RESUMEN
Acquired or inherited genetic alterations either alone or in combination with epigenetic alterations are associated with prostate carcinogenesis and its progression toward advance metastatic or castration-resistant disease. A major objective of translational cancer research in post-genome era is to discover the repertoire of genetic and epigenetic variations associated with prostate cancer. Genome-wide association studies have been at least partially successful in identifying potential germline polymorphisms and allelic imbalances such as microsatellite instability and loss of heterozygosity associated with prostate cancer susceptibility. Epigenetic mechanisms such as DNA hyper- or hypomethylation and histone modifications are reversible genetic alterations which allow stable inheritance of cellular phenotypes without any changes in the DNA sequence or quantity. Epigenetic modifications can potentially be used for the molecular classification, detection, and risk assessment in prostate cancer. Chemical inhibitors of DNA methyltransferases and histone deacetylases have been used in different clinical trials and hold promise as novel chemotherapeutics to be effective alone or in combination with other therapeutic interventions in prostate cancer.
RESUMEN
Recent studies have introduced prosaposin (PSAP) as a pleiotrophic growth factor for prostate cancer (PCa). We have previously reported that PSAP or one of its known active molecular derivatives, saposin C functions as an androgen-agonist and androgen-regulated gene (ARG) for androgen-sensitive (AS) PCa cell lines. Due to the potential significance of androgen receptor (AR)-expressing stroma in PCa, we evaluated a possible bi-directional paracrine regulatory interactions between DHT and PSAP in AR-positive prostate stromal (PrSt) cells. We report that saposin C in a ligand-independent manner increased AR expression, its nuclear content, and tyrosine phosphorylation. DHT treatment of PrSt cells increased PSAP expression. We also demonstrated both serum- and androgen-inducibility of a previously characterized hormone-responsive element (HRE) located in the proximal region of PSAP promoter. In addition, conditioned-media derived from PrSt cells and bone fibroblasts (i.e., MSF) differentially increased PSAP-promoter activity in androgen-independent (AI) PC-3 and AS LNCaP cells. Our data for the first time demonstrate that not only saposin C or PSAP regulates AR expression/activity, but also function as an ARG in PrSt. Ligand-independent activation of AR by PSAP or saposin C in PCa and stromal cells may contribute not only to prostate carcinogenesis at an early stage, but also in AI progression of the disease in an androgen-deprived tumor microenvironment.
Asunto(s)
Factores de Crecimiento Nervioso/química , Próstata/citología , Receptores Androgénicos/metabolismo , Saposinas/química , Saposinas/genética , Células del Estroma/metabolismo , Regulación hacia Arriba/genética , Andrógenos/farmacología , Animales , Huesos/citología , Línea Celular , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Medios de Cultivo Condicionados , Dihidrotestosterona/farmacología , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Humanos , Masculino , Ratones , Fosfotirosina/metabolismo , Neoplasias de la Próstata/metabolismo , Estructura Terciaria de Proteína , Elementos de Respuesta/genética , Saposinas/metabolismo , Suero , Células del Estroma/efectos de los fármacos , Transcripción Genética/efectos de los fármacos , Regulación hacia Arriba/efectos de los fármacosRESUMEN
Androgen-regulated genes (ARG) are implicated in normal and neoplastic growth of the prostate. Recently, we reported genomic amplification and/or overexpression of a previously known neurotrophic factor, prosaposin, in androgen-independent (AI) or metastatic prostate cancer (PCa) cells and tissues. Prosaposin and/or its known active molecular derivatives (e.g., saposin C) function as a pluripotent growth factor with diverse biological activities that favor malignant phenotypes in PCa cells. In addition, prosaposin or saposin C upregulates androgen receptor (AR) and AR-target genes (i.e., prostate-specific antigen, Probasin) expression and activity in LNCaP cells. Here, we examined prosaposin as an ARG. We report that DHT treatment of LNCaP cells increases prosaposin expression. In addition, we demonstrate androgen-responsiveness of prosaposin promoter and AR occupancy to a hormone-responsive element located in the proximal region of the prosaposin promoter. Our data for the first time identify prosaposin as an ARG. This observation, together with the pleiotropic growth factor activity of prosaposin, might suggest a role for this molecule in AR-dependent progression of prostate cancer at its early or late AI-state.
