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1.
FASEB J ; 38(10): e23670, 2024 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-38747803

RESUMO

HPSE2, the gene-encoding heparanase 2 (Hpa2), is mutated in urofacial syndrome (UFS), a rare autosomal recessive congenital disease attributed to peripheral neuropathy. Hpa2 lacks intrinsic heparan sulfate (HS)-degrading activity, the hallmark of heparanase (Hpa1), yet it exhibits a high affinity toward HS, thereby inhibiting Hpa1 enzymatic activity. Hpa2 regulates selected genes that promote normal differentiation, tissue homeostasis, and endoplasmic reticulum (ER) stress, resulting in antitumor, antiangiogenic, and anti-inflammatory effects. Importantly, stress conditions induce the expression of Hpa2, thus establishing a feedback loop, where Hpa2 enhances ER stress which, in turn, induces Hpa2 expression. In most cases, cancer patients who retain high levels of Hpa2 survive longer than patients bearing Hpa2-low tumors. Experimentally, overexpression of Hpa2 attenuates the growth of tumor xenografts, whereas Hpa2 gene silencing results in aggressive tumors. Studies applying conditional Hpa2 knockout (cHpa2-KO) mice revealed an essential involvement of Hpa2 contributed by the host in protecting against cancer and inflammation. This was best reflected by the distorted morphology of the Hpa2-null pancreas, including massive infiltration of immune cells, acinar to adipocyte trans-differentiation, and acinar to ductal metaplasia. Moreover, orthotopic inoculation of pancreatic ductal adenocarcinoma (PDAC) cells into the pancreas of Hpa2-null vs. wild-type mice yielded tumors that were by far more aggressive. Likewise, intravenous inoculation of cancer cells into cHpa2-KO mice resulted in a dramatically increased lung colonization reflecting the involvement of Hpa2 in restricting the formation of a premetastatic niche. Elucidating Hpa2 structure-activity-relationships is expected to support the development of Hpa2-based therapies against cancer and inflammation.


Assuntos
Glucuronidase , Inflamação , Neoplasias , Humanos , Animais , Inflamação/metabolismo , Inflamação/patologia , Neoplasias/patologia , Neoplasias/metabolismo , Neoplasias/genética , Glucuronidase/metabolismo , Glucuronidase/genética , Camundongos , Estresse do Retículo Endoplasmático
2.
Proc Natl Acad Sci U S A ; 119(31): e2203167119, 2022 08 02.
Artigo em Inglês | MEDLINE | ID: mdl-35881786

RESUMO

Heparan sulfate proteoglycans (HSPGs) mediate essential interactions throughout the extracellular matrix (ECM), providing signals that regulate cellular growth and development. Altered HSPG composition during tumorigenesis strongly aids cancer progression. Heparanase (HPSE) is the principal enzyme responsible for extracellular heparan sulfate catabolism and is markedly up-regulated in aggressive cancers. HPSE overactivity degrades HSPGs within the ECM, facilitating metastatic dissemination and releasing mitogens that drive cellular proliferation. Reducing extracellular HPSE activity reduces cancer growth, but few effective inhibitors are known, and none are clinically approved. Inspired by the natural glycosidase inhibitor cyclophellitol, we developed nanomolar mechanism-based, irreversible HPSE inhibitors that are effective within physiological environments. Application of cyclophellitol-derived HPSE inhibitors reduces cancer aggression in cellulo and significantly ameliorates murine metastasis. Mechanism-based irreversible HPSE inhibition is an unexplored anticancer strategy. We demonstrate the feasibility of such compounds to control pathological HPSE-driven malignancies.


Assuntos
Glucuronidase , Inibidores de Glicosídeo Hidrolases , Metástase Neoplásica , Animais , Proliferação de Células/efeitos dos fármacos , Glucuronidase/antagonistas & inibidores , Inibidores de Glicosídeo Hidrolases/farmacologia , Inibidores de Glicosídeo Hidrolases/uso terapêutico , Proteoglicanas de Heparan Sulfato/metabolismo , Heparitina Sulfato/metabolismo , Humanos , Camundongos , Metástase Neoplásica/tratamento farmacológico
3.
J Med Virol ; 96(5): e29630, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38659368

