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Richard A Rippe

Publications and source records attributed to Richard A Rippe.

18 recordsLinked to original sources
PubMed2005

NF-kappaB activation in Kupffer cells after partial hepatectomy.

The transcription factor nuclear factor-kappaB (NF-kappaB) is activated during liver regeneration after partial hepatectomy. However, the physiological role and cellular localization of NF-kappaB activation are unresolved. In this study, we used an adenoviral vector expressing a mutated form of IkappaBalpha to inhibit NF-kappaB activity during liver regeneration. After partial hepatectomy in mice, introduction of Ad5IkappaB, but not a control virus (Ad5GFP), resulted in increased liver injury and decreased hepatocyte proliferation. Hepatocyte apoptosis was not observed. To investigate the kinetics and cellular localization of NF-kappaB-induced transcription during liver regeneration, we generated a transgenic mouse expressing enhanced green fluorescent protein (EGFP) under the transcriptional control of NF-kappaB cis elements (cis-NF-kappaB-EGFP). During liver regeneration, EGFP expression was detected within 12 h and was primarily located in Kupffer cells. Our data demonstrate that activation of NF-kappaB initially occurs in Kupffer cells after partial hepatectomy in mice.

Adenoviridae↗
PubMed2005

The role of p70S6K in hepatic stellate cell collagen gene expression and cell proliferation.

During fibrosis the hepatic stellate cell (HSC) undergoes a complex activation process characterized by increased proliferation and extracellular matrix deposition. The 70-kDa ribosomal S6 kinase (p70S6K) is activated by mitogens, growth factors, and hormones in a phosphatidylinositol 3-kinase-dependent manner. p70S6K regulates protein synthesis, proliferation, and cell cycle control. Because these processes are involved in HSC activation, we investigated the role of p70S6K in HSC proliferation, cell cycle control, and type I collagen expression. Platelet-derived growth factor (PDGF) stimulated p70S6K phosphorylation, which was blocked by LY294002, an inhibitor of phosphatidylinositol 3-kinase. Rapamycin blocked phosphorylation of p70S6K but had no affect on PDGF-induced Akt phosphorylation, positioning p70S6K downstream of Akt. Transforming growth factor-beta, which inhibits HSC proliferation, did not affect PDGF-induced p70S6K phosphorylation. Rapamycin treatment did not affect alpha1(I) collagen mRNA but reduced type I collagen protein secretion. Expression of smooth muscle alpha-actin was not affected by rapamycin treatment, indicating that HSC activation was not altered. Rapamycin inhibited serum-induced DNA synthesis approximately 2-fold. Moreover, rapamycin decreased expression of cyclins D1, D3, and E but not cyclin D2, Rb-Ser780, and Rb-Ser795. Together, p70S6K plays a crucial role in HSC proliferation, collagen expression, and cell cycle control, thus representing a potential therapeutic target for liver fibrosis.

Animals↗
PubMed2005

SMAD and p38 MAPK signaling pathways independently regulate alpha1(I) collagen gene expression in unstimulated and transforming growth factor-beta-stimulated hepatic stellate cells.

The hepatic stellate cell (HSC) is the predominant cell type responsible for excess collagen deposition during liver fibrosis. Both transforming growth factor-beta (TGF-beta), the most potent fibrogenic cytokine for HSCs, which classically activates Smad signaling, and p38 MAPK signaling have been shown to influence collagen gene expression; however, the relative contribution and mechanisms that these two signaling pathways have in regulating collagen gene expression have not been investigated. The aim of this study was to investigate the relative roles and mechanisms of both Smad and p38 MAPK signaling in alpha1(I) collagen gene expression in HSCs. Inhibiting either p38 MAPK or Smad signaling reduced alpha1(I) collagen mRNA expression in untreated or TGF-beta-treated HSCs, and when both signaling pathways were simultaneously inhibited, alpha1(I) collagen gene expression was essentially blocked. Both signaling pathways were found to independently and additively increase alpha1(I) collagen gene expression by transcriptional mechanisms. TGF-beta treatment increased alpha1(I) collagen mRNA half-life, mediated by increased stability of alpha1(I) collagen mRNA through p38 MAPK signaling but not through Smad signaling. In conclusion, both p38 MAPK and Smad signaling independently and additively regulate alpha1(I) collagen gene expression by transcriptional activation, whereas p38 MAPK and not Smad signaling increased alpha1(I) collagen mRNA stability.