Asunto(s)
Andrógenos/farmacología , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Saposinas/genética , Western Blotting , Línea Celular Tumoral , Dihidrotestosterona/farmacología , Humanos , Factor I del Crecimiento Similar a la Insulina/farmacología , Interleucina-6/farmacología , Luciferasas/genética , Luciferasas/metabolismo , Masculino , Plásmidos/genética , Regiones Promotoras Genéticas/genética , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/patología , Unión Proteica/efectos de los fármacos , Receptores Androgénicos/genética , Receptores Androgénicos/metabolismo , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Saposinas/metabolismo , Transducción de Señal/efectos de los fármacos , TransfecciónRESUMEN
Lethal factor is a protease, one component of Bacillus anthracis exotoxin, which cleaves many of the mitogen-activated protein kinase kinases (MEKs). Given the importance of MEK signaling in tumorigenesis, we assessed the effects of anthrax lethal toxin (LeTx) on tumor cells. LeTx was very effective in inhibiting mitogen-activated protein kinase activation in V12 H-ras-transformed NIH 3T3 cells. In vitro, treatment of transformed cells with LeTx caused them to revert to a nontransformed morphology, and inhibited their abilities to form colonies in soft agar and to invade Matrigel without markedly affecting cell proliferation. In vivo, LeTx inhibited growth of ras-transformed cells implanted in athymic nude mice (in some cases causing tumor regression) at concentrations that caused no apparent animal toxicity. Unexpectedly, LeTx also greatly decreased tumor neovascularization. These results demonstrate that LeTx potently inhibits ras-mediated tumor growth and is a potential antitumor therapeutic.
Asunto(s)
Antígenos Bacterianos , Toxinas Bacterianas/farmacología , Transformación Celular Neoplásica/efectos de los fármacos , Quinasas de Proteína Quinasa Activadas por Mitógenos/antagonistas & inhibidores , Neovascularización Fisiológica/efectos de los fármacos , Células 3T3 , Animales , Toxinas Bacterianas/uso terapéutico , Pruebas de Carcinogenicidad , División Celular/efectos de los fármacos , Línea Celular Transformada , Modelos Animales de Enfermedad , Ratones , Ratones Desnudos , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Invasividad Neoplásica , Trasplante de Neoplasias , Neoplasias Experimentales/prevención & control , Péptido Hidrolasas/efectos de los fármacos , Péptido Hidrolasas/metabolismo , Proteínas ras/fisiologíaRESUMEN
We have mapped and characterized the human homolog of Drm/Gremlin (CKTFS1B1), a member of a family of BMP antagonists that have been linked to both developmental and transformation-related functions. By screening a human cDNA library, we isolated a 3.3-kb cDNA containing the 552-bp region encoding the human DRM protein. CKTFS1B1 was localized on human chromosome 15q13--> q15 by somatic cell hybrid analysis and, more precisely, using radiation hybrids, to a region of markers linked to SGNE1, secretory granule neuroendocrine protein 1 and RYR3, the ryanodyne receptor 3. Northern blot analysis showed the presence of a single DRM-specific mRNA expressed in different human tissues, including brain, ovary, intestine and colon. In the brain, DRM expression is associated with the region localized around the internal capsule in the large subcortical nuclei. DRM appears to be predominantly expressed in normal cells and tissues, including normal neurons, astrocytes and fibroblasts. Interestingly, we detected DRM expression in normal cells obtained from several patients, but not in tumor cell lines established from the same patients. The data suggest that down-regulation of DRM is associated with tumor progression, and support the hypothesis that human DRM may play an important role during both neuroembryological development and carcinogenesis.
