pinecone/core-2020-05-10-deduplication huggingface dataset:
Cathepsins B and D drive hepatic stellate cell proliferation and promote their fibrogenic potential

El pdf del artículo es el manuscrito de autor.-- PubMed: PMCID:PMC2670444Cathepsins have been best characterized in tumorigenesis and cell death and implicated in liver ﬁbrosis; however, whether cathepsins directly regulate hepatic stellate cell (HSC) activation and proliferation, hence modulating their ﬁbrogenic potential, is largely unknown. Here, we show that expression of cathepsin B (CtsB) and cathepsin D (CtsD) is negligible in quiescent HSCs but parallels the increase of -smooth muscle actin and transforming growth factor- during in vitro mouse HSC activation. Both cathepsins are necessary for HSC transdifferentiation into myoﬁbroblasts, because their silencing or inhibition decreasedHSC proliferation and the expression of phenotypicmarkers ofHSC activation, with similar results observed with the human HSC cell line LX2. CtsB inhibition blunted AKT phosphorylation in activated HSCs in response to platelet-derived growth factor.Moreover, during in vivo liver ﬁbrogenesis caused by CCl4 administration, CtsB expression increased in HSCs but not in hepatocytes, and its inactivation mitigated CCl4-induced inﬂammation, HSC activation, and collagen deposition. Conclusion: These ﬁndings support a critical role for cathepsins inHSC activation, suggesting that the antagonismof cathepsins inHSCsmay be of relevance for the treatment of liver ﬁbrosis.Financial support: The work was supported by CIBEREHD and grant PI070193 (Instituto de Salud Carlos III); by grant SAF2006-06780 (Plan Nacional de I+D), Spain; and by grant P50-AA-11999 (Research Center for Liver and\ud
Pancreatic Diseases, US National Institute on Alcohol Abuse and Alcoholism).Peer reviewe
ABC-transporters are localized in caveolin-1-positive and reggie-1-negative and reggie-2-negative microdomains of the canalicular membrane in rat hepatocytes

The canalicular plasma membrane is constantly exposed to bile acids acting as detergents. Bile acids are essential to mediate release of biliary lipids from the canalicular membrane. Membrane microdomains (previously called lipid rafts) are biochemically defined by their resistance to detergent solubilization at cold temperature. We aimed to investigate the canalicular plasma membrane for the presence of microdomains, which could protect this membrane against the detergent action of bile acids. Highly purified rat liver canalicular plasma membrane vesicles were extracted with 1% Triton X-100 or 1% Lubrol WX at 4 degrees C and subjected to flotation through sucrose step gradients. Both detergents yielded detergent-resistant membranes containing the microdomain markers alkaline phosphatase and sphingomyelin. However, cholesterol was resistant to Lubrol WX solubilization, whereas it was only marginally resistant to solubilization by Triton X-100. The microdomain marker caveolin-1 was localized to the canalicular plasma membrane domain and was resistant to Lubrol WX, but to a large extent solubilized by Triton X-100. The two additional microdomain markers, reggie-1 and reggie-2, were localized to the basolateral and canalicular plasma membrane and were partially resistant to Lubrol WX but resistant to Triton X-100. The canalicular transporters bile salt export pump, multidrug resistance protein 2, multidrug resistance-associated protein 2, and Abcg5 were largely resistant to Lubrol WX but were solubilized by Triton X-100. Conclusion: These results indicate the presence of two different types of microdomains in the canalicular plasma membrane: "Lubrol-microdomains" and "Triton-microdomains". "Lubrol-microdomains" contain the machinery for canalicular bile formation and may be the starting place for canalicular lipid secretion
Golgi phosphoprotein 2 (GOLPH2) expression in liver tumors and its value as a serum marker in hepatocellular carcinomas