RESUMO

The human papillomavirus type 16 (HPV16) causes a large fraction of genital and oropharyngeal carcinomas. To maintain the transformed state, the tumor cells must continuously synthesize the E6 and E7 viral oncoproteins, which makes them tumor-specific antigens. Indeed, specific T cell responses against them have been well documented and CD8+ T cells engineered to express T cell receptors (TCRs) that recognize epitopes of E6 or E7 have been tested in clinical studies with promising results, yet with limited clinical success. Using CD8+ T cells from peripheral blood of healthy donors, we have identified two novel TCRs reactive to an unexplored E618-26 epitope. These TCRs showed limited standalone cytotoxicity against E618-26-HLA-A*02:01-presenting tumor cells. However, a single-signaling domain chimeric antigen receptor (ssdCAR) targeting L1CAM, a cell adhesion protein frequently overexpressed in HPV16-induced cancer, prompted a synergistic effect that significantly enhanced the cytotoxic capacity of NK-92/CD3/CD8 cells armored with both TCR and ssdCAR when both receptors simultaneously engaged their respective targets, as shown by live microscopy of 2-D and 3-D co-cultures. Thus, virus-specific TCRs from the CD8+ T cell repertoire of healthy donors can be combined with a suitable ssdCAR to enhance the cytotoxic capacity of the effector cells and, indirectly, their specificity.


Assuntos
Linfócitos T CD8-Positivos , Proteínas Oncogênicas Virais , Receptores de Antígenos de Linfócitos T , Receptores de Antígenos Quiméricos , Proteínas Repressoras , Humanos , Proteínas Oncogênicas Virais/imunologia , Proteínas Oncogênicas Virais/genética , Receptores de Antígenos Quiméricos/imunologia , Receptores de Antígenos Quiméricos/genética , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/genética , Proteínas Repressoras/imunologia , Proteínas Repressoras/genética , Linfócitos T CD8-Positivos/imunologia , Células Matadoras Naturais/imunologia , Papillomavirus Humano 16/imunologia , Papillomavirus Humano 16/genética , Citotoxicidade Imunológica , Linhagem Celular Tumoral
4.
Trends Biochem Sci ; 43(1): 18-31, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-29162390

RESUMO

Heparanase, the sole heparan sulfate (HS)-degrading endoglycosidase, regulates multiple biological activities that enhance tumor growth, metastasis, angiogenesis, and inflammation. Heparanase accomplishes this by degrading HS and thereby regulating the bioavailability of heparin-binding proteins; priming the tumor microenvironment; mediating tumor-host crosstalk; and inducing gene transcription, signaling pathways, exosome formation, and autophagy that together promote tumor cell performance and chemoresistance. By contrast, heparanase-2, a close homolog of heparanase, lacks enzymatic activity, inhibits heparanase activity, and regulates selected genes that promote normal differentiation, endoplasmic reticulum stress, tumor fibrosis, and apoptosis, together resulting in tumor suppression. The emerging premise is that heparanase is a master regulator of the aggressive phenotype of cancer, while heparanase-2 functions as a tumor suppressor.


Assuntos
Neoplasias/metabolismo , Neoplasias/patologia , Polissacarídeo-Liases/metabolismo , Animais , Progressão da Doença , Humanos
5.
Cell Mol Life Sci ; 78(6): 2771-2780, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33051777

RESUMO

Heparanase is the predominant enzyme that cleaves heparan sulfate, the main polysaccharide in the extracellular matrix. While the role of heparanase in sustaining the pathology of autoimmune diabetes is well documented, its association with metabolic syndrome/type 2 diabetes attracted less attention. Our research was undertaken to elucidate the significance of heparanase in impaired glucose metabolism in metabolic syndrome and early type 2 diabetes. Here, we report that heparanase exerts opposite effects in insulin-producing (i.e., islets) vs. insulin-target (i.e., skeletal muscle) compartments, sustaining or hampering proper regulation of glucose homeostasis depending on the site of action. We observed that the enzyme promotes macrophage infiltration into islets in a murine model of metabolic syndrome, and fosters ß-cell-damaging properties of macrophages activated in vitro by components of diabetogenic/obese milieu (i.e., fatty acids). On the other hand, in skeletal muscle (prototypic insulin-target tissue), heparanase is essential to ensure insulin sensitivity. Thus, despite a deleterious effect of heparanase on macrophage infiltration in islets, the enzyme appears to have beneficial role in glucose homeostasis in metabolic syndrome. The dichotomic action of the enzyme in the maintenance of glycemic control should be taken into account when considering heparanase-targeting strategies for the treatment of diabetes.


