heparitin-sulfate has been researched along with Carcinoma--Hepatocellular* in 36 studies
2 review(s) available for heparitin-sulfate and Carcinoma--Hepatocellular
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Heparanase-1: From Cancer Biology to a Future Antiviral Target.
Heparan sulfate proteoglycans (HSPGs) are a major constituent of the extracellular matrix (ECM) and are found to be implicated in viral infections, where they play a role in both cell entry and release for many viruses. The enzyme heparanase-1 is the only known endo-beta-D-glucuronidase capable of degrading heparan sulphate (HS) chains of HSPGs and is thus important for regulating ECM homeostasis. Heparanase-1 expression is tightly regulated as the uncontrolled cleavage of HS may result in abnormal cell activation and significant tissue damage. The overexpression of heparanase-1 correlates with pathological scenarios and is observed in different human malignancies, such as lymphoma, breast, colon, lung, and hepatocellular carcinomas. Interestingly, heparanase-1 has also been documented to be involved in numerous viral infections, e.g., HSV-1, HPV, DENV. Moreover, very recent reports have demonstrated a role of heparanase-1 in HCV and SARS-CoV-2 infections. Due to the undenied pro-carcinogenic role of heparanase-1, multiple inhibitors have been developed, some reaching phase II and III in clinical studies. However, the use of heparanase inhibitors as antivirals has not yet been proposed. If it can be assumed that heparanase-1 is implicated in numerous viral life cycles, its inhibition by specific heparanase-acting compounds should result in a blockage of viral infection. This review addresses the perspectives of using heparanase inhibitors, not only for cancer treatment, but also as antivirals. Eventually, the development of a novel class antivirals targeting a cellular protein could help to alleviate the resistance problems seen with some current antiretroviral therapies. Topics: Antiviral Agents; Biology; Carcinoma, Hepatocellular; COVID-19; Glucuronidase; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Humans; Liver Neoplasms; SARS-CoV-2 | 2023 |
PI-88 and novel heparan sulfate mimetics inhibit angiogenesis.
The heparan sulfate (HS) mimetic PI-88 is a promising inhibitor of tumor growth and metastasis expected to commence phase III clinical evaluation in 2007 as an adjuvant therapy for postresection hepatocellular carcinoma. Its anticancer properties are attributed to inhibition of angiogenesis via antagonism of the interactions of angiogenic growth factors and their receptors with HS. It is also a potent inhibitor of heparanase, an enzyme that plays a key role in both metastasis and angiogenesis. A series of PI-88 analogs have been prepared with enhanced chemical and biological properties. The new compounds consist of single, defined oligosaccharides with specific modifications designed to improve their pharmacokinetic properties. These analogs all inhibit heparanase and bind to the angiogenic fibroblast growth factor 1 (FGF-1), FGF-2, and vascular endothelial growth factor with similar affinity to PI-88. However, compared with PI-88, some of the newly designed compounds are more potent inhibitors of growth factor-induced endothelial cell proliferation and of endothelial tube formation on Matrigel. Representative compounds were also tested for antiangiogenic activity in vivo and were found to reduce significantly blood vessel formation. Moreover, the pharmacokinetic profile of several analogs was also improved, as evidenced primarily by lower clearance in comparison with PI-88. The current data support the development of HS mimetics as potent antiangiogenic anticancer agents. Topics: Animals; Biomimetic Materials; Carcinoma, Hepatocellular; Cell Proliferation; Chemotherapy, Adjuvant; Clinical Trials, Phase III as Topic; Endothelial Cells; Fibroblast Growth Factor 1; Fibroblast Growth Factor 2; Heparin Lyase; Heparitin Sulfate; Humans; Neoplasm Metastasis; Neovascularization, Pathologic; Oligosaccharides; Vascular Endothelial Growth Factor A | 2007 |
34 other study(ies) available for heparitin-sulfate and Carcinoma--Hepatocellular
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Expression of glycosaminoglycans in cirrhotic liver and hepatocellular carcinoma-a pilot study including etiology.
Chronic liver diseases have both high incidence and mortality rates; therefore, a deeper understanding of the underlying molecular mechanisms is essential. We have determined the content and sulfation pattern of chondroitin sulfate (CS) and heparan sulfate (HS) in human hepatocellular carcinoma and cirrhotic liver tissues, considering the etiology of the diseases. A variety of pathological conditions such as alcoholic liver disease, hepatitis B and C virus infections, and primary sclerosing cholangitis were studied. Major differences were observed in the total abundance and sulfation pattern of CS and HS chains. For example, the 6-O-sulfation of CS is fundamentally different regarding etiologies of cirrhosis, and a 2-threefold increase in HS N-sulfation/O-sulfation ratio was observed in hepatocellular carcinoma compared to cirrhotic tissues. Topics: Carcinoma, Hepatocellular; Chondroitin Sulfates; Glycosaminoglycans; Heparitin Sulfate; Humans; Liver Cirrhosis; Liver Neoplasms; Pilot Projects | 2022 |
Syndecan-1 Promotes Hepatocyte-Like Differentiation of Hepatoma Cells Targeting Ets-1 and AP-1.
Syndecan-1 is a transmembrane heparan sulfate proteoglycan which is indispensable in the structural and functional integrity of epithelia. Normal hepatocytes display strong cell surface expression of syndecan-1; however, upon malignant transformation, they may lose it from their cell surfaces. In this study, we demonstrate that re-expression of full-length or ectodomain-deleted syndecan-1 in hepatocellular carcinoma cells downregulates phosphorylation of ERK1/2 and p38, with the truncated form exerting an even stronger effect than the full-length protein. Furthermore, overexpression of syndecan-1 in hepatoma cells is associated with a shift of heparan sulfate structure toward a highly sulfated type specific for normal liver. As a result, cell proliferation and proteolytic shedding of syndecan-1 from the cell surface are restrained, which facilitates redifferentiation of hepatoma cells to a more hepatocyte-like phenotype. Our results highlight the importance of syndecan-1 in the formation and maintenance of differentiated epithelial characteristics in hepatocytes partly via the HGF/ERK/Ets-1 signal transduction pathway. Downregulation of Ets-1 expression alone, however, was not sufficient to replicate the phenotype of syndecan-1 overexpressing cells, indicating the need for additional molecular mechanisms. Accordingly, a reporter gene assay revealed the inhibition of Ets-1 as well as AP-1 transcription factor-induced promoter activation, presumably an effect of the heparan sulfate switch. Topics: Carcinoma, Hepatocellular; Cell Differentiation; Cell Proliferation; Gene Expression Regulation, Neoplastic; Hep G2 Cells; Heparitin Sulfate; Hepatocyte Growth Factor; Hepatocytes; Humans; Liver Neoplasms; MAP Kinase Signaling System; Proto-Oncogene Protein c-ets-1; Signal Transduction; Syndecan-1; Transcription Factor AP-1 | 2020 |
Heparanase-1-induced shedding of heparan sulfate from syndecan-1 in hepatocarcinoma cell facilitates lymphatic endothelial cell proliferation via VEGF-C/ERK pathway.