Hepatocellular carcinomas (HCCs) and bile duct carcinomas (BDCs) have a poor prognosis. Therefore, surveillance strategies including sensitive and specific serum markers for early detection are needed. Recently, Golgi Phosphoprotein 2 (GOLPH2) has been proposed as a serum marker for HCC, but GOLPH2 expression data in liver tissues was not available. Using tissue microarrays and immunohistochemistry, we semiquantitatively analyzed GOLPH2 protein expression in patients with HCC (n = 170), benign liver tumors (n = 22), BDC (n = 114) and normal liver tissue (n = 105). A newly designed sandwich enzyme-linked immunoassay (ELISA) was used to analyze GOLPH2 levels in the sera of patients with HCC (n = 62), hepatitis C virus (HCV) (n = 29), BDC (n = 10), and healthy control persons (n = 12). By immunohistochemistry 121/170 (71%) of HCC showed strong GOLPH2 expression, which was significantly associated with a higher tumor grade (P = 0.01). A total of 97/114 (85%) BDCs showed a strong GOLPH2 expression which proved to be an independent prognostic factor for overall survival (P < 0.05). Serum levels of GOLPH2 measured by ELISA were significantly elevated in patients with HCC with underlying HCV infection (median 18 mg/L, P < 0.05) and patients with BDC (median = 14.5 mg/L, P < 0.01) in comparison to healthy controls (median 4 mg/L). Conclusion: GOLPH2 protein is highly expressed in tissues of HCC and BDC. GOLPH2 protein levels are detectable and quantifiable in sera by ELISA. In patients with hepatitis C, serial ELISA measurements in the course of the disease appear to be a promising complementary serum marker in the surveillance of HCC. GOLPH2 should be further evaluated as a serum tumor marker in BDC on a larger scale
S6K1 deficiency protects against apoptosis in hepatocytes

PMCID: PMC2752632The mammalian target of rapamycin (mTOR)/S6K1 signaling pathway controls cell growth and proliferation. To assess the importance of S6K1 in the balance between death and survival in the liver, we have generated immortalized hepatocyte cell lines from wild-type and S6K1-deficient (S6K1(-/-)) mice. In S6K1(-/-) hepatocytes, caspase-8 and the pro-apoptotic protein Bid were constitutively down-regulated as compared with wild-type. Moreover, S6K1(-/-) hepatocytes failed to respond to the apoptotic trigger of death receptor activation. Neither caspase-8 activation nor FLIP(L) degradation in response to tumor necrosis factor alpha (TNF-α) or anti-Fas antibody (Jo2) was observed in cells lacking S6K1. Downstream events such as Bid cleavage, cytochrome C release, caspase-3 activation, DNA laddering, as well as the percentage of apoptotic cells were attenuated as compared with wild-type. In addition, the anti-apoptotic protein Bclx(L) was down-regulated in TNF-α-treated or Jo2-treated wild-type hepatocytes, but this response was abolished in S6K1(-/-) cells. In vivo, S6K1-deficient mice were protected against concanavalin A-induced apoptosis. The withdrawal of growth factors strongly induced apoptosis in wild-type, but not in S6K1(-/-) hepatocytes. S6K1 deficiency did not decrease Bclx(L)/Bim ratio on serum withdrawal, thereby protecting cells from cytochrome C release and DNA fragmentation. At the molecular level, the lack of S6K1-mediated negative feedback decreased insulin receptor substrate-1 (IRS-1) serine phosphorylation, resulting in activation of survival pathways mediated by phosphatidylinositol 3-kinase/Akt and extracellular signal-regulated kinase (ERK). However, S6K1(-/-) hepatocytes underwent apoptosis on serum withdrawal in combination with phosphatidylinositol 3-kinase (PI3K) or ERK inhibitors.[Conclusion] This finding might explain the mechanism of resistance to mTOR inhibitors in cancer treatments and strongly suggests that the inhibition of S6K1 could protect against acute liver failure and, in combination with inhibitors that abrogate the sustained activation of Akt and ERK, could improve the efficacy of hepatocarcinoma (HCC) treatment.Supported by grants BFU 2008-02420 and CIT-090100-2007-35 (MEC, Spain) and Centro de Investigación Biomédica en Red de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM), Instituto Carlos III, MSC, (Spain). S.C.K. and V.Z. are supported by NIH grants CA120475 and DK073802.Peer reviewe
Microenvironmental regulation of the sinusoidal endothelial cell phenotype in vitro

Liver sinusoidal endothelial cells (LSECs) differ, both structurally and functionally, from endothelial cells (ECs) lining blood vessels of other tissues. For example, in contrast to other ECs, LSECs possess fenestrations, have low detectable levels of platelet endothelial cell adhesion molecule 1 expression, and in rat tissue, they distinctively express a cell surface marker recognized by the SE-1 antibody. These unique phenotypic characteristics seen in hepatic tissue are lost over time upon culture in vitro; therefore, this study sought to systematically examine the effects of microenvironmental stimuli—namely, extracellular matrix and neighboring cells, on the LSEC phenotype in vitro. In probing the role of the underlying extracellular matrix, we identified collagen I and collagen III as well as mixtures of collagen I/collagen IV/fibronectin as having a positive effect on LSEC survival. Furthermore, using a stable hepatocellular model (hepatocyte–fibroblast) we were able to prolong the expression of both SE-1 and phenotypic functions of LSEC such as factor VIII activity and AcLOL uptake in cocultured LSECs through the production of short-range paracrine signals. In the course of these experiments, we identified the antigen recognized by SE-1 as CD32b. Conclusion: Collectively, this study has identified several microenvironmental regulators of liver sinusoidal endothelial cells that prolong their phenotypic functions for up to 2 weeks in culture, enabling the development of better in vitro models of liver physiology and disease.Author Manuscript: 2010 June 23
Increased tumor necrosis factor alpha-converting enzyme activity induces insulin resistance and hepatosteatosis in mice