Adenoviridae↗
PubMed2005

Systemic infusion of angiotensin II exacerbates liver fibrosis in bile duct-ligated rats.

Recent evidence indicates that the renin-angiotensin system (RAS) plays a major role in liver fibrosis. Here, we investigate whether the circulatory RAS, which is frequently activated in patients with chronic liver disease, contributes to fibrosis progression. To test this hypothesis, we increased circulatory angiotensin II (Ang II) levels in rats undergoing biliary fibrosis. Saline or Ang II (25 ng/kg/h) were infused into bile duct-ligated rats for 2 weeks through a subcutaneous pump. Ang II infusion increased serum levels of Ang II and augmented bile duct ligation-induced liver injury, as assessed by elevated liver serum enzymes. Moreover, it increased the hepatic concentration of inflammatory proteins (tumor necrosis factor alpha and interleukin 1beta) and the infiltration of CD43-positive inflammatory cells. Ang II infusion also favored the development of vascular thrombosis and increased the procoagulant activity of tissue factor in the liver. Livers from bile duct-ligated rats infused with Ang II showed increased transforming growth factor beta1 content, collagen deposition, accumulation of smooth muscle alpha-actin-positive cells, and lipid peroxidation products. Moreover, Ang II infusion stimulated phosphorylation of c-Jun and p42/44 mitogen-activated protein kinase and increased proliferation of bile duct cells. In cultured rat hepatic stellate cells (HSCs), Ang II (10(-8) mol/L) increased intracellular calcium and stimulated reactive oxygen species formation, cellular proliferation and secretion of proinflammatory cytokines. Moreover, Ang II stimulated the procoagulant activity of HSCs, a newly described biological function for these cells. In conclusion, increased systemic Ang II augments hepatic fibrosis and promotes inflammation, oxidative stress, and thrombogenic events.

Angiotensin II↗
PubMed2005

Attenuated hepatic inflammation and fibrosis in angiotensin type 1a receptor deficient mice.

BACKGROUND/AIMS: Pharmacological blockade of the renin-angiotensin system (RAS) attenuates liver fibrogenesis in rats. Here, we provide genetic evidence implicating angiotensin type 1 (AT1) receptors in liver fibrogenesis. METHODS: Wild type (WT) and AT1a knockout [AT1a (-/-)] mice were subjected to either sham operation or bile-duct ligation. Fibrosis was assessed by Sirius Red staining and hydroxyproline hepatic content. Fibrogenic and inflammatory cytokines were measured by ELISA. RESULTS: Bile duct ligation-induced elevation of serum liver enzymes was similar in WT and AT1a (-/-) mice. Bile duct ligated WT mice showed inflammatory changes and severe septal fibrosis. In contrast, AT1a (-/-) mice showed minor fibrotic lesions. Collagen accumulation was lower in AT1a (-/-) mice compared to WT mice. The increase in hepatic concentration of TGFbeta1 and pro-inflammatory cytokines was attenuated in AT1a (-/-) mice compared to WT mice. Immunohistochemistry analysis revealed decreased infiltration by inflammatory cells, lipid peroxidation products as well as decreased phosphorylation of c-Jun and p42/44 MAPK in AT1a (-/-) mice compared to AT1 (+/+) mice. CONCLUSIONS: AT1 receptors play an important role in the development of fibrosis. Pharmacological blockade of AT1 receptors appears to be a promising approach to treat liver fibrosis.

Animals↗
PubMed2004

PPAR Gamma and Hepatic Stellate Cells.