Heparanase-1/syndecan-1 axis plays critical roles in tumorigenesis and development. The main mechanism includes heparanase-1 (HPA-1) degrades the heparan sulfate chain of syndecan-1 (SDC-1), and the following shedding of heparan sulfate from tumor cell releases and activates SDC-1 sequestered growth factors. However, the significance of Heparanase-1/syndecan-1 axis and its effects on the microenvironment of lymphatic metastasis in hepatocellular carcinogenesis (HCC) procession have not been reported. Herein, we found that HPA-1 could degrade the heparan sulfate on hepatocarcinoma cell surface. Importantly, HPA-1-induced shedding of heparan sulfate chain from SDC-1 facilitated the release of vascular endothelial growth factor C (VEGF-C) from SDC-1/VEGF-C complex into the medium of hepatocarcinoma cell. Further studies indicated that VEGF-C secretion from hepatocarcinoma cell promoted lymphatic endothelial cell growth through activating extracellular signal-regulated kinase (ERK) signaling. Taken together, this study reveals a novel existence of Heparanase-1/syndecan-1 axis in hepatocarcinoma cell and its roles in the cross-talking with the microenvironment of lymphatic metastasis. Topics: Animals; Blotting, Western; Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Proliferation; Endothelial Cells; Glucuronidase; Heparitin Sulfate; Liver Neoplasms; Lymphatic Metastasis; Lymphatic Vessels; MAP Kinase Signaling System; Mice; Protein Binding; Reverse Transcriptase Polymerase Chain Reaction; RNA Interference; Syndecan-1; Tumor Microenvironment; Vascular Endothelial Growth Factor C | 2017 |
Evaluating Tumor-Associated Activity of Extracellular Sulfatase by Analyzing Naturally Occurring Substrate in Tumor Microenvironment of Hepatocellular Carcinoma.
The progress of cancer is intimately connected with the activity of the extracellular matrix (ECM) enzymes. To evaluate the promoting effect of these enzymes on tumor development in a pathological biocontext, we propose in this work to analyze their natural substrates in the ECM. This strategy is demonstrated by studying heparan sulfate (HS), the substrate of ECM sulfatase, in the development of hepatocellular carcinoma (HCC). An assay is designed to study the abundance and sulfation of HS and to evaluate the interactions between HS and the growth factors, such as fibroblast growth factor 2 (FGF2). Peptides derived from the amyloid peptide and various growth factors are employed to detect HS and evaluate their affinity toward the growth factors, whereas the ruthenium polypyridyl complex is taken as a photocatalyst to achieve a more sensitive signal readout. Applying this method to HepG2 cells, correlated changes between the activity of sulfatase 2 in regulating FGF2-induced cell proliferation and the abundance, degree of sulfation, and growth factor binding of HS can be observed. This method has also been applied to analyze clinical tissue samples of HCC. The results may suggest tumor-progress-related alterations in the above-studied biochemical features of HS. These results may point to the prospect of using this method to facilitate the diagnosis and prognosis of HCC in the future. Topics: Carcinoma, Hepatocellular; Fibroblast Growth Factor 2; Hep G2 Cells; Heparitin Sulfate; Humans; Liver Neoplasms; Models, Molecular; Substrate Specificity; Sulfatases; Sulfotransferases; Tumor Microenvironment | 2016 |
Human Monoclonal Antibody Targeting the Heparan Sulfate Chains of Glypican-3 Inhibits HGF-Mediated Migration and Motility of Hepatocellular Carcinoma Cells.
Heparan sulfate proteoglycans (HSPGs) participate in many processes related to tumor development, including tumorigenesis and metastasis. HSPGs contain one or more heparan sulfate (HS) chains that are covalently linked to a core protein. Glypican-3 (GPC3) is a cell surface-associated HSPG that is highly expressed in hepatocellular carcinoma (HCC). GPC3 is involved in Wnt3a-dependent HCC cell proliferation. Our previous study reported that HS20, a human monoclonal antibody targeting the HS chains on GPC3, inhibited Wnt3a/β-catenin activation. In the current study, we showed that the HS chains of GPC3 could mediate HCC cells' migration and motility. Knocking down GPC3 or targeting the HS chains by HS20 inhibited HCC cell migration and motility. However, HS20 had no effect on GPC3 knockdown cells or GPC3 negative cells. In addition, an antibody that recognizes the core protein of GPC3 did not change the rate of cell motility. HCC cell migration and motility did not respond to either canonical or non-canonical Wnt induction, but did increase under hepatocyte growth factor (HGF) treatment. HS20-treated HCC cells exhibited less ability for HGF-mediated migration and motility. Furthermore, HS20 inhibited in vitro HCC spheroid formation and liver tumor growth in mice. GPC3 interacted with HGF; however, a mutant GPC3 lacking the HS chain showed less interaction with HGF. Blocking the HS chains on GPC3 with HS20 reduced c-Met activation in HGF-treated HCC cells and 3D-cultured spheroids. Taken together, our study suggests that GPC3 is involved in HCC cell migration and motility through HS chain-mediated cooperation with the HGF/Met pathway, showing how HS targeting has potential therapeutic implications for liver cancer. Topics: Antibodies, Monoclonal; Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Movement; Glypicans; Heparitin Sulfate; Hepatocyte Growth Factor; Humans; Liver Neoplasms | 2015 |
Inactivation of Wnt signaling by a human antibody that recognizes the heparan sulfate chains of glypican-3 for liver cancer therapy.
Wnt signaling is important for cancer pathogenesis and is often up-regulated in hepatocellular carcinoma (HCC). Heparan sulfate proteoglycans (HSPGs) function as coreceptors or modulators of Wnt activation. Glypican-3 (GPC3) is an HSPG that is highly expressed in HCC, where it can attract Wnt proteins to the cell surface and promote cell proliferation. Thus, GPC3 has emerged as a candidate therapeutic target in liver cancer. While monoclonal antibodies to GPC3 are currently being evaluated in preclinical and clinical studies, none have shown an effect on Wnt signaling. Here, we first document the expression of Wnt3a, multiple Wnt receptors, and GPC3 in several HCC cell lines, and demonstrate that GPC3 enhanced the activity of Wnt3a/β-catenin signaling in these cells. Then we report the identification of HS20, a human monoclonal antibody against GPC3, which preferentially recognized the heparan sulfate chains of GPC3, both the sulfated and nonsulfated portions. HS20 disrupted the interaction of Wnt3a and GPC3 and blocked Wnt3a/β-catenin signaling. Moreover, HS20 inhibited Wnt3a-dependent cell proliferation in vitro and HCC xenograft growth in nude mice. In addition, HS20 had no detectable undesired toxicity in mice. Taken together, our results show that a monoclonal antibody primarily targeting the heparin sulfate chains of GPC3 inhibited Wnt/β-catenin signaling in HCC cells and had potent antitumor activity in vivo.. An antibody directed against the heparan sulfate of a proteoglycan shows efficacy in blocking Wnt signaling and HCC growth, suggesting a novel strategy for liver cancer therapy. Topics: Animals; Antibodies, Monoclonal; beta Catenin; Carcinoma, Hepatocellular; Cell Surface Display Techniques; Female; Glypicans; Hep G2 Cells; Heparitin Sulfate; Humans; Liver Neoplasms; Mice; Mice, Inbred BALB C; Mice, Nude; Wnt Signaling Pathway; Xenograft Model Antitumor Assays | 2014 |
WNT/β-catenin signaling and hepatocellular carcinoma.
Topics: Animals; Antibodies, Monoclonal; Carcinoma, Hepatocellular; Female; Glypicans; Heparitin Sulfate; Humans; Liver Neoplasms; Wnt Signaling Pathway | 2014 |
Granulin-epithelin precursor interacts with heparan sulfate on liver cancer cells.