Tumor necrosis factor alpha-converting enzyme (TACE, also known as ADAM17) was recently involved in the pathogenesis of insulin resistance. We observed that TACE activity was significantly higher in livers of mice fed a high-fat diet (HFD) for 1 month, and this activity was increased in liver > white adipose tissue > muscle after 5 months compared with chow control. In mouse hepatocytes, C(2)C(12) myocytes, and 3T3F442A adipocytes, TACE activity was triggered by palmitic acid, lipolysaccharide, high glucose, and high insulin. TACE overexpression significantly impaired insulin-dependent phosphorylation of AKT, GSK3, and FoxO1 in mouse hepatocytes. To test the role of TACE activation in vivo, we used tissue inhibitor of metalloproteinase 3 (Timp3) null mice, because Timp3 is the specific inhibitor of TACE and Timp3(-/-) mice have higher TACE activity compared with wild-type (WT) mice. Timp3(-/-) mice fed a HFD for 5 months are glucose-intolerant and insulin-resistant; they showed macrovesicular steatosis and ballooning degeneration compared with WT mice, which presented only microvesicular steatosis. Shotgun proteomics analysis revealed that Timp3(-/-) liver showed a significant differential expression of 38 proteins, including lower levels of adenosine kinase, methionine adenosysltransferase I/III, and glycine N-methyltransferase and higher levels of liver fatty acid-binding protein 1. These changes in protein levels were also observed in hepatocytes infected with adenovirus encoding TACE. All these proteins play a role in fatty acid uptake, triglyceride synthesis, and methionine metabolism, providing a molecular explanation for the increased hepatosteatosis observed in Timp3(-/-) compared with WT mice. Conclusion: We have identified novel mechanisms, governed by the TACE-Timp3 interaction, involved in the determination of insulin resistance and liver steatosis during overfeeding in mice
Ghrelin attenuates hepatocellular injury and liver fibrogenesis in rodents and influences fibrosis progression in humans

El pdf del artículo es la versión pre-print.-- et al-There are no effective antifibrotic therapies for patients with liver diseases. We performed an experimental and translational study to investigate whether ghrelin, an orexigenic hormone with pleiotropic properties, modulates liver fibrogenesis. Recombinant ghrelin was administered to rats with chronic (bile duct ligation) and acute (carbon tetrachloride) liver injury. Hepatic gene expression was analyzed by way of microarray analysis and quantitative polymerase chain reaction. The hepatic response to chronic injury was also evaluated in wild-type and ghrelin-deficient mice. Primary human hepatic stellate cells were used to study the effects of ghrelin in vitro. Ghrelin hepatic gene expression and serum levels were assessed in patients with chronic liver diseases. Ghrelin gene polymorphisms were analyzed in patients with chronic hepatitis C. Recombinant ghrelin treatment reduced the fibrogenic response, decreased liver injury and myofibroblast accumulation, and attenuated the altered gene expression profile in bile duct-ligated rats. Moreover, ghrelin reduced the fibrogenic properties of hepatic stellate cells. Ghrelin also protected rats from acute liver injury and reduced the extent of oxidative stress and inflammation. Ghrelin-deficient mice developed exacerbated hepatic fibrosis and liver damage after chronic injury. In patients with chronic liver diseases, ghrelin serum levels decreased in those with advanced fibrosis, and ghrelin gene hepatic expression correlated with expression of fibrogenic genes. In patients with chronic hepatitis C, polymorphisms of the ghrelin gene (994CT and 604GA) influenced the progression of liver fibrosis. Conclusion: Ghrelin exerts antifibrotic effects in the liver and may represent a novel antifibrotic therapy. Copyright © 2010 by the American Association for the Study of Liver Diseases.Supported by grants from the Ministerio de Ciencia e Investigación (SAF2005-06245), from the Instituto de Salud Carlos III (FIS2005-050567, FIS 2008-PI08/0237 and PI070497), and from the European Community FP6 (LSHB-CT-2007-036644 - DIALOK) and by fellowships from Institut d’Investigacions Biomèdiques August Pi i Sunyer (to M. M. and M. D.), the Fundación Bilbao Vizcaya Argentaria (to M. D.) and the Fundació Clínic (to P. S. B.).Peer Reviewe
Inhibition of hepatitis C virus infection by anti-claudin-1 antibodies is mediated by neutralization of E2-CD81-claudin-1 associations.