Activation of Hepatic stellate cells (HSC) in fibrogenesis involves distinct morphological and biochemical changes. This activation requires the coordinated changes in activity of several transcription factors. Peroxisome proliferator-activated receptor gamma (PPAR gamma) is one such factor whose activity is decreased in activated HSC. PPAR gamma ligands suppress several markers of HSC activation such as expression of collagen and alpha smooth muscle actin (alpha-SMA), cell proliferation and migration. Expression of PPAR gamma, per se, also inhibits HSC activation. These findings support the role of PPAR gamma in reversion of activated HSC toward their quiescent state.

Journal Article↗
PubMed2004

Primary cirrhotic hepatocytes resist TGFbeta-induced apoptosis through a ROS-dependent mechanism.

BACKGROUND/AIMS: The cirrhotic liver manifests dysregulated hepatocyte growth by poor regenerative capacity, formation of regenerative nodules, and malignant transformation to hepatocellular carcinoma. The purpose of this study was to determine if dysregulated hepatocyte growth occurs through deficient apoptosis. METHODS: Hepatocytes were isolated from normal and CCl(4)-treated mice and treated with TGFbeta, TNFalpha, and UV-C, known apoptotic agents. RESULTS: Cirrhotic hepatocytes were less sensitive to TGFbeta- (45+/-5 vs. 15+/-3%; P<0.003), TNFalpha- (59+/-21 vs. 21+/-8%; P=0.02), and UV-C-induced (31+/-4 vs. 17+/-4%; P<0.03) apoptosis compared to normal hepatocytes. In normal hepatocytes, TGFbeta-induced apoptosis occurred through a ROS-, MPT-, and caspase-dependent pathway. Cirrhotic hepatocytes lacked caspase activation, had decreased procaspase-8 expression, failed to undergo the MPT, and had increased basal ROS activity compared to normal hepatocytes. After treatment with trolox, an antioxidant that reduced basal ROS activity, cirrhotic hepatocytes underwent apoptosis in response to TGFbeta treatment. CONCLUSIONS: These findings suggest that increased ROS activity in cirrhotic hepatocytes plays a critical role in mediating cirrhotic hepatocyte resistance to apoptosis. Cirrhotic hepatocyte resistance to TGFbeta-induced apoptosis is ROS-dependent and is a mechanism of dysregulated growth in the chronically inflamed liver.

Animals↗
PubMed2004

TNF alpha-induced hepatocyte apoptosis is associated with alterations of the cell cycle and decreased stem loop binding protein.

BACKGROUND: Inhibition of nuclear factor kappa B (NF kappa B) during liver regeneration induces hepatocyte apoptosis associated with normal DNA synthesis but decreased mitosis, suggesting that inhibition of NF kappa B impairs progression from S-phase through the G(2)/M phase of the cell cycle. Our aim was to determine if inhibition of NF kappa B alters cell cycle characteristics in hepatocytes treated with tumor necrosis factor alpha (TNF alpha). METHODS: Primary hepatocytes from BALB/c mice were infected with adenoviruses expressing luciferase (control; AdLuc) or the I kappa B super-repressor (AdI kappa B) and treated with or without TNF alpha (30 ng/ml). Flow cytometry was performed (0 to 40 hours) to determine apoptosis and cell cycle progression. Reverse transcriptase-polymerase chain reaction and immunoblots assessed changes in cell cycle mediators and antiapoptotic factors. RESULTS: Primary hepatocytes treated with AdI kappa B and TNF alpha demonstrated significantly more S-phase cells (14% +/- 3% vs 6% +/- 2%, P<.05) at 14 hours compared with controls. Inhibition of NF kappa B with or without TNFalpha was associated with decreased expression of stem loop bind protein, a marker of cell cycle progression through S-phase. The NF kappa B-induced antiapoptotic proteins, iNOS and TRAF2, had decreased message at 9 and 12 hours, respectively, in TNF alpha- and AdI kappa B-treated cells. CONCLUSION: Inhibition of NF kappa B in TNF alpha-treated primary mouse hepatocytes is associated with increased S-phase cell cycle retention and decreased stem loop bind protein.