Asunto(s)
Encéfalo/embriología , Encéfalo/metabolismo , Cromosomas Humanos Par 15/genética , Péptidos y Proteínas de Señalización Intercelular , Proteínas/genética , Envejecimiento/genética , Envejecimiento/metabolismo , Proteínas Morfogenéticas Óseas/metabolismo , Encéfalo/citología , Encéfalo/crecimiento & desarrollo , Línea Celular , Clonación Molecular , Perfilación de la Expresión Génica , Ligamiento Genético/genética , Humanos , Células Híbridas , Datos de Secuencia Molecular , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patología , Mapeo Físico de Cromosoma , Proteínas/metabolismo , ARN Mensajero/análisis , ARN Mensajero/genética , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Células Tumorales CultivadasRESUMEN
The Met receptor tyrosine kinase and its ligand, hepatocyte growth factor/scatter factor (HGF/SF), have been implicated in human tumor development and metastasis. HGF/SF induces the expression of urokinase plasminogen activator (uPA) and the uPA receptor (uPAR), important mediators of cell invasion and metastasis. We have developed a cell-based assay to screen for inhibitors of this signaling system using the induction of endogenous uPA and uPAR and the subsequent conversion of plasminogen to plasmin as the biological end point. Assay validation was established using a neutralizing antiserum to HGF/SF and a uPA inhibitor (B428), as well as inhibitors of the MKK-MAPK1/2 pathway, shown previously to be important in the induction of uPA and uPAR. Using this assay, we found several classes of molecules that exhibited inhibition of HGF/SF-dependent plasmin activation. However, we discovered that certain members of the geldanamycin family of anisamycin antibiotics are potent inhibitors of HGF/SF-mediated plasmin activation, displaying inhibitory properties at femtomolar concentrations and nine orders of magnitude below their growth inhibitory concentrations. At nanomolar concentrations, the geldanamycins down-regulate Met protein expression, inhibit HGF/SF-mediated cell motility and invasion, and also revert the phenotype of both autocrine HGF/SF-Met transformed cells as well as those transformed by Met proteins with activating mutations. Thus, the geldanamycins may have important therapeutic potential for the treatment of cancers in which Met activity contributes to the invasive/metastatic phenotype.
Asunto(s)
Antibióticos Antineoplásicos/toxicidad , Fibrinolisina/metabolismo , Factor de Crecimiento de Hepatocito/metabolismo , Proteínas Proto-Oncogénicas c-met/fisiología , Quinonas/toxicidad , Receptores de Superficie Celular/metabolismo , Activador de Plasminógeno de Tipo Uroquinasa/metabolismo , Células 3T3 , Animales , Benzoquinonas , División Celular/efectos de los fármacos , Línea Celular , Línea Celular Transformada , Humanos , Lactamas Macrocíclicas , Ratones , Receptores del Activador de Plasminógeno Tipo Uroquinasa , Proteínas Recombinantes/metabolismo , Transducción de Señal , Relación Estructura-Actividad , Transfección , Células Tumorales CultivadasRESUMEN
Loss of function in the von Hippel-Lindau (VHL) tumor suppressor gene occurs in familial and most sporadic renal cell carcinomas (RCCs). VHL has been linked to the regulation of cell cycle cessation (G(0)) and to control of expression of various mRNAs such as for vascular endothelial growth factor. RCC cells express the Met receptor tyrosine kinase, and Met mediates invasion and branching morphogenesis in many cell types in response to hepatocyte growth factor/scatter factor (HGF/SF). We examined the HGF/SF responsiveness of RCC cells containing endogenous mutated (mut) forms of the VHL protein (VHL-negative RCC) with that of isogenic cells expressing exogenous wild-type (wt) VHL (VHL-positive RCC). We found that VHL-negative 786-0 and UOK-101 RCC cells were highly invasive through growth factor-reduced (GFR) Matrigel-coated filters and exhibited an extensive branching morphogenesis phenotype in response to HGF/SF in the three-dimensional (3D) GFR Matrigel cultures. In contrast, the phenotypes of A498 VHL-negative RCC cells were weaker, and isogenic RCC cells ectopically expressing wt VHL did not respond at all. We found that all VHL-negative RCC cells expressed reduced levels of tissue inhibitor of metalloproteinase 2 (TIMP-2) relative to the wt VHL-positive cells, implicating VHL in the regulation of this molecule. However, consistent with the more invasive phenotype of the 786-0 and UOK-101 VHL-negative RCC cells, the levels of TIMP-1 and TIMP-2 were reduced and levels of the matrix metalloproteinases 2 and 9 were elevated compared to the noninvasive VHL-positive RCC cells. Moreover, recombinant TIMPs completely blocked HGF/SF-mediated branching morphogenesis, while neutralizing antibodies to the TIMPs stimulated HGF/SF-mediated invasion in vitro. Thus, the loss of the VHL tumor suppressor gene is central to changes that control tissue invasiveness, and a more invasive phenotype requires additional genetic changes seen in some but not all RCC lines. These studies also demonstrate a synergy between the loss of VHL function and Met signaling.