Assuntos
Glucuronidase/metabolismo , Síndrome Metabólica/patologia , Animais , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/patologia , Dieta Hiperlipídica , Modelos Animais de Doenças , Ácidos Graxos Insaturados/farmacologia , Teste de Tolerância a Glucose , Glucuronidase/genética , Resistência à Insulina , Células Secretoras de Insulina/citologia , Células Secretoras de Insulina/metabolismo , Interleucina-1beta/metabolismo , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/metabolismo , Masculino , Síndrome Metabólica/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Obesidade/metabolismo , Obesidade/patologia , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo
6.
Int J Mol Sci ; 23(9)2022 Apr 27.
Artigo em Inglês | MEDLINE | ID: mdl-35563215

RESUMO

Heparin is a polysaccharide expressed in animal connective tissue-type mast cells. Owing to the special pentasaccharide sequence, heparin specifically binds to antithrombin (AT) and increases the inhibitory activity of AT towards coagulation enzymes. Heparin isolated from porcine intestinal mucosa has an average molecular weight of 15 kDa, while heparins recovered from rat skin and the peritoneal cavity were 60-100 kDa and can be fragmented by the endo-glucuronidase heparanase in vitro. In this study, we have examined heparin isolated from in vitro matured fetal skin mast cells (FSMC) and peritoneal cavity mast cells (PCMC) collected from wildtype (WT), heparanase knockout (Hpa-KO), and heparanase overexpressing (Hpa-tg) mice. The metabolically 35S-labeled heparin products from the mast cells of WT, Hpa-KO, and Hpa-tg mice were compared and analyzed for molecular size and AT-binding activity. The results show that PCMC produced heparins with a size similar to heparin from porcine intestinal mast cells, whilst FSMC produced much longer chains. As expected, heparanase overexpression resulted in the generation of smaller fragments in both cell types, while heparins recovered from heparanase knockout cells were slightly longer than heparin from WT cells. Unexpectedly, we found that heparanase expression affected the production of total glycosaminoglycans (GAGs) and the proportion between heparin and other GAGs but essentially had no effect on heparin catabolism.


Assuntos
Glucuronidase , Mastócitos , Animais , Anticoagulantes/metabolismo , Antitrombinas/metabolismo , Glucuronidase/metabolismo , Glicosaminoglicanos/metabolismo , Heparina/química , Mastócitos/metabolismo , Camundongos , Ratos , Suínos
7.
Int J Mol Sci ; 23(20)2022 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-36293542

RESUMO

The kidney glomerular filtration barrier (GFB) is enriched with heparan sulfate (HS) proteoglycans, which contribute to its permselectivity. The endoglycosidase heparanase cleaves HS and hence appears to be involved in the pathogenesis of kidney injury and glomerulonephritis. We have recently reported, nonetheless, that heparanase overexpression preserved glomerular structure and kidney function in an experimental model of Adriamycin-induced nephropathy. To elucidate mechanisms underlying heparanase function in podocytes-key GFB cells, we utilized a human podocyte cell line and transgenic mice overexpressing heparanase. Notably, podocytes overexpressing heparanase (H) demonstrated significantly higher survival rates and viability after exposure to Adriamycin or hydrogen peroxide, compared with mock-infected (V) podocytes. Immunofluorescence staining of kidney cryo-sections and cultured H and V podocytes as well as immunoblotting of proteins extracted from cultured cells, revealed that exposure to toxic injury resulted in a significant increase in autophagic flux in H podocytes, which was reversed by the heparanase inhibitor, Roneparstat (SST0001). Heparanase overexpression was also associated with substantial transcriptional upregulation of autophagy genes BCN1, ATG5, and ATG12, following Adriamycin treatment. Moreover, cleaved caspase-3 was attenuated in H podocytes exposed to Adriamycin, indicating lower apoptotic cell death in H vs. V podocytes. Collectively, these findings suggest that in podocytes, elevated levels of heparanase promote cytoprotection.


Assuntos
Podócitos , Camundongos , Animais , Humanos , Podócitos/metabolismo , Doxorrubicina/toxicidade , Caspase 3/metabolismo , Peróxido de Hidrogênio/metabolismo , Glucuronidase/genética , Glucuronidase/metabolismo , Autofagia , Camundongos Transgênicos , Heparitina Sulfato/metabolismo , Proteoglicanas/metabolismo
8.
Molecules ; 27(20)2022 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-36296711

RESUMO

In this study, superparamagnetic iron oxide nanoparticles (SPIONs) were engineered with an organic coating composed of low molecular weight heparin (LMWH) and bovine serum albumin (BSA), providing heparin-based nanoparticle systems (LMWH@SPIONs). The purpose was to merge the properties of the heparin skeleton and an inorganic core to build up a targeted theranostic nanosystem, which was eventually enhanced by loading a chemotherapeutic agent. Iron oxide cores were prepared via the co-precipitation of iron salts in an alkaline environment and oleic acid (OA) capping. Dopamine (DA) was covalently linked to BSA and LMWH by amide linkages via carbodiimide coupling. The following ligand exchange reaction between the DA-BSA/DA-LMWH and OA was conducted in a biphasic system composed of water and hexane, affording LMWH@SPIONs stabilized in water by polystyrene sulfonate (PSS). Their size and morphology were investigated via dynamic light scattering (DLS) and transmission electron microscopy (TEM), respectively. The LMWH@SPIONs' cytotoxicity was tested, showing marginal or no toxicity for samples prepared with PSS at concentrations of 50 µg/mL. Their inhibitory activity on the heparanase enzyme was measured, showing an effective inhibition at concentrations comparable to G4000 (N-desulfo-N-acetyl heparin, a non-anticoagulant and antiheparanase heparin derivative; Roneparstat). The LMWH@SPION encapsulation of paclitaxel (PTX) enhanced the antitumor effect of this chemotherapeutic on breast cancer cells, likely due to an improved internalization of the nanoformulated drug with respect to the free molecule. Lastly, time-domain NMR (TD-NMR) experiments were conducted on LMWH@SPIONs obtaining relaxivity values within the same order of magnitude as currently used commercial contrast agents.