Animals↗
PubMed2003

Peroxisome proliferator-activated receptor gamma induces a phenotypic switch from activated to quiescent hepatic stellate cells.

Depletion of peroxisome proliferator-activated receptor gamma (PPARgamma) accompanies myofibroblastic transdifferentiation of hepatic stellate cells (HSC), the primary cellular event underlying liver fibrogenesis. The treatment of activated HSC in vitro or in vivo with synthetic PPARgamma ligands suppresses the fibrogenic activity of HSC. However, it is uncertain whether PPARgamma is indeed a molecular target of this effect, because the ligands are also known to have receptor-independent actions. To test this question, the present study examined the effects of forced expression of PPARgamma via an adenoviral vector on morphologic and biochemical features of culture-activated HSC. The vector-mediated expression of PPARgamma itself is sufficient to reverse the morphology of activated HSC to the quiescent phenotype with retracted cytoplasm, prominent dendritic processes, reduced stress fibers, and accumulation of retinyl palmitate. These effects are abrogated by concomitant expression of a dominant negative mutant of PPARgamma that prevents transactivation of but not binding to the PPAR response element. PPARgamma expression also inhibits the activation markers such as the expression of alpha-smooth muscle actin, type I collagen, and transforming growth factor beta1; DNA synthesis; and JunD binding to the activator protein-1 (AP-1) site and AP-1 promoter activity. Inhibited JunD activity by PPARgamma is not due to reduced JunD expression or JNK activity or to a competition for p300. But it is due to a JunD-PPARgamma interaction as demonstrated by co-immunoprecipitation and glutathione S-transferase pull-down analysis. Further, the use of deletion constructs reveals that the DNA binding region of PPARgamma is the JunD interaction domain. In summary, our results demonstrate that the restoration of PPARgamma reverses the activated HSC to the quiescent phenotype and suppresses AP-1 activity via a physical interaction between PPARgamma and JunD.

Animals↗
PubMed2003

Prolonged infusion of angiotensin II into normal rats induces stellate cell activation and proinflammatory events in liver.

Recent evidence indicates that angiotensin II (ANG II) plays an important role in liver fibrogenesis. However, the underlying mechanisms are largely unknown. In advanced chronic liver diseases, circulating levels of ANG II are frequently elevated. We investigated the hepatic effects of prolonged systemic infusion of ANG II in normal rats. Saline or ANG II at subpressor and pressor doses (15 and 50 ng.kg-1.min-1, respectively) were infused to normal rats for 4 wk through a subcutaneous osmotic pump. Infusion of ANG II resulted in liver injury, as assessed by elevated serum liver enzymes. Livers from ANG II-perfused rats showed activation of JNK and ERK as well as increased NF-kappaB and activating protein-1 DNA-binding activity. Moreover, ANG II perfusion induced oxidative stress, increased concentration of proinflammatory cytokines, and upregulated the inflammatory proteins inducible nitric oxide synthase and cyclooxygenase-2. Histological examination of the livers from ANG II-infused rats showed mild portal inflammation as well as thickening and thrombosis of small hepatic vessels. ANG II-treated livers showed accumulation of CD43-positive inflammatory cells and activated hepatic stellate cells (HSCs) at the pericentral areas. A slight increase in collagen synthesis was observed, as assessed by Sirius red staining and hepatic hydroxyproline. All of these effects were observed when ANG II was perfused at subpressor and pressor doses. ANG II also accelerated the activation of primary cultured rat HSCs. In conclusion, increased systemic ANG II can induce liver injury by promoting proinflammatory events and vascular damage. ANG II-induced hepatic effects are not dependent on increase in arterial pressure.

Angiotensin II↗
PubMed2003

Liver fibrosis: signals leading to the amplification of the fibrogenic hepatic stellate cell.