Asunto(s)
Carcinoma de Células Renales/genética , Genes Supresores de Tumor , Factor de Crecimiento de Hepatocito/farmacología , Neoplasias Renales/genética , Ligasas , Proteínas/genética , Proteínas Supresoras de Tumor , Ubiquitina-Proteína Ligasas , Enfermedad de von Hippel-Lindau/genética , Carcinoma de Células Renales/patología , Carcinoma de Células Renales/fisiopatología , Endopeptidasas/metabolismo , Espacio Extracelular/enzimología , Expresión Génica , Factor de Crecimiento de Hepatocito/antagonistas & inhibidores , Factor de Crecimiento de Hepatocito/fisiología , Humanos , Neoplasias Renales/patología , Neoplasias Renales/fisiopatología , Invasividad Neoplásica , Fenotipo , Proteínas Tirosina Quinasas Receptoras/metabolismo , Inhibidor Tisular de Metaloproteinasa-1/genética , Inhibidor Tisular de Metaloproteinasa-1/metabolismo , Inhibidor Tisular de Metaloproteinasa-2/genética , Inhibidor Tisular de Metaloproteinasa-2/metabolismo , Células Tumorales Cultivadas , Proteína Supresora de Tumores del Síndrome de Von Hippel-LindauRESUMEN
The Met tyrosine kinase receptor has been implicated in human cancer. Here we have examined the signaling requirements of three oncogenic forms of this molecule: wild type Met in response to ligand/autocrine stimulation, Met which has been mutationally activated, and Tpr-Met (a constitutively active truncated Met fusion protein). Previous studies have demonstrated the importance of a Grb2 binding site, and of specific tyrosine residues (i.e. Y8,9 and Y14,15) for Met function, and we have now explored the relevance of these and other sites for oncogenic Met signaling. Following substitution of various intracellular tyrosines for phenylalanine, we find that the transforming activity of each Met oncogene is dependent upon tyrosines Y8,9 and Y14,15, in addition to two novel tyrosines (Y6 and Y10) not previously implicated in Met signaling. Tyrosines Y6 and Y10 influence a variety of Met-mediated responses both in vitro (transformation, mitogenicity and invasion), and in vivo (tumorigenicity and metastasis). We also show that Tpr-Met is much more dependent on its Grb2 binding site for biological activity than are the other oncogenic forms of the Met receptor. Thus, although the three Met oncogenes examined are similar in their dependency on a number of specific tyrosines for activity, the signaling strategy employed by Tpr-Met can be differentiated from that of the other two.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales , Sustitución de Aminoácidos , Transformación Celular Neoplásica/genética , Oncogenes , Proteínas Proto-Oncogénicas c-met/fisiología , Transducción de Señal , Células 3T3 , Animales , Sitios de Unión , Femenino , Proteína Adaptadora GRB2 , Ratones , Ratones Desnudos , Modelos Moleculares , Mutagénesis Sitio-Dirigida , Metástasis de la Neoplasia , Fenilalanina/química , Mutación Puntual , Proteínas/metabolismo , Proteínas Proto-Oncogénicas c-met/química , Proteínas Proto-Oncogénicas c-met/genética , Proteínas Recombinantes de Fusión/fisiología , Eliminación de Secuencia , Relación Estructura-Actividad , Tirosina/químicaRESUMEN
Mutations in Met have been identified in human papillary renal carcinomas. We have shown previously that these mutations deregulate the enzymatic activity of Met and that NIH 3T3 cells expressing mutationally activated Met are transformed in vitro and are tumorigenic in vivo. In the present investigation, we find that mutant Met induces the motility of Madin-Darby canine kidney cells in vitro and experimental metastasis of NIH 3T3 cells in vivo, and that the Ras-Raf-MEK-ERK signaling pathway, which has been implicated previously in cellular motility and metastasis, is constitutively activated by the Met mutants. We also report that transgenic mice harboring mutationally activated Met develop metastatic mammary carcinoma. These data confirm the tumorigenic activity of mutant Met molecules and demonstrate their ability to induce the metastatic phenotype.