Granulin-epithelin precursor (GEP) is a pluripotent secretory growth factor which promotes cancer progression in a number of human cancers. However, how cancer cells interact with GEP remains unknown. In this study, we aimed to identify the cell surface-binding partner of GEP on liver cancer cells. Human recombinant GEP (rGEP) was expressed and purified to homogeneity. The rGEP was shown to trigger phosphorylation of AKT and ERK1/2 in liver cancer cells. We demonstrated cell surface attachment of rGEP, which was blocked by prebinding of platelet-derived growth factor-AA, platelet-derived growth factor-BB and fibroblast growth factor-2. Therefore, heparan sulfate (HS) had been reasoned as the binding partner of rGEP. Heparinase digestion validated the role of HS on supporting the attachment. The heparin-binding domain of GEP was mapped to RRH(555-557) in the C-terminal region. Suppression of the HS polymerase exostosin-1 reduced the rGEP binding and rGEP-mediated signaling transduction. Suppression of a specific HS proteoglycan, glypican-3, also showed a partial reduction of rGEP binding and an inhibition on rGEP-mediated activation of AKT. Furthermore, glypican-3 was shown to correlate with the expressions of GEP in clinical samples (Spearman's ρ = 0.363, P = 0.001). This study identified HS, partly through glypican-3, as a novel binding partner of GEP on the surface of liver cancer cells. Topics: Carcinoma, Hepatocellular; Fibroblast Growth Factor 2; Gene Expression Regulation, Neoplastic; Glypicans; Hep G2 Cells; Heparitin Sulfate; Humans; Intercellular Signaling Peptides and Proteins; Liver Neoplasms; MAP Kinase Signaling System; Oncogene Protein v-akt; Progranulins; Protein Binding | 2014 |
Heparin and liver heparan sulfate can rescue hepatoma cells from topotecan action.
Topotecan (TpT) is a major inhibitory compound of topoisomerase (topo) I that plays important roles in gene transcription and cell division. We have previously reported that heparin and heparan sulfate (HS) might be transported to the cell nucleus and they can interact with topoisomerase I. We hypothesized that heparin and HS might interfere with the action of TpT. To test this hypothesis we isolated topoisomerase I containing cell nuclear protein fractions from normal liver, liver cancer tissues, and hepatoma cell lines. The enzymatic activity of these extracts was measured in the presence of heparin, liver HS, and liver cancer HS. In addition, topo I activity, cell viability, and apoptosis of HepG2 and Hep3B cells were investigated after heparin and TpT treatments. Liver cancer HS inhibited topo I activity in vitro. Heparin treatment abrogated topo I enzyme activity in Hep3B cells, but not in HepG2 cells, where the basal activity was higher. Heparin protected the two hepatoma cell lines from TpT actions and decreased the rate of TpT induced S phase block and cell death. These results suggest that heparin and HS might interfere with the function of TpT in liver and liver cancer. Topics: Carcinoma, Hepatocellular; Cell Cycle; Cell Proliferation; DNA Topoisomerases, Type I; DNA, Neoplasm; Hep G2 Cells; Heparin; Heparitin Sulfate; Humans; Liver; Liver Neoplasms; Topotecan | 2014 |
Mutation of herpesvirus Saimiri ORF51 glycoprotein specifically targets infectivity to hepatocellular carcinoma cell lines.
Herpesvirus saimiri (HVS) is a gamma herpesvirus with several properties that make it an amenable gene therapy vector; namely its large packaging capacity, its ability to persist as a nonintegrated episome, and its ability to infect numerous human cell types. We used RecA-mediated recombination to develop an HVS vector with a mutated virion protein. The heparan sulphate-binding region of HVS ORF51 was substituted for a peptide sequence which interacts with somatostatin receptors (SSTRs), overexpressed on hepatocellular carcinoma (HCC) cells. HVS mORF51 showed reduced infectivity in non-HCC human cell lines compared to wild-type virus. Strikingly, HVS mORF51 retained its ability to infect HCC cell lines efficiently. However, neutralisation assays suggest that HVS mORF51 has no enhanced binding to SSTRs. Therefore, mutation of the ORF51 glycoprotein has specifically targeted HVS to HCC cell lines by reducing the infectivity of other cell types; however, the mechanism for this targeting is unknown. Topics: Amino Acid Sequence; Animals; Carcinoma, Hepatocellular; Cell Line, Tumor; Cloning, Molecular; Electrophoresis, Agar Gel; Flow Cytometry; Genetic Vectors; Heparitin Sulfate; Herpesvirus 2, Saimiriine; Humans; Liver Neoplasms; Membrane Glycoproteins; Mice; Molecular Sequence Data; Mutation; Neutralization Tests; Receptors, Somatostatin; Viral Envelope Proteins | 2011 |
Quantitative and qualitative alterations of heparan sulfate in fibrogenic liver diseases and hepatocellular cancer.
Heparan sulfate (HS), due to its ability to interact with a multitude of HS-binding factors, is involved in a variety of physiological and pathological processes. Remarkably diverse fine structure of HS, shaped by non-exhaustive enzymatic modifications, influences the interaction of HS with its partners. Here we characterized the HS profile of normal human and rat liver, as well as alterations of HS related to liver fibrogenesis and carcinogenesis, by using sulfation-specific antibodies. The HS immunopattern was compared with the immunolocalization of selected HS proteoglycans. HS samples from normal liver and hepatocellular carcinoma (HCC) were subjected to disaccharide analysis. Expression changes of nine HS-modifying enzymes in human fibrogenic diseases and HCC were measured by quantitative RT-PCR. Increased abundance and altered immunolocalization of HS was paralleled by elevated mRNA levels of HS-modifying enzymes in the diseased liver. The strong immunoreactivity of the normal liver for 3-O-sulfated epitope further increased with disease, along with upregulation of 3-OST-1. Modest 6-O-undersulfation of HCC HS is probably explained by Sulf overexpression. Our results may prompt further investigation of the role of highly 3-O-sulfated and partially 6-O-desulfated HS in pathological processes such as hepatitis virus entry and aberrant growth factor signaling in fibrogenic liver diseases and HCC. Topics: Agrin; Animals; Carcinoma, Hepatocellular; Chronic Disease; Disaccharides; Focal Nodular Hyperplasia; Glucuronidase; Glypicans; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Humans; Immunohistochemistry; Liver; Liver Cirrhosis; Liver Neoplasms; Male; Rats; Rats, Wistar; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Sulfotransferases; Syndecan-1 | 2010 |
WSS25 inhibits growth of xenografted hepatocellular cancer cells in nude mice by disrupting angiogenesis via blocking bone morphogenetic protein (BMP)/Smad/Id1 signaling.
The highly expressed Id1 (inhibitor of DNA binding/differentiation) protein promotes angiogenesis in HCC and is a well established target for anti-angiogenesis therapeutic strategies. Heparan sulfate (HS) mimetics such as PI-88 can abrogate HS-protein interactions to inhibit angiogenesis. Id1 is the direct downstream effector of bone morphogenetic proteins (BMPs), which are angiogenic and HS-binding proteins. Thus, targeting BMPs by HS mimetics may inhibit angiogenesis via attenuating Id1 expression. We report here that a HS mimetic WSS25 potently inhibited the tube formation of HMEC-1 cells on Matrigel and their migration. Meanwhile, WSS25 (25 μg/ml) nearly completely blocked Id1 expression in the HMEC-1 cells as demonstrated by oligo-angiogenesis microarray analysis and further confirmed by RT-PCR and Western blotting. BMP/Smad/Id1 signaling also was blocked by WSS25 treatment in HMEC-1 cells. Importantly, Id1 knockdown in HMEC-1 cells caused the disruption of their tube formation on Matrigel. By employing quartz crystal microbalance analysis, we found that WSS25 strongly bound to BMP2. Moreover, WSS25 impaired BMP2-induced tube formation of HMEC-1 cells on Matrigel and angiogenesis in Matrigel transplanted into C57BL6 mice. Furthermore, WSS25 (100 mg/kg) abrogated the growth of HCC cells xenografted in male nude mice. Immunohistochemical analysis showed that both the expression of Id1 and the endothelial cell marker CD31 were lower in the WSS25-treated tumor tissue than in the control. Therefore, WSS25 is a potential drug candidate for HCC therapy as a tumor angiogenesis inhibitor. Topics: Animals; Bone Morphogenetic Proteins; Carcinoma, Hepatocellular; Cell Line, Tumor; Glucans; Heparitin Sulfate; Humans; Inhibitor of Differentiation Protein 1; Liver Neoplasms; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Nude; Molecular Structure; Neoplasm Transplantation; Neoplasms, Experimental; Neovascularization, Pathologic; Quartz Crystal Microbalance Techniques; Signal Transduction; Smad Proteins; Transplantation, Heterologous | 2010 |
Glycosaminoglycan mimetics inhibit SDF-1/CXCL12-mediated migration and invasion of human hepatoma cells.