Liver fibrosis represents a major medical problem with significant morbidity and mortality. Worldwide hepatitis viral infections represent the major cause liver fibrosis; however, within the United States chronic ethanol consumption is the leading cause of hepatic fibrosis. Other known stimuli for liver fibrosis include helminthic infection, iron or copper overload and biliary obstruction. Fibrosis can be classified as a wound healing response to a variety of chronic stimuli that is characterized by an excessive deposition of extracellular matrix proteins of which type I collagen predominates. This excess deposition of extracellular matrix proteins disrupts the normal architecture of the liver resulting in pathophysiological damage to the organ. If left untreated fibrosis can progress to liver cirrhosis ultimately leading to organ failure and death if left untreated. This review will discuss the molecular events leading to liver fibrosis. The discussion will include collagen gene regulation and proliferative signals that contribute to the amplification of the hepatic stellate cell, the primary fibrogenic cell type that resides in the liver.

Animals↗
PubMed2003

Activated stellate cells express the TRAIL receptor-2/death receptor-5 and undergo TRAIL-mediated apoptosis.

Apoptosis has emerged as an important mechanism to reduce numbers of activated stellate cells during the resolution phase of hepatic fibrosis. These observations suggest that activated stellate cells may be more susceptible to apoptotic stimuli than their quiescent counterparts. Because other activated cell types are more sensitive than their quiescent phenotypes to apoptosis by tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), we examined the expression of TRAIL death receptors (DRs) and susceptibility to TRAIL cytotoxicity in stellate cells undergoing progressive activation. A spontaneously immortalized human stellate cell line, LX-2, was analyzed during 14 days of progressive activation following plating, during which time alpha-smooth muscle actin (alpha-SMA) and a beta-crystallin (markers of stellate cell activation) messenger RNA (mRNA) increased 7-fold and 5-fold, respectively. During this same interval, TRAIL-R1/DR4 and TRAIL-R2/DR5 mRNA expression increased 18-fold and 17.6-fold, although TRAIL-R2/DR5 expression was quantitatively 103-fold greater than TRAIL-R1/DR4; parallel changes occurred in TRAIL/DR5 protein expression and cellular susceptibility to TRAIL-mediated apoptosis. Similar findings were observed in primary murine stellate cells undergoing activation on a plastic surface. In conclusion, stellate cells show activation-dependent TRAIL-R2/DR5 expression and TRAIL-mediated apoptosis. Because TRAIL-R2/DR5 is not expressed by hepatocytes, TRAIL/DR5 agonists may be useful in reducing fibrosis by inducing stellate cell apoptosis.

Animals↗
PubMed2003

DNase I-hypersensitive sites enhance alpha1(I) collagen gene expression in hepatic stellate cells.

Liver fibrosis is characterized by a dramatic increase in the expression of type I collagen. Several deoxyribonuclease (DNase) I-hypersensitive sites (HS) have been located in the distal 5'-flanking region of the alpha1(I) collagen gene that are specific to collagen-producing cells. To assess the role of the DNase I-HS in regulating alpha1(I) collagen gene expression in hepatic stellate cells (HSCs), 3 transgenic mouse lines expressing collagen-alpha1(I) reporter genes were used (Krempen et al. Gene Expr 1999;8:151-163). The pCol9GFP transgene contains the collagen gene promoter (-3122 to +111) linked to the green fluorescent protein (GFP) reporter gene. The pCol9GFP-HS4,5 transgene contains HS4,5 and pColGFP-HS8,9 contains HS8,9 positioned upstream of the collagen promoter in pCol9GFP. HSCs isolated from transgenic mice containing pCol9GFPHS4,5 and pColGFP-HS8,9 showed earlier and higher GFP expression patterns than HSCs isolated from pCol9GFP mice. HSCs from pCol9GFP-HS4,5 showed the highest levels of GFP expression and culture-induced expression correlated with induction of the endogenous alpha1(I) collagen gene. After CCl(4) administration, pCol9GFP-HS4,5 mice showed increased GFP expression compared with pCol9GFP mice in both whole liver extracts and isolated HSCs. Several sites for DNA-protein interactions in both HS4 and HS5 were identified that included a binding site for activator protein 1. In conclusion, DNase I-HS4,5 enhance expression of the alpha1(I) collagen gene promoter in HSCs both in vitro and in vivo after a fibrogenic stimulus. The collagen-GFP transgenic mice provide a convenient and reliable model system to investigate the molecular mechanisms controlling increased collagen expression during fibrosis.