Asunto(s)
Transformación Celular Neoplásica/genética , Neoplasias Mamarias Experimentales/patología , Proteínas Proto-Oncogénicas c-met/genética , Células 3T3 , Animales , Línea Celular , Movimiento Celular/fisiología , Perros , Femenino , Humanos , Riñón , Neoplasias Mamarias Experimentales/genética , Metalotioneína/biosíntesis , Metalotioneína/genética , Ratones , Ratones Endogámicos C3H , Ratones Endogámicos C57BL , Ratones Transgénicos , Mutagénesis Sitio-Dirigida , Metástasis de la Neoplasia , Mutación Puntual , Proteínas Proto-Oncogénicas c-met/biosíntesis , Proteínas Recombinantes de Fusión/biosíntesis , TransfecciónRESUMEN
Using double immunofluorescence staining and quantitative confocal laser scan microscopy, we show that the intensity of hepatocyte growth factor/scatter factor (HGF/SF) and Met staining in human primary brain tumors increases with the grade of malignancy and is prevalent in both the infiltrating tumor cells and endothelial hyperplastic areas. HGF/SF and Met also are expressed in vitro in glioblastoma multiforme cell lines as well as in normal human astrocyte (NHA) cells. Moreover, HGF/SF stimulates tyrosine phosphorylation of Met in both glioma cell lines and NHA cells, but only the glioma cell lines proliferate and become motile and invasive in response to HGF/SF, whereas the NHA cells are nonresponsive. These results implicate autocrine/paracrine Met-HGF/SF signaling in glioma tumorigenesis and suggest that HGF/SF signaling through Met is negatively regulated in NHA cells.
Asunto(s)
Astrocitos/metabolismo , Neoplasias Encefálicas/metabolismo , Glioma/metabolismo , Factor de Crecimiento de Hepatocito/biosíntesis , Proteínas Proto-Oncogénicas c-met/biosíntesis , Células 3T3 , Animales , Astrocitos/citología , Neoplasias Encefálicas/patología , División Celular/efectos de los fármacos , Glioblastoma/metabolismo , Glioblastoma/patología , Glioma/patología , Factor de Crecimiento de Hepatocito/farmacología , Humanos , Ratones , Morfogénesis , Invasividad Neoplásica , Proteínas Recombinantes/biosíntesis , Transfección , Células Tumorales CultivadasRESUMEN
Expression of seven serotonin or 5-hydroxytryptamine (5-HT) receptors (5-HT1D alpha, 5-HT1E, 5-HT2, 5-HT1A, 5-HT1C, 5-HT1D beta, and 5-HT6) was investigated in human normal fetal astrocytes and eight glioma cell lines by reverse transcription and polymerase chain reaction (RT-PCR). No expression of 5-HT1D beta and 5-HT6 was observed in any of the cell lines studied. The 5-HT1D alpha receptor was found to be expressed in two human glioma cell lines but not in normal astrocytes. In addition, only three glioma cell lines expressed the 5-HT1E receptor. The 5-HT1C receptor was expressed in six glioma cell lines but not in normal astrocytes while the 5-HT1A was found to be expressed in normal astrocytes from the left hemisphere and in six glioma cell lines but not in normal astrocytes from the cerebellum. Interestingly, the 5-HT2 receptor was expressed in all cells studied but very weakly in normal astrocytes. The effect of 5-HT on glioma cell proliferation, migration, and invasion was also investigated. Serotonin was found to positively modulate these three processes in vitro. These results suggest that 5-HT may play an important role in the control of the biological properties of human glioma cells.