We have recently reported that the CXC-chemokine stromal cell-derived factor-1 (SDF-1)/CXCL12 induces proliferation, migration, and invasion of the Huh7 human hepatoma cells through its G-protein-coupled receptor CXCR4 and that glycosaminoglycans (GAGs) are involved in these events. Here, we demonstrate by surface plasmon resonance that the chemokine binds to GAG mimetics obtained by grafting carboxylate, sulfate or acetate groups onto a dextran backbone. We also demonstrate that chemically modified dextrans inhibit SDF-1/CXCL12-mediated in vitro chemotaxis and anchorage-independent cell growth in a dose-dependent manner. The binding of GAG mimetics to the chemokine and their effects in modulating the SDF-1/CXCL12 biological activities are mainly related to the presence of sulfate groups. Furthermore, the mRNA expression of enzymes involved in heparan sulfate biosynthesis, such as exostosin-1 and -2 or N-deacetylase N-sulfotransferases remained unchanged, but heparanase mRNA and protein expressions in Huh7 cells were decreased upon GAG mimetic treatment. Moreover, decreasing heparanase-1 mRNA levels by RNA interference significantly reduced SDF-1/CXCL12-induced extracellular signal-regulated kinase 1/2 (ERK 1/2) phosphorylation. Therefore, we suggest that GAG mimetic effects on SDF-1/CXCL12-mediated hepatoma cell chemotaxis may rely on decreased heparanase expression, which impairs SDF-1/CXCL12's signaling. Altogether, these data suggest that GAG mimetics may compete with cellular heparan sulfate chains for the binding to SDF-1/CXCL12 and may affect heparanase expression, leading to reduced SDF-1/CXCL12 mediated in vitro chemotaxis and growth of hepatoma cells. Topics: Binding, Competitive; Carcinoma, Hepatocellular; Cell Adhesion; Cell Communication; Cell Movement; Chemokine CXCL12; Glucuronidase; Glycosaminoglycans; Heparitin Sulfate; Humans; Liver Neoplasms; Neoplasm Invasiveness; RNA, Small Interfering; Tumor Cells, Cultured | 2009 |
hSulf1 Sulfatase promotes apoptosis of hepatocellular cancer cells by decreasing heparin-binding growth factor signaling.
The heparin-binding growth factors fibroblast growth factor (FGF) and hepatocyte growth factor (HGF) are potent mitogens for hepatocellular carcinomas (HCCs). Heparin-binding growth factor signaling is regulated by sulfation of cell-surface heparan sulfate proteoglycans (HSPGs). We hypothesized that hSulf1, a recently described sulfatase, regulates growth signaling in HCCs.. Expression of hSulf1 in human HCC tumors was determined by real-time PCR. Down-regulation of hSulf1 expression was investigated by analyzing loss of heterozygosity (LOH) at the hSulf1 locus and the effect of the DNA methylation inhibitor 5-aza-deoxycytidine on hSulf1 expression. The subcellular location of hSulf1 and sulfation state of cell-surface HSPGs were assessed by immunocytochemistry. FGF and HGF signaling was examined by phospho-specific immunoblot analysis. Cell growth was measured by trypan blue exclusion, and the MTT assay and apoptosis were quantitated by fluorescence microscopy.. hSulf1 expression was decreased in 29% of HCCs and 82% of HCC cell lines. There was LOH at the hSulf1 locus in 42% of HCCs. Treatment with 5-aza-deoxycytidine reactivated hSulf1 expression in hSulf1-negative cell lines. Low hSulf1-expressing cells showed increased sulfation of cell-surface HSPGs, enhanced FGF and HGF-mediated signaling, and increased HCC cell growth. Conversely, forced expression of hSulf1 decreased sulfation of cell-surface HSPGs and abrogated growth signaling. HCC cells with high-level hSulf1 expression were sensitive to staurosporine- or cisplatin-induced apoptosis, whereas low expressing cells were resistant. Transfection of hSulf1 into hSulf1-negative cells restored staurosporine and cisplatin sensitivity.. Down-regulation of hSulf1 contributes to hepatocarcinogenesis by enhancing heparin-binding growth factor signaling and resistance to apoptosis. Topics: Apoptosis; Carcinoma, Hepatocellular; Cell Division; Cell Line, Tumor; Cell Membrane; Cisplatin; DNA Methylation; Fibroblast Growth Factor 2; Heparitin Sulfate; Hepatocyte Growth Factor; Humans; Liver Neoplasms; Loss of Heterozygosity; Protein Structure, Tertiary; RNA, Messenger; Signal Transduction; Staurosporine; Sulfates; Sulfotransferases | 2004 |
Exploitation of heparanase inhibitors from microbial metabolites using an efficient visual screening system.
In this paper we describe the establishment of an efficient visual method for screening heparanase inhibitors, and we present the results of screening 10,000 microbial culture broths. Heparanase-overexpressing stable clones of the human hepatocellular carcinoma HepG2 cells were established and used as an enzyme source. Digestion of heparan sulfate (HS) was detected using novel HS-containing tablets or SDS-polyacrylamide gel electrophoresis. This method was able to find suramin, a known heparanase inhibitor, from a library of typical enzyme inhibitors. By screening 10,000 culture broths of microorganisms (actinomycetes, fungi, and bacteria) an actinomycete strain, RK99-A234, was found to have heparanase inhibitory activity. RK-682 was identified in the fermentation broth as a heparanase inhibitor, IC50 = 17 microM. Topics: Actinobacteria; Animals; Blotting, Western; Carcinoma, Hepatocellular; Cell Line, Tumor; Clone Cells; Culture Media; DNA, Complementary; Drug Evaluation, Preclinical; Electrophoresis, Polyacrylamide Gel; Enzyme Inhibitors; Glucuronidase; Heparitin Sulfate; Humans; Liver Neoplasms; Phosphoprotein Phosphatases; Transfection | 2004 |
Expression of the CD44v2-10 isoform confers a metastatic phenotype: importance of the heparan sulfate attachment site CD44v3.
We expressed the full-length CD44v2-10 isoform in SKHep1 cells, a nonmetastatic human hepatocellular carcinoma cell line that does not express any endogenous CD44v isoforms. In SCID mice, expression of CD44v2-10 by SKHep1 cells had no effect on s.c. primary tumor development but caused pulmonary metastases in 41% (7 of 17) of animals compared with control SKHep1 cells (0 of 16; P < 0.01). CD44v2-10 expression by SKHep1 cells resulted in enhanced heparan sulfate (HS) attachment and an enhanced capacity to bind heparin-binding growth factors. Mutation of the v3 domain to prevent HS attachment and growth factor binding abolished the metastatic phenotype, demonstrating that HS modification of CD44v2-10 plays a critical role in the development of metastases in this model. However, in vitro proliferation, motility, and invasion were not altered by CD44v2-10 expression. Topics: Alternative Splicing; Binding Sites; Carcinoma, Hepatocellular; Cell Adhesion; Cell Division; Cell Movement; Growth Substances; Heparitin Sulfate; Humans; Hyaluronan Receptors; Hyaluronic Acid; Liver Neoplasms; Lung Neoplasms; Neoplasm Metastasis; Protein Isoforms; Tumor Cells, Cultured | 2003 |
Effect of heparin and liver heparan sulphate on interaction of HepG2-derived transcription factors and their cis-acting elements: altered potential of hepatocellular carcinoma heparan sulphate.