Animals↗
PubMed2002

The role of focal adhesion kinase-phosphatidylinositol 3-kinase-akt signaling in hepatic stellate cell proliferation and type I collagen expression.

Following a fibrogenic stimulus, the hepatic stellate cell (HSC) undergoes a complex activation process associated with increased cell proliferation and excess deposition of type I collagen. The focal adhesion kinase (FAK)-phosphatidylinositol 3-kinase (PI3K)-Akt signaling pathway is activated by platelet-derived growth factor (PDGF) in several cell types. We investigated the role of the FAK-PI3K-Akt pathway in HSC activation. Inhibition of FAK activity blocked HSC migration, cell attachment, and PDGF-induced PI3K and Akt activation. Both serum- and PDGF-induced Akt phosphorylation was inhibited by LY294002, an inhibitor of PI3K. A constitutively active form of Akt stimulated HSC proliferation in serum-starved HSCs, whereas LY294002 and dominant-negative forms of Akt and FAK inhibited PDGF-induced proliferation. Transforming growth factor-beta, an inhibitor of HSC proliferation, did not block PDGF-induced Akt phosphorylation, suggesting that transforming growth factor-beta mediates its antiproliferative effect downstream of Akt. Expression of type I collagen protein and alpha1(I) collagen mRNA was increased by Akt activation and inhibited when PI3K activity was blocked. Therefore, FAK is important for HSC migration, cell attachment, and PDGF-induced cell proliferation. PI3K is positioned downstream of FAK. Signals for HSC proliferation are transduced through FAK, PI3K, and Akt. Finally, expression of type I collagen is regulated by the PI3K-Akt signaling pathway.

Animals↗
PubMed2002

Cholestasis induces murine hepatocyte apoptosis and DNA synthesis with preservation of the immediate-early gene response.

BACKGROUND: Major hepatic resection in patients with unrelieved obstructive jaundice carries an increased risk of postoperative liver failure. We hypothesized that cholestasis induces hepatocyte apoptosis and impairs hepatic regeneration by inhibiting up-regulation of the known immediate-early response genes, nuclear factor kappa B (NF-kappaB) and activating protein-1 (AP-1). The aim of this study was to determine whether the immediate-early gene response in hepatic regeneration remains intact in extrahepatic cholestasis. METHODS: Eight-week-old BALB/c mice underwent either sham operation (SO) or common bile duct ligation (BDL). Two-thirds partial hepatectomy (PH) was performed at 4 and 7 days, with remnant liver harvested 0, 15, 30, or 60 minutes after PH. Serum analysis for markers of cholestasis and histopathology was obtained. Proliferating cell nuclear antigen and terminal deoxyuridine triphosphate nick end labeling (TUNEL) immunohistochemistry for detection of DNA synthesis and apoptosis, respectively, was performed 4, 7, or 10 days after SO or BDL. Liver samples from 0, 15, 30, or 60 minutes after PH were analyzed for NF-kappaB and AP-1 DNA binding activity by using electrophoretic mobility shift assays. RESULTS: Increased serum bilirubin level and hematoxylin-eosin-stained liver sections confirmed cholestasis in BDL mice. BDL induced marked DNA synthesis and hepatocyte apoptosis in prehepatectomy liver at both 4 and 7 days. Substantially higher basal levels of both NF-kappaB and AP-1 binding activity were present in BDL compared with SO mice. Fold induction of NF-kappaB and AP-1, however, was similar between BDL and SO mice. Cholestasis induced hepatocyte DNA synthesis and apoptosis. Basal NF-kappaB and AP-1 DNA binding activity was increased in BDL mice, but fold induction of these immediate-early genes did not differ from controls. CONCLUSIONS: Although basal NF-kappaB and AP-1 DNA binding is increased in cholestasis, the immediate-early gene response to PH remains intact in BDL mice.