Asunto(s)
Astrocitos/metabolismo , Neoplasias Encefálicas/metabolismo , Glioma/metabolismo , Proteínas de Neoplasias/biosíntesis , Proteínas del Tejido Nervioso/biosíntesis , Receptores de Serotonina/biosíntesis , Astrocitos/efectos de los fármacos , División Celular/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Proteínas Fetales/biosíntesis , Proteínas Fetales/genética , Regulación Neoplásica de la Expresión Génica , Glioma/clasificación , Glioma/patología , Humanos , Invasividad Neoplásica/fisiopatología , Proteínas de Neoplasias/genética , Proteínas del Tejido Nervioso/genética , Reacción en Cadena de la Polimerasa , ARN Mensajero/análisis , ARN Neoplásico/análisis , Receptores de Serotonina/clasificación , Receptores de Serotonina/efectos de los fármacos , Receptores de Serotonina/genética , Células Tumorales Cultivadas/efectos de los fármacosRESUMEN
A major hallmark of gliomas is their intense neovascularisation. Ganglioside GD3, is one of the major gangliosides which has been implicated in tumour angiogenesis. Recently we reported that GD3 was a potent stimulator of vascular endothelial growth factor release in human glioma cell lines. In the present study we were able to detect GD3-immunoreactivity in 10 out of 10 cases of glioblastoma multiforme and 7 out of 10 cases of anaplastic astrocytoma while low grade tumours were negative. Interestingly, GD3 was intensively expressed in hypervascularised areas of high grade gliomas. These data support the involvement of this ganglioside in brain tumour angiogenesis.
Asunto(s)
Neoplasias Encefálicas/irrigación sanguínea , Gangliósidos/análisis , Glioma/irrigación sanguínea , Neovascularización Patológica/metabolismo , Vasos Sanguíneos/química , Vasos Sanguíneos/metabolismo , Factores de Crecimiento Endotelial/biosíntesis , Factores de Crecimiento Endotelial/metabolismo , Humanos , Inmunohistoquímica , Linfocinas/biosíntesis , Linfocinas/metabolismo , Factor A de Crecimiento Endotelial Vascular , Factores de Crecimiento Endotelial VascularRESUMEN
Matrix metalloproteinases (MMPs) are zinc-dependent peptidases and are amongst those enzymes responsible for extracellular matrix (ECM) degradation during tumour-cell migration. Gangliosides are a family of acidic membrane glycolipids thought to play a role during cell development, differentiation and oncogenic transformation. In this descriptive study, we investigated the effects of six exogenous gangliosides (GM1, GM3, GD1a, GD1b, GD3 and GT1b) on the secretion of MMP-2 (72 kDa gelatinase or gelatinase-A) and MMP-9 (92 kDa gelatinase or gelatinase-B). Cell-conditioned media from eight human glioma-derived cell-lines served as the source of MMPs and were investigated using SDS-PAGE zymography. Six of the cell lines showed upregulation of secretion of both enzymes by all six gangliosides. Of the remaining two cell lines, one showed inhibition of MMP secretion by all gangliosides and the other had a small but differential response to the range of gangliosides investigated. These results suggest that gangliosides may stimulate glioma cell invasiveness by promoting MMP expression.