Proteoglycan assembly in malignant tumours is subject to profound changes. The significance of these alterations is not well understood; especially, their role in nuclear regulation is a topic for debate. The capacity of heparin and liver carcinoma heparan sulphate (HS) to alter DNA-transcription factor interactions has been studied to provide further evidence concerning the regulatory potential of glycosaminoglycan (GAG) in the nucleus. Experiments both in vitro and in vivo indicated that heparin and HS are capable of inhibiting the interaction of transcription factors with their consensus oligonucleotide elements. Among five transcription factors studied, AP-1, SP-1, ETS-1 and nuclear factor kappaB proved to be sensitive to heparin and heparan sulphate, whereas TFIID was hardly inhibited in either in vitro or in vivo systems. Interestingly, HS from peritumoral liver was five times more effective than heparin. Liver carcinoma HS was less effective than liver HS, but its activity was comparable with that of heparin. These results indicate that the structural differences of GAG chains strongly influence their biological behaviour. The loss of their recognized functional activity in malignant tumours might promote the development of uncontrolled growth and gene expression favouring the neoplastic process. Topics: Carcinoma, Hepatocellular; DNA-Binding Proteins; Heparin; Heparitin Sulfate; Humans; Liver; Liver Neoplasms; Protein Binding; Transcription Factors; Tumor Cells, Cultured | 2000 |
The HepG2 extracellular matrix contains separate heparinase- and lipid-releasable pools of ApoE. Implications for hepatic lipoprotein metabolism.
We have examined the association of apoE with the extracellular matrix (ECM) of HepG2 cells. Comparison of ECM prepared by previously published methods demonstrated that cytochalasin B-prepared material yielded the highest endogenous apoE, representing 23.6% of that in cell monolayers. ECM prepared with EDTA or Triton X-100 exhibited decreased levels of apoE, 3 and 6%, respectively. ECM bound very low density lipoprotein poorly (5-6% of the monolayer capacity); however, these incubations dramatically increased the apoE content of the ECM. Heparinase or suramin decreased apoE of the ECM by 19.6 and 37.3%, respectively, suggesting association with heparin sulfate proteoglycans. EDTA or EGTA also displaced 35% of the apoE, suggesting a Ca2+-dependent association. Incubation with phosphatidylcholine vesicles (PCV) displaced 30% of the apoE, suggesting that lipid content affects association of apoE with the ECM. Data derived from sequential incubations with combinations of suramin, EGTA, and PCV were consistent with the presence of two distinct pools of apoE on the HepG2 ECM, one releasable with suramin and EGTA and the other releasable with lipids. Exogenously applied lipid-free apoE readily bound to the ECM; however, increasing the lipid content decreased its association. Lipid-free apoE could be equally displaced from the ECM with PCV or suramin. When lipid-free apoE adsorbed to microtiter wells was incubated with a triglyceride emulsion or palmitoyloleyl phosphatidylcholine micelles, the immunoreactivity of 3H1 (but not other antibodies), a monoclonal antibody against an epitope in the C-terminal domain of apoE, increased about 4-fold. In a similar manner, incubation of ECM with lipid dramatically increased the immunoreactivity of 3H1, indicating that apoE of the ECM exists in a lipid-poor form. Scatchard analysis demonstrated that the increased immunoreactivity was due to an increase in the number of antibody binding sites. In conclusion, the ECM contains two pools of lipid-poor apoE. One pool associates with the ECM through heparin sulfate proteoglycans- and Ca2+-dependent interactions. A second pool of apoE dissociates from the ECM upon lipidation. The lipid-sensitive pool of apoE may participate in secretion or efflux of lipids or in the capture of lipoproteins by providing the apoE needed for receptor-mediated uptake. Topics: Antibodies, Monoclonal; Apolipoproteins E; Carcinoma, Hepatocellular; Egtazic Acid; Emulsions; Extracellular Matrix; Fat Emulsions, Intravenous; Heparin Lyase; Heparitin Sulfate; Humans; Lipids; Lipoproteins; Phosphatidylcholines; Phospholipids; Protein Binding; Safflower Oil; Soybean Oil; Suramin; Tumor Cells, Cultured | 1998 |
Substrate specificity of heparanases from human hepatoma and platelets.
Heparan sulfate proteoglycans, attached to cell surfaces or in the extracellular matrix, interact with a multitude of proteins via their heparan sulfate side chains. Degradation of these chains by limited (endoglycosidic) heparanase cleavage is believed to affect a variety of biological processes. Although the occurrence of heparanase activity in mammalian tissues has been recognized for many years, the molecular characteristics and substrate recognition properties of the enzyme(s) have remained elusive. In the present study, the substrate specificity and cleavage site of heparanase from human hepatoma and platelets were investigated. Both enzyme preparations were found to cleave the single beta-D-glucuronidic linkage of a heparin octasaccharide. A capsular polysaccharide from Escherichia coli K5, with the same (-GlcUAbeta1,4-GlcNAcalpha1,4-)n structure as the unmodified backbone of heparan sulfate, resisted heparanase degradation in its native state as well as after chemical N-deacetylation/N-sulfation or partial enzymatic C-5 epimerization of beta-D-GlcUA to alpha-L-IdceA. By contrast, a chemically O-sulfated (but still N-acetylated) K5 derivative was susceptible to heparanase cleavage. O-Sulfate groups, but not N-sulfate or IdceA residues, thus are essential for substrate recognition by the heparanase(s). In particular, selective O-desulfation of the heparin octasaccharide implicated a 2-O-sulfate group on a hexuronic acid residue located two monosaccharide units from the cleavage site, toward the reducing end. Topics: Animals; Antithrombin III; Blood Platelets; Carbohydrate Sequence; Carcinoma, Hepatocellular; Escherichia coli; Glucuronidase; Glycoside Hydrolases; Heparin; Heparitin Sulfate; Humans; Isoenzymes; Liver Neoplasms; Mast-Cell Sarcoma; Mice; Microsomes; Molecular Sequence Data; Substrate Specificity; Tumor Cells, Cultured | 1998 |
Structural characterization and functional effects of a circulating heparan sulfate in a patient with hepatocellular carcinoma.
A circulating anticoagulant was isolated from the plasma of a 42-year-old man with cirrhosis and hepatocellular carcinoma who had an unusual coagulation test profile. The patient developed a fatal coagulopathy, unresponsive to protamine therapy or plasma exchange following liver biopsy. However, at presentation, routine hemostasis assays were normal. The patient had mucocutaneous bleeding but the sole laboratory abnormality was a prolonged thrombin time (TT = 99 s, normal 25-35 s). Protamine titration indicated activity equivalent to a heparin concentration of 6-7 U/ml. Antithrombin III (AT III) antigen and activity were markedly elevated. The anticoagulant activity, purified from plasma by DEAE chromatography, was identified as a glycosaminoglycan (GAG). GAG anti-thrombin activity was completely abolished by heparin lyase III. Based on the degree of sulfation and HPLC pattern, the GAG was classified as heparan sulfate. Low levels (4 microM) of purified GAG markedly prolonged the TT (>120 s) but not the activated partial thromboplastin time (PTT) (31.4 s). In a Factor Xa assay, the GAG exhibited a potency equivalent to 0.06 U of low molecular weight heparin per nmol of uronic acid. Patients with endogenous circulating glycosaminoglycans can present with unusual laboratory coagulation test profiles. These reflect complex dysfunction of hemostasis, leading to difficulty in providing diagnosis and effective care. Topics: Adult; Blood Coagulation; Blood Platelets; Carcinoma, Hepatocellular; Disaccharides; Factor Xa; Glycosaminoglycans; Heparitin Sulfate; Humans; Male; Partial Thromboplastin Time; Protein Conformation; Receptors, Thrombin; Sulfuric Acid Esters; Thrombin; Thrombin Time | 1998 |
Heparan sulphate proteoglycan expression in human primary liver tumours.