Assuntos
Nanopartículas de Magnetita , Nanopartículas , Nanopartículas de Magnetita/química , Soroalbumina Bovina , Hexanos , Meios de Contraste , Ácido Oleico , Medicina de Precisão , Ligantes , Heparina de Baixo Peso Molecular/farmacologia , Dopamina , Sais , Compostos Férricos/química , Nanopartículas/química , Heparina , Nanopartículas Magnéticas de Óxido de Ferro , Paclitaxel , Ferro , Água , Carbodi-Imidas , Amidas
9.
Semin Thromb Hemost ; 47(3): 240-253, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33794549

RESUMO

Cell surface proteoglycans are important constituents of the glycocalyx and participate in cell-cell and cell-extracellular matrix (ECM) interactions, enzyme activation and inhibition, and multiple signaling routes, thereby regulating cell proliferation, survival, adhesion, migration, and differentiation. Heparanase, the sole mammalian heparan sulfate degrading endoglycosidase, acts as an "activator" of HS proteoglycans, thus regulating tissue hemostasis. Heparanase is a multifaceted enzyme that together with heparan sulfate, primarily syndecan-1, drives signal transduction, immune cell activation, exosome formation, autophagy, and gene transcription via enzymatic and nonenzymatic activities. An important feature is the ability of heparanase to stimulate syndecan-1 shedding, thereby impacting cell behavior both locally and distally from its cell of origin. Heparanase releases a myriad of HS-bound growth factors, cytokines, and chemokines that are sequestered by heparan sulfate in the glycocalyx and ECM. Collectively, the heparan sulfate-heparanase axis plays pivotal roles in creating a permissive environment for cell proliferation, differentiation, and function, often resulting in the pathogenesis of diseases such as cancer, inflammation, endotheliitis, kidney dysfunction, tissue fibrosis, and viral infection.


Assuntos
Doença , Glucuronidase/metabolismo , Heparitina Sulfato/metabolismo , Humanos
10.
Anticancer Drugs ; 31(9): 885-889, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32304406

RESUMO

Heparanase is an endoglycosidase that degrades heparan sulfate side chains of heparan sulfate-proteoglycans. It liberates heparan sulfate-bound growth factors and thereby promotes blood vessel sprouting and angiogenesis. The subterranean blind mole rat, Spalax, is a wild mammal that lives most of its life in underground tunnels where it experiences sharp fluctuations in oxygen and carbon dioxide levels. We described two splice variants of heparanase from Spalax, Splice 7 and splice 36, both devoid of heparanase enzymatic activity. Splice 7 increases tumor growth, while splice 36 functions as a dominant negative to wild-type heparanase and decreases tumor growth and metastasis. Here, we describe two novel splice variants of Spalax heparanase, splice 67 and splice 612. These splice variants result in production of a shorter heparanase proteins that are similar to the wild-type native heparanase in their N-terminal but have unique C-terminals. Both splice 67 and 612 lack heparan sulfate degradation activity.


Assuntos
Glucuronidase/genética , Glucuronidase/metabolismo , Spalax/genética , Spalax/metabolismo , Sequência de Aminoácidos , Animais , Clonagem Molecular , Células HEK293 , Humanos , Isoenzimas , Transfecção
11.
Adv Exp Med Biol ; 1221: 3-59, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32274705

RESUMO

This review summarizes key developments in the heparanase field obtained 20 years prior to cloning of the HPSE gene and nearly 20 years after its cloning. Of the numerous publications and review articles focusing on heparanase, we have selected those that best reflect the progression in the field as well as those we regard important accomplishments with preference to studies performed by scientists and groups that contributed to this book. Apart from a general 'introduction' and 'concluding remarks', the abstracts of these studies are presented essentially as published along the years. We apologize for not being objective and not being able to include some of the most relevant abstracts and references, due to space limitation. Heparanase research can be divided into two eras. The first, initiated around 1975, dealt with identifying the enzyme, establishing the relevant assay systems and investigating its biological activities and significance in cancer and other pathologies. Studies performed during the first area are briefly introduced in a layman style followed by the relevant abstracts presented chronologically, essentially as appears in PubMed. The second era started in 1999 when the heparanase gene was independently cloned by 4 research groups [1-4]. As expected, cloning of the heparanase gene boosted heparanase research by virtue of the readily available recombinant enzyme, molecular probes, and anti-heparanase antibodies. Studies performed during the second area are briefly introduced followed by selected abstracts of key findings, arranged according to specific topics.