International audienceThe tight junction protein claudin-1 (CLDN1) has been shown to be essential for hepatitis C virus (HCV) entry-the first step of viral infection. Due to the lack of neutralizing anti-CLDN1 antibodies, the role of CLDN1 in the viral entry process is poorly understood. In this study, we produced antibodies directed against the human CLDN1 extracellular loops by genetic immunization and used these antibodies to investigate the mechanistic role of CLDN1 for HCV entry in an infectious HCV cell culture system and human hepatocytes. Antibodies specific for cell surface-expressed CLDN1 specifically inhibit HCV infection in a dose-dependent manner. Antibodies specific for CLDN1, scavenger receptor B1, and CD81 show an additive neutralizing capacity compared with either agent used alone. Kinetic studies with anti-CLDN1 and anti-CD81 antibodies demonstrate that HCV interactions with both entry factors occur at a similar time in the internalization process. Anti-CLDN1 antibodies inhibit the binding of envelope glycoprotein E2 to HCV permissive cell lines in the absence of detectable CLDN1-E2 interaction. Using fluorescent-labeled entry factors and fluorescence resonance energy transfer methodology, we demonstrate that anti-CLDN1 antibodies inhibit CD81-CLDN1 association. In contrast, CLDN1-CLDN1 and CD81-CD81 associations were not modulated. Taken together, our results demonstrate that antibodies targeting CLDN1 neutralize HCV infectivity by reducing E2 association with the cell surface and disrupting CD81-CLDN1 interactions. Conclusion: These results further define the function of CLDN1 in the HCV entry process and highlight new antiviral strategies targeting E2-CD81-CLDN1 interactions
A meta-analysis of survival rates of untreated patients in randomized clinical trials of hepatocellular carcinoma.

Knowing the spontaneous outcome of hepatocellular carcinoma (HCC) is important for designing randomized controlled trials (RCTs) of new therapeutic approaches; however, survival of patients in the absence of treatment is highly variable, and prognostic factors influencing outcomes are incompletely defined. The aims of this meta-analysis were to estimate the 1-year and 2-year survival rates of untreated HCC patients enrolled in RCTs of palliative treatments, and to identify prognostic factors. RCTs evaluating therapies for HCC with placebo or no-treatment arms were identified on MEDLINE through April 2009. Data were combined in a random effect model. Primary outcomes were 1-year and 2-year survival. Thirty studies met the inclusion criteria. The pooled estimates of the survival rates were 17.5% at 1 year (95% confidence interval [95%CI], 11%-27%; range, 0%-75%) and 7.3% at 2 years (95%CI, 3.9%-13%; range, 0%-50%). Heterogeneity among studies was highly significant (P < 0.0001) both for 1-year and 2-year survival, and persisted when RCTs were stratified according to all patient and study features. Through meta-regression, impaired performance status, Child-Pugh B-C class, and presence of portal vein thrombosis were all independently associated with shorter survival. Ascites was strongly linked to a worse outcome in intermediate/advanced Barcelona Clinic Liver Cancer stages. Conclusion: This meta-analysis confirms the heterogeneity of behavior of untreated HCC and provides a sound basis for stratifying patients with HCC according to expected survival in future trials of new anti-cancer agents
Themes of liver transplantation

Liver transplantation was the product of five interlocking themes. These began in 1958-1959 with canine studies of then theoretical hepatotrophic molecules in portal venous blood (Theme I) and with the contemporaneous parallel development of liver and multivisceral transplant models (Theme II). Further Theme I investigations showed that insulin was the principal, although not the only, portal hepatotrophic factor. In addition to resolving long-standing controversies about the pathophysiology of portacaval shunt, the hepatotrophic studies blazed new trails in the regulation of liver size, function, and regeneration. They also targeted inborn metabolic errors (e.g., familial hyperlipoproteinemia) whose palliation by portal diversion presaged definitive correction with liver replacement. Clinical use of the Theme II transplant models depended on multiple drug immunosuppression (Theme III, Immunology), guided by an empirical algorithm of pattern recognition and therapeutic response. Successful liver replacement was first accomplished in 1967 with azathioprine, prednisone, and antilymphoid globulin. With this regimen, the world's longest surviving liver recipient is now 40 years postoperative. Incremental improvements in survival outcome occurred (Theme IV) when azathioprine was replaced by cyclosporine (1979), which was replaced in turn by tacrolimus (1989). However, the biologic meaning of alloengraftment remained enigmatic until multilineage donor leukocyte microchimerism was discovered in 1992 in long-surviving organ recipients. Seminal mechanisms were then identified (clonal exhaustion-deletion and immune ignorance) that linked organ engraftment and the acquired tolerance of bone marrow transplantation and eventually clarified the relationship of transplantation immunology to the immunology of infections, neoplasms, and autoimmune disorders. With this insight, better strategies of immunosuppression have evolved. As liver and other kinds of organ transplantation became accepted as healthcare standards, the ethical, legal, equity, and the other humanism issues of Theme V have been resolved less conclusively than the medical-scientific problems of Themes I-IV. Copyright © 2010 by the American Association for the Study of Liver Diseases
Fatty liver and fibrosis in glycine N-methyltransferase knockout mice is prevented by nicotinamide