Heparan sulphate proteoglycans (HSPGs) play important biological roles in cell-matrix adhesion processes and are essential regulators of growth factor actions (e.g., as co-receptor for hepatocyte growth factor). Since in liver carcinogenesis, interactions between cells, the matrix, and growth factors play a major role, the aim of this study was to investigate whether the distribution pattern of HSPGs is altered in human primary liver tumours. Twenty-two primary liver tumours and five normal liver biopsies were studied, using specific monoclonal antibodies against syndecans-1, -2, -3, and -4; glypican; perlecan; and heparan sulphate chains. Cholangiocarcinomas as well as hepatocellular carcinomas showed an altered immunoreactivity pattern of the different HSPGs in comparison with normal liver parenchyma, probably reflecting the growth regulatory roles of HSPGs. Intracellular positivity for integral membrane HSPGs syndecan-1 and especially syndecan-4 was a constant finding in most tumours, suggesting increased synthesis or internalization of these HSPGs. Syndecan-3 and perlecan expression in tumours was found in an expected distribution pattern. The strong reactivity for syndecan-3 and perlecan in tumoral stromal vessels might suggest a role for these HSPGs in tumoral angiogenesis. In addition, perlecan probably exerts its known growth factor reservoir function also in the stroma of primary liver tumours. Topics: Bile Duct Neoplasms; Bile Ducts, Intrahepatic; Carcinoma, Hepatocellular; Cholangiocarcinoma; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Humans; Immunohistochemistry; Liver; Liver Neoplasms; Membrane Glycoproteins; Microscopy, Immunoelectron; Proteoglycans; Retrospective Studies; Syndecan-1; Syndecans | 1998 |
Protein kinase C bound to the Golgi apparatus supports the formation of constitutive transport vesicles.
Constitutive secretion of heparan sulphate proteoglycans (HSPGs) was stimulated in human hepatoma HepG2 cells by phorbol 12-myristate 13-acetate (PMA) and inhibited by calphostin C, a specific inhibitor of protein kinase C (PKC). To delineate more closely the site of PKC action, the packaging in vitro of 35SO4-labelled HSPGs into transport vesicles was investigated. Formation of transport vesicles at the trans-Golgi network was stimulated by PMA and inhibited by calphostin C or Ro 31-8220 by using a post-nuclear supernatant. Treatment of either isolated Golgi-enriched membranes or cytosolic proteins with calphostin C provided evidence that membrane-bound PKC forms strongly supported vesicle formation, whereas cytosolic PKC forms showed a marginal effect. The PKC isoforms PKC-alpha and PKC-zeta were attached to highly purified Golgi membranes, as shown by Western blotting. Both isoforms were localized by confocal immunofluorescence microscopy in the Golgi area of HepG2 cells. Immunoelectron microscopy of ultrathin cryosections of HepG2 cells showed that PKC-zeta predominantly attaches to the trans-Golgi region, whereas PKC-alpha binds to the cis- and trans-Golgi area. Topics: Carcinoma, Hepatocellular; Cell Fractionation; Cell Line; Cell-Free System; Cytoplasmic Granules; Egtazic Acid; Enzyme Inhibitors; Golgi Apparatus; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Humans; Indoles; Intracellular Membranes; Isoenzymes; Kinetics; Liver Neoplasms; Naphthalenes; Protein Binding; Protein Kinase C; Protein Kinase C-alpha; Proteoglycans | 1996 |
Role of heparan sulfate proteoglycans in the binding and uptake of apolipoprotein E-enriched remnant lipoproteins by cultured cells.
Addition of apolipoprotein (apo) E to rabbit beta-very low density lipoproteins (beta-VLDL) has been shown to result in a marked enhancement of their binding and uptake by various cell types. Apolipoprotein E binds to lipoprotein receptors and proteoglycans. To distinguish between apoE binding to these sites, cells were treated with heparinase. Heparinase treatment of receptor-negative familial hypercholesterolemic (FH) fibroblasts and human hepatoma cells (HepG2) released 30-40% of newly synthesized cell surface 35S-labeled proteoglycans and decreased the binding of beta-VLDL+apoE to FH and normal fibroblasts and HepG2 cells by more than 80%. Furthermore, heparinase treatment significantly decreased the uptake of fluorescently labeled beta-VLDL+apoE by HepG2 cells and decreased cholesteryl ester synthesis in FH fibroblasts by 75%. Likewise, canine chylomicron remnants enriched in apoE demonstrated enhanced binding that was 80% inhibited by heparinase treatment of HepG2 cells. Heparinase treatment did not affect beta-VLDL (without added apoE) or low density lipoprotein (LDL) binding to these cells or the binding activity of beta-VLDL+apoE to the LDL receptor-related protein (LRP) or to the LDL receptor on ligand blots. Chinese hamster ovary (CHO) mutant cells lacking the synthesis of either heparan sulfate (pgsD-677) or all proteoglycans (pgsA-745) did not display any enhanced binding of the beta-VLDL+apoE. By comparison, wild-type CHO cells demonstrated enhanced binding of beta-VLDL+apoE that could be abolished by treatment with heparinase. These mutant cells and wild-type CHO cells possessed a similar amount of LRP, as determined by ligand blot analyses and by alpha 2-macroglobulin binding, and possessed a similar amount of LDL receptor activity, as determined by LDL binding. Therefore, we would interpret these data as showing that heparan sulfate proteoglycan may be involved in the initial binding of the apoE-enriched remnants with the subsequent involvement of the LRP in the uptake of these lipoproteins. It remains to be determined whether the heparan sulfate proteoglycan can function by itself in both the binding and internalization of the apoE-enriched remnants or whether the proteoglycan is part of a complex with LRP that mediates a two-step process, i.e. binding and subsequent internalization by the receptor. Topics: alpha-Macroglobulins; Animals; Apolipoproteins E; Biological Transport; Carcinoma, Hepatocellular; Cell Membrane; Cells, Cultured; CHO Cells; Cholesterol; Cholesterol Esters; Cricetinae; Diet, Atherogenic; Fibroblasts; Heparan Sulfate Proteoglycans; Heparin Lyase; Heparitin Sulfate; Humans; Hyperlipoproteinemia Type II; Lipoproteins, LDL; Lipoproteins, VLDL; Liver; Liver Neoplasms; Membrane Proteins; Polysaccharide-Lyases; Protein Binding; Proteoglycans; Rabbits; Rats; Receptors, LDL; Transfection; Tumor Cells, Cultured | 1993 |
Potential markers (enzymes, proteoglycans) for human liver tumors.
In the present work the activities of GGT and G-6-Pase and the content of Cyt P-450 were determined in surgically removed liver specimens (16 hepatocellular carcinomas, 8 focal nodular hyperplasias and 4 adenomas). The activities were compared to the surrounding seemingly normal liver tissue. In the adenomas neither of the enzymes studied showed alterations, characteristic for hepatocarcinogenesis. Four out of 8 FNHs had the enzyme pattern that was found in experimental liver carcinogenesis. Liver carcinoma specimens proved to be heterogenous. Neither elevated GGT nor reduced G-6-Pase activity was consistent in these samples although the average of G-6-Pase activity decreased to 50 percent. Cytochrome P-450 was significantly reduced in the majority of cases, showing the best agreement with the tendency observed in experimental models. As an other approach, the qualitative and quantitative alterations of proteoglycans (PG) were analized in the same tumor samples. The amount of sugar components of PGs the glycosaminoglycans (GAG) increased by many times in liver tumors. Carcinoma samples were characterized by about twentyfold increase in chondroitin sulfate content, compared to normal liver. The enhancement of GAGs is partly the consequence of a selective alteration in PG expression. The amount of perlecan and decorin was found to be increased, while syndecan disappeared from liver carcinomas. These data suggest that malignant transformation in liver is accompanied by specific alteration in the content, composition and structure of PGs. Presumably, these changes have significance in tumor progression and have also the potential to be used as markers for liver tumors. Topics: Adenoma; Biomarkers, Tumor; Carcinoma, Hepatocellular; Clinical Enzyme Tests; Cytochrome P-450 Enzyme System; Diagnosis, Differential; gamma-Glutamyltransferase; Glucose-6-Phosphatase; Glycosaminoglycans; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Humans; Hyperplasia; Liver; Liver Neoplasms; Membrane Glycoproteins; Proteoglycans; Syndecans | 1993 |
Low density lipoprotein receptor internalizes low density and very low density lipoproteins that are bound to heparan sulfate proteoglycans via lipoprotein lipase.