Assuntos
Glucuronidase/história , Pesquisa Translacional Biomédica/história , Glucuronidase/genética , História do Século XX , História do Século XXI , Humanos , Neoplasias
12.
Adv Exp Med Biol ; 1221: 231-249, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32274712

RESUMO

Single Nucleotide Polymorphisms (SNPs) is the substitution of a single nucleotide, stably inherited, highly abundant, and distributed throughout the genome. Up today 9746 SNPs were found in the HPSE gene. During 12 years 21 SNPs were analyzed in normal and pathological samples. The most prominent SNPs are rs4693608, rs11099592, rs4693084, and rs4364254. These SNPs were found in correlation with heparanase mRNA and protein expression among healthy persons. Moreover, an association of the HPSE gene SNPs with inflammatory processes, cancer development and progression was detected. SNP investigation allowed the identification of strong HPSE gene enhancer in the intron 2. In normal leukocytes, heparanase binds to the enhancer region and regulates HPSE gene expression via negative feedback in rs4693608 SNP-dependent manner. In malignant cells, heparanase halted self-regulation of the enhancer region. Instead of heparanase, the helicase-like transcription factor (HLTF) binds to the regulatory region. These and subsequent studies will elucidate how modification in the HPSE enhancer region could be applied to develop new approaches for cancer treatment.


Assuntos
Carcinogênese/genética , Glucuronidase/genética , Inflamação/genética , Neoplasias/genética , Neoplasias/patologia , Polimorfismo de Nucleotídeo Único , Progressão da Doença , Humanos , Íntrons , RNA Mensageiro
13.
Adv Exp Med Biol ; 1221: 253-283, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32274713

RESUMO

Two decades following the cloning of the heparanase gene, the significance of this enzyme for tumor growth and metastasis cannot be ignored. Compelling pre-clinical and clinical evidence tie heparanase with all steps of tumor formation namely, initiation, growth, metastasis, and chemo resistance, thus confirming and significantly expanding earlier observations that coupled heparanase activity with the metastatic capacity of tumor cells. This collective effort has turned heparanase from an obscure enzyme to a valid target for the development of anti-cancer drugs, and led basic researchers and biotech companies to develop heparanase inhibitors as anti-cancer therapeutics, some of which are currently examined clinically. As expected, the intense research effort devoted to understanding the biology of heparanase significantly expanded the functional repertoire of this enzyme, but some principle questions are still left unanswered or are controversial. For example, many publications describe increased heparanase levels in human tumors, but the mechanism underlying heparanase induction is not sufficiently understood. Moreover, heparanase is hardly found to be increased in many studies utilizing methodologies (i.e., gene arrays) that compare tumors vs (adjacent) normal tissue. The finding that heparanase exert also enzymatic activity-independent function significantly expands the mode by which heparanase can function outside, but also inside the cell. Signaling aspects, and a role of heparanase in modulating autophagy are possibly as important as its enzymatic aspect, but these properties are not targeted by heparanase inhibitors, possibly compromising their efficacy. This Book chapter review heparanase function in oncology, suggesting a somewhat different interpretation of the results.


Assuntos
Glucuronidase/metabolismo , Neoplasias , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Glucuronidase/antagonistas & inibidores , Humanos , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Transdução de Sinais
14.
J Mol Cell Cardiol ; 131: 29-40, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31004678

RESUMO

Although cancer cells use heparanase for tumor metastasis, favourable effects of heparanase have been reported in the management of Alzheimer's disease and diabetes. Indeed, we previously established a protective function for heparanase in the acutely diabetic heart, where it conferred cardiomyocyte resistance to oxidative stress and apoptosis by provoking changes in gene expression. In this study, we tested if overexpression of heparanase can protect the heart against chemically induced or ischemia/reperfusion (I/R) injury. Transcriptomic analysis of Hep-tg hearts reveal that 240 genes related to the stress response, immune response, cell death, and development were altered in a pro-survival direction encompassing genes promoting the unfolded protein response (UPR) and autophagy, as well as those protecting against oxidative stress. The observed UPR activation was adaptive and not apoptotic, was mediated by activation of ATF6α, and when combined with mTOR inhibition, induced autophagy. Subjecting wild type (WT) mice to increasing concentrations of the ER stress inducer thapsigargin evoked a transition from adaptive to apoptotic UPR, an effect that was attenuated in Hep-tg mouse hearts. Consistent with these observations, when exposed to I/R, the infarct size and markers of apoptosis were significantly lower in the Hep-tg heart compared to WT. Finally, UPR and autophagy inhibitors reduced the protective effects of heparanase overexpression during I/R. Our data suggest that the mechanisms that underlie the role of heparanase in promoting cell survival could be uniquely beneficial to the heart by providing protection against cellular stresses, and could be useful for exploitation as a therapeutic target for the treatment of heart disease.