Deletion of glycine N-methyltransferase (GNMT) in mice, the main gene involved in liver S-adenosylmethionine (SAMe) catabolism, leads to the hepatic accumulation of this molecule and the development of fatty liver and fibrosis. To demonstrate that the excess of hepatic SAMe is the main agent contributing to liver disease in GNMT-KO mice, we treated 1.5-month old GNMT-KO mice for 6 weeks with nicotinamide (NAM), a substrate of the enzyme NAM N-methyltransferase. NAM administration markedly reduced hepatic SAMe content, prevented DNA-hypermethylation and normalized the expression of critical genes involved in fatty acid metabolism, oxidative stress, inflammation, cell proliferation, and apoptosis. More important, NAM treatment prevented the development of fatty liver and fibrosis in GNMT-KO mice. Because GNMT expression is down-regulated in patients with cirrhosis and there are subjects with GNMT mutations who have spontaneous liver disease, the clinical implication of the present findings is obvious at least with respect to these latter individuals. Especially since NAM has been used for many years to treat a broad spectrum of diseases including pellagra and diabetes without significant side effects, it should be considered in subjects with GNMT mutations.ConclusionsThese results indicate that the anomalous accumulation of SAMe in GNMT-KO mice can be corrected by NAM treatment leading to the normalization of the expression of many genes involved in fatty acid metabolism, oxidative stress, inflammation, cell proliferation and apoptosis, and to the reversion of the appearance of the pathologic phenotype
Transforming Growth Factor-Beta Induces Senescence in Hepatocellular Carcinoma Cells and Inhibits Tumor Growth

Cataloged from PDF version of article.Senescence induction could be used as an effective treatment for hepatocellular carcinoma (HCC). However, major senescence inducers (p53 and p16Ink4a) are frequently inactivated in these cancers.We tested whether transforming growth factor-β (TGF-β) could serve as a potential senescence inducer in HCC. First, we screened for HCC cell lines with intact TGF-β signaling that leads to small mothers against decapentaplegic (Smad)-targeted gene activation. Five cell lines met this condition, and all of them displayed a strong senescence response to TGF-β1 (1-5 ng/mL) treatment. Upon treatment, c-myc was down-regulated, p21Cip1 and p15Ink4b were up-regulated, and cells were arrested at G1. The expression of p16Ink4a was not induced, and the senescence response was independent of p53 status. A short exposure of less than 1 minute was sufficient for a robust senescence response. Forced expression of p21 Cip1 and p15Ink4b recapitulated TGF-β1 effects. Senescence response was associated with reduced nicotinamide adenine dinucleotide phosphate oxidase 4 (Nox4) induction and intracellular reactive oxygen species (ROS) accumulation. The treatment of cells with the ROS scavenger N-acetyl-L-cysteine, or silencing of the NOX4 gene, rescued p21Cip1 and p15Ink4b accumulation as well as the growth arrest in response to TGF-β. Human HCC tumors raised in immunodeficient mice also displayed TGF-β1-induced senescence. More importantly, peritumoral injection of TGF-β1 (2 ng) at 4-day intervals reduced tumor growth by more than 75%. In contrast, the deletion of TGF-β receptor 2 abolished in vitro senescence response and greatly accelerated in vivo tumor growth. Conclusion: TGF-β induces p53-independent and p16Ink4a-independent, but Nox4-dependent, p21Cip1-dependent, p15Ink4b-dependent, and ROS-dependent senescence arrest in well-differentiated HCC cells. Moreover, TGF-β-induced senescence in vivo is associated with a strong antitumor response against HCC. Copyright © 2010 by the American Association for the Study of Liver Diseases