induction of PAI-1 transcript by TGF-, were observed in HSCs. Thereafter, Smad 7 induced by TGF-, negatively regulated the Smad 2 action. These results indicated that endogenous TGF,,mediated Smad 7 in HSCs terminated the fibrotic signals mediated by signal-transducing Smads, and might be involved in the transient response to autocrine TGF-, signal after acute liver injury. By contrast, Smad 7 was not induced by the autocrine TGF-, signal, and constitutive Smad 2 activation was observed in MFBs throughout chronic liver injury, although Smad 7 could inhibit the TGF-, signal requiring Smad 2 phosphorylation by activated TGF-, receptor in cultured MFBs. This constitutive phosphorylation of Smad 2 by endogenous TGF-, under a low level of Smad 7 could be involved in the progression of liver fibrosis. [source] Superoxide anions and hydrogen peroxide inhibit proliferation of activated rat stellate cells and induce different modes of cell deathLIVER INTERNATIONAL, Issue 6 2009Sandra Dunning Abstract Background: In chronic liver injury, hepatic stellate cells (HSCs) proliferate and produce excessive amounts of connective tissue causing liver fibrosis and cirrhosis. Oxidative stress has been implicated as a driving force of HSC activation and proliferation, although contradictory results have been described. Aim: To determine the effects of oxidative stress on activated HSC proliferation, survival and signalling pathways. Methods: Serum-starved culture-activated rat HSCs were exposed to the superoxide anion donor menadione (5,25 ,mol/L) or hydrogen peroxide (0.2,5 mmol/L). Haem oxygenase-1 mRNA expression, glutathione status, cell death, phosphorylation of mitogen-activated protein (MAP) kinases and proliferation were investigated. Results: Menadione induced apoptosis in a dose- and time-dependent, but caspase-independent manner. Hydrogen peroxide induced necrosis only at extremely high concentrations. Both menadione and hydrogen peroxide activated Jun N-terminal kinase (JNK) and p38. Hydrogen peroxide also activated extracellular signal-regulated protein. Menadione, but not hydrogen peroxide, reduced cellular glutathione levels. Inhibition of JNK or supplementation of glutathione reduced menadione-induced apoptosis. Non-toxic concentrations of menadione or hydrogen peroxide inhibited platelet-derived growth factor- or/and serum-induced proliferation. Conclusion: Reactive oxygen species (ROS) inhibit HSC proliferation and promote HSC cell death in vitro. Different ROS induce different modes of cell death. Superoxide anion-induced HSC apoptosis is dependent on JNK activation and glutathione status. [source] Atorvastatin induces apoptosis by a caspase-9-dependent pathway: an in vitro study on activated rat hepatic stellate cellsLIVER INTERNATIONAL, Issue 4 2008Isabella Aprigliano Abstract Background: Statins are shown to have cholesterol-independent properties such as anti-inflammation and immunomodulation. Activated hepatic stellate cells (HSCs) acquire the capacity to synthesize matrix proteins in damaged liver. We tested the hypothesis that atorvastatin may be capable of inducing apoptosis in HSCs. Methods: Primary cultures of rat HSCs were exposed to atorvastatin, mevalonic acid and U0126. Quantification of living, apoptotic and necrotic HSCs was performed by flow cytometry and laser-scan microscopy. Cell-cycle analysis was performed by flow cytometry. Pro- and anti-apoptotic factors were investigated by Western blot and electrophoresis mobility shift assay. Protease activity of caspases was calculated using a colorimetric kit. Results: Atorvastatin leads to a G2-arrest and induces apoptosis in activated HSCs. Atorvastatin-mediated apoptosis could be blocked by co-administration of mevalonic acid and U0126. No effects of atorvastatin on gene expression of CD95, CD95L, NF-,B, p53 and p21WAF1 could be observed. Atorvastatin-induced apoptosis in activated HSCs is related to an increased protease activity of caspase-9 and -3. Gene expression of the major proteins of the bcl-system shows that truncated Bid is involved in apoptosis