Assuntos
Glucuronidase/metabolismo , Traumatismo por Reperfusão Miocárdica/metabolismo , Miocárdio/metabolismo , Miócitos Cardíacos/metabolismo , Substâncias Protetoras/metabolismo , Animais , Apoptose/fisiologia , Autofagia/fisiologia , Sobrevivência Celular/fisiologia , Coração/fisiologia , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ratos , Ratos Wistar , Tapsigargina/metabolismo , Resposta a Proteínas não Dobradas/fisiologia
15.
Int J Cancer ; 145(6): 1596-1608, 2019 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-31032901

RESUMO

Heparanase is an endo-ß-d-glucuronidase that cleaves heparan sulfate (HS) side chains of heparan sulfate proteoglycans. Compelling evidence tie heparanase levels with all steps of tumor formation including tumor initiation, growth, metastasis and chemo-resistance, likely involving augmentation of signaling pathways and gene transcription. In order to reveal the molecular mechanism(s) underlying the protumorigenic properties of heparanase, we established an inducible (Tet-on) system in U87 human glioma cells and applied gene array methodology in order to identify genes associated with heparanase induction. We found that CD24, a mucin-like cell adhesion protein, is consistently upregulated by heparanase and by heparanase splice variant devoid of enzymatic activity, whereas heparanase gene silencing was associated with decreased CD24 expression. This finding was further substantiated by a similar pattern of heparanase and CD24 immunostaining in glioma patients (Pearson's correlation; R = 0.66, p = 0.00001). Noteworthy, overexpression of CD24 stimulated glioma cell migration, invasion, colony formation in soft agar and tumor growth in mice suggesting that CD24 functions promote tumor growth. Likewise, anti-CD24 neutralizing monoclonal antibody attenuated glioma tumor growth, and a similar inhibition was observed in mice treated with a neutralizing mAb directed against L1 cell adhesion molecule (L1CAM), a ligand for CD24. Importantly, significant shorter patient survival was found in heparanase-high/CD24-high tumors vs. heparanase-high/CD24-low tumors for both high-grade and low-grade glioma (p = 0.02). Our results thus uncover a novel heparanase-CD24-L1CAM axis that plays a significant role in glioma tumorigenesis.


Assuntos
Neoplasias Encefálicas/patologia , Antígeno CD24/metabolismo , Glioma/patologia , Glucuronidase/metabolismo , Animais , Neoplasias Encefálicas/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Progressão da Doença , Feminino , Glioma/metabolismo , Xenoenxertos , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Molécula L1 de Adesão de Célula Nervosa/metabolismo , Transdução de Sinais
16.
Proc Natl Acad Sci U S A ; 113(48): E7808-E7817, 2016 11 29.
Artigo em Inglês | MEDLINE | ID: mdl-27849593

RESUMO

The emerging role of heparanase in tumor initiation, growth, metastasis, and chemoresistance is well recognized and is encouraging the development of heparanase inhibitors as anticancer drugs. Unlike the function of heparanase in cancer cells, very little attention has been given to heparanase contributed by cells composing the tumor microenvironment. Here we used a genetic approach and examined the behavior and function of macrophages isolated from wild-type (WT) and heparanase-knockout (Hpa-KO) mice. Hpa-KO macrophages express lower levels of cytokines (e.g., TNFα, IL1-ß) and exhibit lower motility and phagocytic capacities. Intriguingly, inoculation of control monocytes together with Lewis lung carcinoma (LLC) cells into Hpa-KO mice resulted in nearly complete inhibition of tumor growth. In striking contrast, inoculating LLC cells together with monocytes isolated from Hpa-KO mice did not affect tumor growth, indicating that heparanase is critically required for activation and function of macrophages. Mechanistically, we describe a linear cascade by which heparanase activates Erk, p38, and JNK signaling in macrophages, leading to increased c-Fos levels and induction of cytokine expression in a manner that apparently does not require heparanase enzymatic activity. These results identify heparanase as a key mediator of macrophage activation and function in tumorigenesis and cross-talk with the tumor microenvironment.