Asunto(s)
Colagenasas/metabolismo , Gangliósidos/farmacología , Gelatinasas/metabolismo , Glioma/enzimología , Metaloendopeptidasas/metabolismo , Medios de Cultivo Condicionados , Electroforesis en Gel de Poliacrilamida , Humanos , Metaloproteinasa 2 de la Matriz , Metaloproteinasa 9 de la Matriz , Células Tumorales Cultivadas/efectos de los fármacos , Células Tumorales Cultivadas/enzimologíaRESUMEN
Vascular endothelial growth factor (VEGF) is an angiogenic factor which is known to be expressed in several malignancies including glioma. The effect of transforming growth factor-beta (TGF-beta) isoforms as well as gangliosides on VEGF production was investigated in human glioma cell lines. TGF-beta isoforms and gangliosides were found to differentially stimulate VEGF production by these cells. The ganglioside GD3 enhanced this release to the greatest extent and the stimulation was more marked in a glioblastoma cell line than in the two other anaplastic astrocytoma cell lines. These results suggest that both TGF-betas and gangliosides may act as indirect angiogenic factors by stimulating VEGF secretion.
Asunto(s)
Factores de Crecimiento Endotelial/biosíntesis , Gangliósidos/farmacología , Glioma/metabolismo , Linfocinas/biosíntesis , Factor de Crecimiento Transformador beta/farmacología , Humanos , Isomerismo , Estimulación Química , Factor de Crecimiento Transformador beta/farmacocinética , Células Tumorales Cultivadas , Factor A de Crecimiento Endotelial Vascular , Factores de Crecimiento Endotelial VascularRESUMEN
In a recent study we showed that hyaluronic acid (HA) induces cell detachment and promotes migration and invasion of glioma cells in vitro through its interaction with the cell surface molecule CD44H. In this study, the role of HA in proliferation in eight human glioma-derived cell Lines was investigated. We demonstrate that HA exerts a dose dependent proliferative and anti-proliferative effect on glioma cells. This effect was found to be partially counteracted by a CD44H monoclonal antibody, suggesting the involvement of its high affinity receptor, CD44H and other HA receptors in this process.
RESUMEN
The mechanisms underlying the invasive properties of gliomas, the major form of intrinsic brain tumours in humans, are poorly understood. We have reported that CD44 plays an important role in this behaviour in vitro. In the present work, we investigated the role of its ligand, hyaluronic acid (HA), in invasion in 8 human glioma cell lines. We found that HA mediates cell detachment via its interaction with its high affinity receptor, CD44H. Using 8 microns porosity polycarbonate filter transwells, we demonstrate that HA strongly stimulates migration in all 8 cell lines. This effect was found to be partially counteracted by a CD44H monoclonal antibody (MAb), suggesting the involvement of CD44H, as well as other HA receptors, in this process. Furthermore, incorporation of increasing concentrations of HA in Matrigel in an in vitro invasion assay resulted in a substantial increase in the invasive propensity of the glioma cell lines. Moreover, blocking experiments with the CD44H MAb suggest that CD44H and other receptors interact with HA to promote cell invasion in vitro. Our results show that HA induces cell detachment, stimulates migration and promotes invasion via its interaction with CD44H and other HA receptors in vitro. These effects could be prevented by use of specific HA receptor antibodies.
Asunto(s)
Glioma/patología , Receptores de Hialuranos/fisiología , Ácido Hialurónico/farmacología , Animales , Adhesión Celular/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Humanos , Ratones , Invasividad Neoplásica , Células Tumorales CultivadasRESUMEN
Gliomas, the most common form of intrinsic brain tumor, are characterized by diffuse local invasion of the normal brain structures, irrespective of their histological grade of malignancy; a feature that is a major obstacle to successful therapy. They generally infiltrate the central nervous system (CNS) as individual tumor cells several centimeters beyond the macroscopic tumor margin and consequently often recur, after subtotal surgical resection. Factors involved in the control of both their proliferation and invasiveness are poorly documented. In this work, the role of gangliosides on proliferation of both human fetal human brain cells and five cell lines derived from human gliomas with different grades of malignancy was investigated. In addition, 8 microns-porosity polycarbonate filters were used to study cell motility. In addition, these filters were coated with the reconstituted extracellular matrix (ECM) composite, Matrigel, to assess invasiveness. The results presented show that gangliosides generally exert a proliferation inhibitory effect on fetal brain cells and glioma cell lines in vitro and play an important role in promoting glioma cell motility and invasiveness. The molecular mechanisms involved in the action of gangliosides may prove useful in identifying new targets for an anti-invasion therapy.