It has previously been shown that lipoprotein lipase (LPL) enhances the binding of low density lipoproteins (LDL) and very low density lipoproteins (VLDL) to HepG2 cells and fibroblasts, up to 80-fold. This increase in binding is LDL receptor-independent and is due to a bridging of LPL between extracellular heparan sulfate proteoglycans (HSPG) and the lipoproteins. In the present paper, we show that preincubation of the cells with LPL, followed by washing prior to the binding experiment, increased binding to the same extent as occurs when the binding is performed in the presence of LPL. This indicates that the formation of a complex of LPL with the lipoproteins is not a prerequisite of binding. Binding curves and Scatchard analyses reveal that both the number of binding sites and the affinity of the binding are increased 20-30-fold by the addition of 3.4 micrograms/ml LPL. The addition of LPL also resulted in an enhanced uptake and subsequent lysosomal degradation of both LDL and VLDL when compared with binding, although to a lesser extent (up to 25-fold when measured after 5 h at 37 degrees C). Strikingly, enhanced uptake did not occur in LDL receptor-negative fibroblasts. In addition, down-regulation of the LDL receptor activity by preincubation of the cells for 48 h with either LDL or beta-VLDL resulted in a parallel decrease in the uptake of LPL-mediated HSPG-bound LDL, whereas the LPL-mediated binding itself was not diminished. These observations indicate that the uptake of LPL-mediated HSPG-bound LDL and VLDL mainly proceeds via the LDL receptor. Binding of labeled LDL to the cells at 4 degrees C for 2 h followed by a chase period at 37 degrees C revealed that in absolute terms, the initial rate of internalization of HSPG-bound LDL is comparable with that of LDL receptor-bound LDL (0.58 and 0.44 ng/min/mg of cell protein, respectively). We conclude that in LDL receptor-positive cells, the LPL-mediated binding of LDL and VLDL to HSPG is followed by internalization of the lipoproteins mainly through the rapid process of the classical LDL receptor recycling system, whereas only a minor portion is internalized via the much slower process of HSPG uptake. Topics: Animals; Biological Transport; Carcinoma, Hepatocellular; Cattle; Female; Fibroblasts; Heparan Sulfate Proteoglycans; Heparin Lyase; Heparitin Sulfate; Humans; Kinetics; Lipoprotein Lipase; Lipoproteins, LDL; Lipoproteins, VLDL; Liver Neoplasms; Milk; Models, Biological; Polysaccharide-Lyases; Protein Binding; Proteoglycans; Receptors, LDL; Tumor Cells, Cultured | 1993 |
Mechanisms by which lipoprotein lipase alters cellular metabolism of lipoprotein(a), low density lipoprotein, and nascent lipoproteins. Roles for low density lipoprotein receptors and heparan sulfate proteoglycans.
We sought to investigate effects of lipoprotein lipase (LpL) on cellular catabolism of lipoproteins rich in apolipoprotein B-100. LpL increased cellular degradation of lipoprotein(a) (Lp(a)) and low density lipoprotein (LDL) by 277% +/- 3.8% and 32.5% +/- 4.1%, respectively, and cell association by 509% +/- 8.7% and 83.9% +/- 4.0%. The enhanced degradation was entirely lysosomal. Enhanced degradation of Lp(a) had at least two components, one LDL receptor-dependent and unaffected by heparitinase digestion of the cells, and the other LDL receptor-independent and heparitinase-sensitive. The effect of LpL on LDL degradation was entirely LDL receptor-independent, heparitinase-sensitive, and essentially absent from mutant Chinese hamster ovary cells that lack cell surface heparan sulfate proteoglycans. Enhanced cell association of Lp(a) and LDL was largely LDL receptor-independent and heparitinase-sensitive. The ability of LpL to reduce net secretion of apolipoprotein B-100 by HepG2 cells by enhancing cellular reuptake of nascent lipoproteins was also LDL receptor-independent and heparitinase-sensitive. None of these effects on Lp(a), LDL, or nascent lipoproteins required LpL enzymatic activity. We conclude that LpL promotes binding of apolipoprotein B-100-rich lipoproteins to cell surface heparan sulfate proteoglycans. LpL also enhanced the otherwise weak binding of Lp(a) to LDL receptors. The heparan sulfate proteoglycan pathway represents a novel catabolic mechanism that may allow substantial cellular and interstitial accumulation of cholesteryl ester-rich lipoproteins, independent of feedback inhibition by cellular sterol content. Topics: Animals; Apolipoprotein B-100; Apolipoproteins B; Carcinoma, Hepatocellular; Cattle; CHO Cells; Cricetinae; Cricetulus; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Humans; Iodine Radioisotopes; Lipoprotein Lipase; Lipoprotein(a); Lipoproteins; Lipoproteins, LDL; Polysaccharide-Lyases; Proteoglycans; Receptors, LDL; Tumor Cells, Cultured | 1992 |
Heparin and hormonal regulation of mRNA synthesis and abundance of autocrine growth factors: relevance to clonal growth of tumors.
Highly sulfated, heparinlike species of heparan sulfate proteoglycans, with heparinlike glycosaminoglycan chains, are extracellular matrix components that are plasma membrane bound in growth-arrested liver cells. Heparins were found to inhibit the growth and lower the clonal growth efficiency of HepG2, a minimally deviant, human hepatoma cell line. Heparan sulfates, closely related glycosaminoglycans present in the extracellular matrix around growing liver cells, had no effect on the growth rate or clonal growth efficiency of HepG2 cells. Neither heparins nor heparan sulfates had any effect on the growth rate or clonal growth efficiency of two poorly differentiated, highly metastatic hepatoma cell lines, SK-Hep-1 and PLC/PRF/5. Heparin's inhibition of growth of HepG2 cells correlated with changes in the mRNA synthesis and abundance of insulinlike growth factor II (IGF II) and transforming growth factor beta (TGF beta). HepG2 cells expressed high basal levels of mRNAs encoding IGF II and TGF beta that were inducible, through transcriptional and posttranscriptional mechanisms, to higher levels by specific heparin-hormone combinations. For both IGF II and TGF beta, the regulation was multifactorial. Transcriptionally, IGF II was regulated by the additive effects of insulin, glucagon, and growth hormone in combination with heparin; TGF beta was regulated primarily by the synergistic effects of insulin and growth hormone in combination with heparin. Posttranscriptionally, the mRNA abundance of the IGF II 4.5- and 3.7-kb transcripts was affected by insulin. Heparin induction of all IGF II transcripts was also dependent on triiodotyronine and prolactin, but it is unknown whether their induction by heparin was via transcriptional or posttranscriptional mechanisms. Heparin-insulin combinations regulated TGF beta posttranscriptionally. The poorly differentiated hepatoma cell lines PLC/PRF/5 and SK-Hep-1 either did not express or constitutively expressed low basal levels of IGF I, IGF II, and TGF beta, whose mRNA synthesis and abundance showed no response to any heparin-hormone combination. We discuss the data as evidence that matrix chemistry is a variable determining the expression of autocrine growth factor genes and the biological responses to them. Topics: Blotting, Northern; Carcinoma, Hepatocellular; Cell Differentiation; Cell Division; Clone Cells; Growth Substances; Heparin; Heparitin Sulfate; Hormones; Humans; In Vitro Techniques; Insulin-Like Growth Factor II; Liver Neoplasms; Neoplasm Metastasis; RNA, Messenger; RNA, Neoplasm; Transforming Growth Factor beta; Tumor Cells, Cultured | 1991 |
Biochemical composition and heterogeneity of heparan sulfates isolated from AH-130 ascites hepatoma cells and fluid.