Assuntos
Carcinoma Pulmonar de Lewis/enzimologia , Glucuronidase/fisiologia , Ativação de Macrófagos , Macrófagos/enzimologia , Animais , Carcinoma Pulmonar de Lewis/imunologia , Carcinoma Pulmonar de Lewis/patologia , Linhagem Celular , Citocinas/metabolismo , Regulação Neoplásica da Expressão Gênica , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transplante de Neoplasias , Transdução de Sinais , Fatores de Transcrição/metabolismo , Ativação Transcricional , Carga Tumoral , Microambiente Tumoral
17.
Proc Natl Acad Sci U S A ; 113(3): 704-9, 2016 Jan 19.
Artigo em Inglês | MEDLINE | ID: mdl-26729870

RESUMO

Heparanase is an endoglycosidase that cleaves heparan sulfate side chains of proteoglycans, resulting in disassembly of the extracellular matrix underlying endothelial and epithelial cells and associating with enhanced cell invasion and metastasis. Heparanase expression is induced in carcinomas and sarcomas, often associating with enhanced tumor metastasis and poor prognosis. In contrast, the function of heparanase in hematological malignancies (except myeloma) was not investigated in depth. Here, we provide evidence that heparanase is expressed by human follicular and diffused non-Hodgkin's B-lymphomas, and that heparanase inhibitors restrain the growth of tumor xenografts produced by lymphoma cell lines. Furthermore, we describe, for the first time to our knowledge, the development and characterization of heparanase-neutralizing monoclonal antibodies that inhibit cell invasion and tumor metastasis, the hallmark of heparanase activity. Using luciferase-labeled Raji lymphoma cells, we show that the heparanase-neutralizing monoclonal antibodies profoundly inhibit tumor load in the mouse bones, associating with reduced cell proliferation and angiogenesis. Notably, we found that Raji cells lack intrinsic heparanase activity, but tumor xenografts produced by this cell line exhibit typical heparanase activity, likely contributed by host cells composing the tumor microenvironment. Thus, the neutralizing monoclonal antibodies attenuate lymphoma growth by targeting heparanase in the tumor microenvironment.


Assuntos
Anticorpos Neutralizantes/farmacologia , Anticorpos Neutralizantes/uso terapêutico , Glucuronidase/imunologia , Linfoma/patologia , Animais , Anticorpos Monoclonais/farmacologia , Proliferação de Células/efeitos dos fármacos , Glucuronidase/isolamento & purificação , Células HEK293 , Humanos , Luciferases/metabolismo , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos NOD , Camundongos SCID , Peso Molecular , Metástase Neoplásica , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/farmacologia , Proteínas Recombinantes/uso terapêutico , Saponinas/farmacologia , Carga Tumoral/efeitos dos fármacos , Microambiente Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
18.
Glycobiology ; 28(5): 269-275, 2018 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-29471321

RESUMO

Heparanase, an endo-glucuronidase that specifically cleaves heparan sulfate (HS), is upregulated in several pathological conditions. In this study, we aimed to find a correlation of heparanase expression and platelets production. In the transgenic mice overexpressing human heparanase (Hpa-tg), hematological analysis of blood samples revealed a significantly higher number of platelets in comparison with wild-type (Ctr) mice, while no significant difference was found in leukocytes and red blood cell number between the two groups. Total number of thiazole orange positive platelets was increased in Hpa-tg vs. Ctr blood, reflecting a higher rate of platelets production. Concomitantly, megakaryocytes from Hpa-tg mice produced more and shorter HS fragments that were shed into the medium. Further, thrombopoietin (TPO) level was elevated in the liver and plasma of Hpa-tg mice. Together, the data indicate that heparanase expression promoted megakaryopoiesis, which may be through upregulated expression of TPO and direct effect of released HS fragments expressed in the megakaryocytes.


Assuntos
Glucuronidase/genética , Megacariócitos/metabolismo , Animais , Células Cultivadas , Perfilação da Expressão Gênica , Glucuronidase/metabolismo , Megacariócitos/citologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos
19.
Am J Physiol Heart Circ Physiol ; 314(1): H82-H94, 2018 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-28986359