mediated by atorvastatin. By blocking the extracellular signal-regulated protein kinase (ERK1/2) activation by adding U0126, we could prevent the apoptosis induced by atorvastatin. By Western blot we could not detect any change in the activation of c-jun N-terminal kinase (JNK). Conclusions: Atorvastatin induces apoptosis in activated HSCs acting through an ERK-dependent cleavage of Bid and a highly increased protease activity of caspase-9 and -3. JNK is not involved in atorvastatin-mediated apoptosis in HSCs. [source] Ganoderma lucidum extract attenuates the proliferation of hepatic stellate cells by blocking the PDGF receptorPHYTOTHERAPY RESEARCH, Issue 6 2009Guei-Jane Wang Abstract Hepatic fibrosis is an outcome of chronic liver diseases. The activation and proliferation of hepatic stellate cells (HSCs) is a key event in liver injury. The fruiting body of Ganoderma lucidum has long been a popular oriental medicine for treating liver diseases. The aim of this present study was to investigate the antiproliferative effects of the triterpenoid-rich extract (GLT) of G. lucidum in a cell line of rat HSCs (HSC-T6) stimulated with platelet-derived growth factor (PDGF)-BB. DNA synthesis was investigated by bromodeoxyuridine (BrdU) incorporation. Flow cytometry using propidium iodide (PI) labeling was carried out to analyse the cell cycle distribution and apoptosis. , -Smooth muscle actin (, -SMA) was used to evaluate extracellular matrix deposition, and western blotting was performed to measure cyclins D1 and D2, and phosphorylation of the PDGF, -receptor (PDGF,R), Akt and JNK. The results indicated that the GLT attenuated BrdU incorporation in a concentration-dependent manner with an IC50 of 8.52 ± 0.33 µg/mL. The inhibitory effect of the GLT was associated with downregulation of cyclins D1 and D2, and PDGF,R and Akt phosphorylation, upregulation of JNK phosphorylation, and a reduction in , -SMA expression. These results indicated that G. lucidum inhibits PDGF-BB-activated HSC proliferation possibly through blocking PDGF,R phosphorylation, thereby indicating its efficacy for preventing and treating hepatic fibrosis. Copyright © 2008 John Wiley & Sons, Ltd. [source] In vivo and in vitro Interactions between Human Colon Carcinoma Cells and Hepatic Stellate CellsCANCER SCIENCE, Issue 12 2000Sadatoshi Shimizu Stromal reaction is important for the growth of cancer both in primary and metastatic sites. To demonstrate this reaction during the hepatic metastasis of human colon carcinoma, we histologically investigated alterations to the distribution and phenotype of hepatic stellate cells (HSCs), the only mesenchymal cells in the liver parenchyma, using a nude mouse model. Intrasplenically injected colon carcinoma LM-H3 cells migrated into the space of Disse and underwent proliferation, in close association with hepatocytes and HSCs, at 2 days. At 14 days, HSCs were accumulated around the tumor mass and expressed ,-smooth muscle actin, a marker for HSC activation. We next investigated in vitro the growth factors involved in the interactions between LM-H3 cells and HSCs. Conditioned medium of rat HSCs which underwent culture-induced activation contained platelet-derived growth factor (PDGF)-AB, hepatocyte growth factor (HGF) and transforming growth factor (TGF),, and could augment LM-H3-cell proliferation and migration. Neutralizing antibodies against PDGF-AA and PDGF-BB and those against PDGF-BB and HGF inhibited proliferation and migration, respectively, of LM-H3 cells, whereas antibody against TGF-, had no effect. LM-H3 cells expressed PDGF receptors-, and -, and c-met. Conditioned medium of LM-H3 cells contained PDGF-AB, and could enhance HSC proliferation and migration. This augmenting effect was suppressed by treatment with anti-PDGF-AB antibody. The present study has demonstrated that bidirectional interactions involving PDGF and HGF take place in vitro between colon carcinoma cells and HSCs, raising the possibility that similar interactions might be involved in the stromal reaction during hepatic metastasis. [source] | ||||||||||