The glycosaminoglycan composition of AH-130 ascites hepatoma cells and fluid were examined using enzymatic digestion, electrophoresis, and sequential partition fractionation. The cell-associated glycosaminoglycans were found to consist of 93% heparan sulfate, with the remainder consisting primarily of chondroitin sulfate. The glycosaminoglycans isolated from the ascitic fluid were found to consist of 58% heparan sulfate, 26% hyaluronic acid and 16% chondroitin sulfate. Dermatan sulfate was not detected in either cells or fluid. The heparan sulfate isolated from AH-130 cells in low-sulfate and highly heterogeneous with respect to biochemical composition. Fractions isolated by partition fractionation varied from 0.14 mol sulfate/mol uronic acid to 0.6 mol sulfate/mol uronic acid. Of the total sulfate 70--80% is N-sulfate in the former and 50% in the latter. Electrophoresis in 0.1 M HCl showed a highly heterogeneous material with mobility between that of hyaluronic acid and beef lung heparan sulfate. The heparan sulfate isolated from the fluid was similar to that isolated from the cells but was, however, somewhat more homogeneous with respect to charge. Topics: Animals; Ascitic Fluid; Carcinoma, Hepatocellular; Cattle; Glycosaminoglycans; Heparitin Sulfate; Hexosamines; Liver; Liver Neoplasms; Lung; Male; Rats; Sulfates; Uronic Acids | 1978 |
Release of glycopeptides and mucopolysaccharides from ascites hepatoma cells by tryptic treatment.
Two types of ascites hepatoma cells, AH 66 and AH 130 FN, were treated with trypsin to observe the release of complex carbohydrates constituting the plasma membranes. From AH 66 cells, mucopolysaccharide (heparan sulfate) was preferentially released. From AH 130 FN cells, N-glycosidic glycopeptides were preferentially released whereas no mucopolysaccharide (chondroitin sulfate A) was released. Topics: Animals; Carcinoma, Hepatocellular; Cells, Cultured; Female; Glucosamine; Glycopeptides; Glycosaminoglycans; Heparitin Sulfate; Liver Neoplasms; Rats; Sulfates; Trypsin | 1978 |
35S incorporation into sulfated mucopolysaccharides from embryonic, normal, and neoplastic liver cells.
The incorporation of 35S into the sulfated mucopolysaccharides from hepatoma, normal and embryonic liver cells has been studied by means of anion exchange chromatography of the isolated mucopolysaccharide. Comparatively to liver cells, while showing a higher labeling of low sulfated mucopolysaccharides the hepatoma cells incorporate much less 35S into the heparin fraction. Topics: Animals; Ascitic Fluid; Carcinoma, Hepatocellular; Chondroitin; Female; Glycosaminoglycans; Heparin; Heparitin Sulfate; Liver; Liver Neoplasms; Pregnancy; Rats; Sulfates | 1976 |
The glycosaminoglycans in human hepatic cancer.
A method is proposed for the analysis of glycosaminoglycans that were isolated from human liver, combining cellulose acetate electrophoresis and enzymatic digestion with mucopolysaccharidases. The major constituent of glycossaminoglycans in the healthy liver is heparin sulfate and/or heparin (about 65%), with approximately equal quantities of dermatan sulfate and hyalauronic acid (about 13.5 and 13%, respectively) and a small amount of chondroitin sulfate. These components, especially chondroitin sulfate and hyaluronic acid, are markedly increased in hepatic carcinomas. Topics: Adenoma, Bile Duct; Carcinoma, Hepatocellular; Chondroitin; Dermatan Sulfate; Electrophoresis; Glycosaminoglycans; Heparin; Heparitin Sulfate; Humans; Hyaluronic Acid; Hydrolysis; Liver; Liver Neoplasms | 1975 |
Stimulation of synthesis of free chondroitin sulfate chains by beta-D-xylosides in cultured cells.
Beta-Xylosides stimulate 2- to 6-fold the synthesis of glycosaminoglycans by three types of nonconnective tissue cells (RG-C6, NB41A, and rat hepatoma cells, and normal and simian virus 40 (SV40)-transformed normal human skin fibroblasts. The effect, which is specific for the anomeric linkage and the glycone, is observed in the presence and absence of puromycin. Beta-Xylosides may substitute for xylosylated core protein as initiators of synthesis of chondroitin sulfate chains. No stimulation of synthesis of heparan sulfate was observed. With the use of a fluorogenic xyloside, 4-methylumbelliferyl-beta-D-xyloside, it was demonstrated that the free chondroitin sulfate chains secreted into the medium bear the xyloside at the reducing end, and have an average molecular weight of 16,500. Topics: Animals; Carcinoma, Hepatocellular; Cells, Cultured; Chondroitin; Chromatography, Gel; Dermatan Sulfate; Fibroblasts; Galactose; Glycosaminoglycans; Glycosides; Heparitin Sulfate; Humans; Hyaluronic Acid; Hymecromone; Liver Neoplasms; Mice; Neuroblastoma; Neuroglia; Nitrophenols; Rats; Simian virus 40; Skin; Spectrometry, Fluorescence; Sulfur Radioisotopes; Sulfuric Acids; Tritium; Xylose | 1975 |
The isolation and characterization of glycopeptides and mucopolysaccharides from plasma membranes of an ascites hepatoma, AH 130.
Plasma membranes were isolated from an ascites hepatoma, AH 130, by the fluorescein mercuric acetate (FMA) method. Glycopeptides and mucopolysaccharides were prepared by digesting the membranes with pronase, then by fractionating the digest chromatographically and electrophoretically. Isolated fractions were analyzed for their amino acid and carbohydrate compositions. Results were compared with those for corresponding fractions from AH 66 (J. Biochem. 76, 319-333 (1974)). Mucopolysaccharides and a series of glycopeptides were isolated from the fraction excluded from Sephadex G-50. The mucopolysaccharides were identified as a family of heparan sulfates with different electrophoretic mobilities. The glycopeptides contained serine, threonine, galactose, galactosamine, glucosamine, and sialic acid as the major constituents as aspartic acid and mannose as minor ones. This suggests that most of the carbohydrate moieties are linked to serine or threonine (O-glycosidic), and that some are linked to asparagine (N-glycosidic). No nearly purely O-glycosidic glycopeptides were found in this fraction from AH 130, through they were the major glycopeptides from the AH 66 plasma membranes. In the fraction included in the gel, glycopeptides containing fucose, galactose, mannose, glucosamine, glaactosamine, and sialic acid were found. The presence of galactosamine suggests that some of the glycopeptides are O-glycosidic though most are N-glycosidic. In the corresponding fraction from AH 66, nearly purely N-glycosidic glycopeptides were found. Topics: Animals; Aspartic Acid; Carcinoma, Hepatocellular; Cell Membrane; Female; Glycopeptides; Glycosaminoglycans; Heparitin Sulfate; Hexosamines; Hexoses; Liver Neoplasms; Rats; Serine; Sialic Acids; Sulfuric Acid Esters; Threonine | 1975 |
The isolation and characterization of glycopeptides from plasma membranes of an ascites hepatoma, AH 66.
Topics: Amino Acids; Animals; Carcinoma, Hepatocellular; Cell Membrane; Centrifugation, Density Gradient; Chromatography, Gel; Chromatography, Ion Exchange; Electrophoresis; Female; Fucose; Galactosamine; Galactose; Glucosamine; Glucose; Glycopeptides; Heparitin Sulfate; Inositol; Liver Neoplasms; Mannose; Microscopy, Phase-Contrast; Molecular Weight; Protein Binding; Rats; Sialic Acids; Sulfur Radioisotopes; Tritium; Uronic Acids | 1974 |