RESUMO

In the diabetic heart, there is excessive dependence on fatty acid (FA) utilization to generate ATP. Lipoprotein lipase (LPL)-mediated hydrolysis of circulating triglycerides is suggested to be the predominant source of FA for cardiac utilization during diabetes. In the heart, the majority of LPL is synthesized in cardiomyocytes and secreted onto cell surface heparan sulfate proteoglycan (HSPG), where an endothelial cell (EC)-releasable ß-endoglycosidase, heparanase cleaves the side chains of HSPG to liberate LPL for its onward movement across the EC. EC glycosylphosphatidylinositol-anchored high-density lipoprotein-binding protein 1 (GPIHBP1) captures this released enzyme at its basolateral side and shuttles it across to its luminal side. We tested whether the diabetes-induced increase of transforming growth factor-ß (TGF-ß) can influence the myocyte and EC to help transfer LPL to the vascular lumen to generate triglyceride-FA. In response to high glucose and EC heparanase secretion, this endoglycosidase is taken up by the cardiomyocyte (Wang Y, Chiu AP, Neumaier K, Wang F, Zhang D, Hussein B, Lal N, Wan A, Liu G, Vlodavsky I, Rodrigues B. Diabetes 63: 2643-2655, 2014) to stimulate matrix metalloproteinase-9 expression and the conversion of latent to active TGF-ß. In the cardiomyocyte, TGF-ß activation of RhoA enhances actin cytoskeleton rearrangement to promote LPL trafficking and secretion onto cell surface HSPG. In the EC, TGF-ß signaling promotes mesodermal homeobox 2 translocation to the nucleus, which increases the expression of GPIHBP1, which facilitates movement of LPL to the vascular lumen. Collectively, our data suggest that in the diabetic heart, TGF-ß actions on the cardiomyocyte promotes movement of LPL, whereas its action on the EC facilitates LPL shuttling. NEW & NOTEWORTHY Endothelial cells, as first responders to hyperglycemia, release heparanase, whose subsequent uptake by cardiomyocytes amplifies matrix metalloproteinase-9 expression and activation of transforming growth factor-ß. Transforming growth factor-ß increases lipoprotein lipase secretion from cardiomyocytes and promotes mesodermal homeobox 2 to enhance glycosylphosphatidylinositol-anchored high-density lipoprotein-binding protein 1-dependent transfer of lipoprotein lipase across endothelial cells, mechanisms that accelerate fatty acid utilization by the diabetic heart.


Assuntos
Glicemia/metabolismo , Diabetes Mellitus Experimental/enzimologia , Cardiomiopatias Diabéticas/enzimologia , Células Endoteliais/enzimologia , Metabolismo Energético , Ácidos Graxos/metabolismo , Lipase Lipoproteica/metabolismo , Miócitos Cardíacos/enzimologia , Animais , Comunicação Celular , Células Cultivadas , Diabetes Mellitus Experimental/sangue , Diabetes Mellitus Experimental/fisiopatologia , Cardiomiopatias Diabéticas/sangue , Cardiomiopatias Diabéticas/fisiopatologia , Glucuronidase/metabolismo , Proteínas de Homeodomínio/metabolismo , Masculino , Metaloproteinase 9 da Matriz/metabolismo , Proteínas Musculares/metabolismo , Ratos Wistar , Receptores de Lipoproteínas/metabolismo , Transdução de Sinais , Fator de Crescimento Transformador beta/metabolismo
20.
Hum Mol Genet ; 24(7): 1991-9, 2015 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-25510506

RESUMO

Urofacial syndrome (UFS) is an autosomal recessive disease with severe dysfunctional urination including urinary incontinence (UI). Biallelic mutations of HPSE2 are discovered from UFS patients, suggesting that HPSE2 is a candidate disease gene. Here, we show that deletion of Hpse2 is sufficient to cause the UFS-like phenotype in mice. Hpse2 knockout mutants display a distended bladder (megacystis) phenotype and abnormal voiding behavior similar to that found in patients. While Hpse2 is largely dispensable for detrusor smooth muscle and urothelial cell fate determination, the mutants have significantly lower rates of cell proliferation than wild-type littermate controls. All Hpse2 mutants have a growth retardation phenotype and die within a month after birth. Comprehensive blood chemistry and urinalysis indicate that Hpse2 mutants have renal dysfunction and malnutrition. We provide evidence that transforming growth factor beta (Tgfß) signaling is attenuated at birth. However, Tgfß activity is significantly enhanced at later stages when the urological phenotype is severe, and the mutant bladders have accumulated excessive amount of fibrotic tissue. Together, these findings strongly suggest that Hpse2 is a causative gene of human UFS and further uncover unexpected roles of Hpse2 in bladder physiology, tissue remodeling and Tgfß signaling.


Assuntos
Modelos Animais de Doenças , Glucuronidase/genética , Camundongos , Doenças Urológicas/enzimologia , Doenças Urológicas/genética , Animais , Fácies , Feminino , Deleção de Genes , Glucuronidase/metabolismo , Humanos , Masculino , Camundongos Knockout , Fenótipo , Transdução de Sinais , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo , Doenças Urológicas/metabolismo , Doenças Urológicas/patologia
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