Asunto(s)
Neoplasias Encefálicas/patología , Gangliósidos/farmacología , Glioma/patología , División Celular/efectos de los fármacos , Quimiotaxis/efectos de los fármacos , Depresión Química , Matriz Extracelular/efectos de los fármacos , Matriz Extracelular/fisiología , Humanos , Células Tumorales CultivadasRESUMEN
Adhesion of eight cell lines, derived from human gliomas of different histological types, to fibronectin, collagen I, vitronectin, and laminin was investigated in vitro. The glioma cell lines were found to attach to these substrates to different extents. Interestingly, all cell lines strongly attached to laminin. In addition, glioma cell adhesion was found to be dose dependent. Moreover, adhesion of three cell lines to fibronectin and collagen I was partially inhibited and to vitronectin completely prevented by GRGDTP peptide, indicating the involvement of integrin receptors in glioma cell adhesion. We have demonstrated, recently, that gangliosides play an important role in promoting glioma cell invasion of the reconstituted basement membrane, Matrigel, in vitro. In order to study the mechanism of action of gangliosides in this process, the role of six gangliosides (GM1, GM3, GD3, GD1a, GD1b, and GT1b) in cell adhesion to the four proteins was investigated in three cell lines. Although all gangliosides, with the exception of GM3, were found to enhance cell adhesion to these proteins to different extents, GD3 proved to be the most effective adhesion-promoting ganglioside in all three cell lines. GM3 was found to inhibit cell adhesion to the four proteins in one cell line but enhanced cell adhesion in two other cell lines. The three cell lines were found to express both GD3 and gangliosides recognised by the A2B5 antibody. Furthermore, adhesion of the three cell lines to fibronectin, vitronectin, laminin, and collagen I was inhibited by incubation with A2B5, demonstrating the involvement of intrinsic cell membrane gangliosides in adhesion of glioma cells to these proteins. Taken together with the observation that gangliosides modulate integrin receptor function, these data suggest that gangliosides may play a central role in the control of the adhesive and invasive properties of human glioma cells.
Asunto(s)
Adhesión Celular/efectos de los fármacos , Proteínas de la Matriz Extracelular/metabolismo , Gangliósidos/farmacología , Glioma/fisiopatología , Fragmentos de Péptidos/metabolismo , Secuencia de Aminoácidos , Membrana Basal/metabolismo , Adhesión Celular/fisiología , Proteínas de la Matriz Extracelular/química , Gangliósidos/fisiología , Humanos , Integrinas/fisiología , Datos de Secuencia Molecular , Peso Molecular , Oligopéptidos/farmacología , Fragmentos de Péptidos/química , Células Tumorales CultivadasRESUMEN
Neoplastic cells from intrinsic, neuroectodermal tumours may migrate up to several millimeters away from the original tumour mass into normal nervous tissue. The biological mechanisms underlying this local invasive behaviour of gliomas are poorly understood. We have demonstrated recently that growth factors and cell surface gangliosides are positively involved in human glioma cell adhesion, migration and invasion in vitro. In order to study the mechanism of action of gangliosides and growth factors in this process, their role in the production of laminin, the major component of glioma vascular basal lamina, was investigated. Both growth factors and gangliosides stimulated laminin production in vitro suggesting that these factors increase laminin production in order to enable glioma cells to adhere and then migrate and invade in vivo.