Animals↗
PubMed2002

Transforming growth factor-beta1 induces hepatocyte apoptosis by a c-Jun independent mechanism.

BACKGROUND: During hepatic regeneration, transforming growth factor (TGF)-beta1 messenger RNA increases after the initial cycle of DNA synthesis, and it may control hepatocyte growth by inducing apoptosis. TGF-beta1 also induces c-Jun, a potential proapoptotic transcription factor. We hypothesized that autocrine expression of activated TGF-beta1 (Ad5aTGF-beta1) would increase c-jun expression in rat liver and limit hepatic regeneration by inducing apoptosis. METHODS: Male rats (175 to 200 g) received portal venous injections with adenoviruses expressing either luciferase (Ad5Luc), as a control, or Ad5aTGF-beta1 at a dose of 6 x 10(9) plaque-forming units. Livers were harvested 24 or 48 hours after injection and nuclear extracts and total RNA isolated. TGF-beta1 expression was confirmed by Northern blot analysis in all TGF-beta1-injected rats. RESULTS: A 2.5-fold increase in c-jun mRNA expression was detected in Ad5aTGF-beta1-infected rats compared with control rats. Transcriptional activity was assessed with an AP-1-responsive-reporter gene that increased 3-fold in rat primary hepatocytes infected with Ad5aTGF-beta1. C-Jun N-terminal kinase activity also increased 6- to 7-fold in Ad5aTGF-beta1-treated rats 24 and 48 hours after injection. Ad5aTGF-beta1-injected rats demonstrated increased AP-1 binding activity compared with Ad5Luc rats. Hepatocytes infected in vitro with Ad5aTGF-beta1 demonstrated increased apoptosis compared with Ad5Luc-infected hepatocytes (47% vs 27%) 36 hours after infection. Dual adenoviral infection with Ad5aTGF-beta1 and a dominant-negative c-Jun (Ad5TAM67) decreased AP-1-induced Ad5Luc activity but not hepatocyte apoptosis (46% with dominant-negative c-Jun and 47% without). CONCLUSIONS: These data demonstrate that TGF-beta1 induces c-Jun, but c-Jun is not proapoptotic in hepatocytes.

Adenoviridae↗
PubMed2002

IL-10 receptor and coreceptor expression in quiescent and activated hepatic stellate cells.

Interleukin (IL)-10 expression is induced in activated hepatic stellate cells (HSC) in vitro and in vivo. We analyzed expression of IL-10 receptor (IL-10R) and coreceptor cytokine receptor family (CRF2-4) in HSC. We aimed to clone and sequence partial cDNA for rat IL-10R and CRF2-4, determine their expression in activated rat HSC in vivo and in vitro, and examine the biological responsiveness of HSC to exogenous IL-10. PCR cloning and sequencing of partial rat IL-10R and CRF2-4 cDNAs revealed 86% homology with corresponding mouse sequences. In hepatic macrophages, Northern blot with cloned IL-10R cDNA detected an expected 3.5-kb transcript, and IL-10R and CRF2-4 mRNAs showed steady constitutive expression after in vitro lipopolysaccharide treatment or cholestatic liver injury. IL-10R mRNA expression, as confirmed by immunohistochemistry, was induced 20.1- and 8.6-fold in HSC from cholestatic livers and 7-day culture-activated HSC, respectively but CRF2-4 mRNA levels were unchanged. Under serum-free conditions, IL-10 had minimal effects on collagen production but reduced DNA synthesis, matrix metalloprotease-2 mRNA levels, and activity in HSC. With serum, IL-10 inhibited both collagen production and DNA synthesis but had no effect on procollagen-alpha(1)(I) mRNA levels. This shows concomitant induction of IL-10R but not CRF2-4 to that of IL-10 by activated HSC in vitro and in vivo and associated acquisition of the responsiveness to IL-10, entailing complex effects on HSC.

Animals↗