neuropeptide-y and Brain-Neoplasms

Ewing sarcoma (ES) develops in bones or soft tissues of children and adolescents. The presence of bone metastases is one of the most adverse prognostic factors, yet the mechanisms governing their formation remain unclear. As a transcriptional target of EWS-FLI1, the fusion protein driving ES transformation, neuropeptide Y (NPY) is highly expressed and released from ES tumors. Hypoxia up-regulates NPY and activates its pro-metastatic functions. To test the impact of NPY on ES metastatic pattern, ES cell lines, SK-ES1 and TC71, with high and low peptide release, respectively, were used in an orthotopic xenograft model. ES cells were injected into gastrocnemius muscles of SCID/beige mice, the primary tumors excised, and mice monitored for the presence of metastases. SK-ES1 xenografts resulted in thoracic extra-osseous metastases (67%) and dissemination to bone (50%) and brain (25%), while TC71 tumors metastasized to the lungs (70%). Bone dissemination in SK-ES1 xenografts associated with increased NPY expression in bone metastases and its accumulation in bone invasion areas. The genetic silencing of NPY in SK-ES1 cells reduced bone degradation. Our study supports the role for NPY in ES bone invasion and provides new models for identifying pathways driving ES metastases to specific niches and testing anti-metastatic therapeutics.

Topics: Animals; Bone Neoplasms; Brain Neoplasms; Cell Line, Tumor; Cell Movement; Culture Media, Conditioned; Disease Models, Animal; Female; Gene Silencing; Humans; Hypoxia; Lung Neoplasms; Mice; Mice, SCID; Neoplasm Metastasis; Neoplasm Transplantation; Neuropeptide Y; Oncogene Proteins, Fusion; Phenotype; Proto-Oncogene Protein c-fli-1; RNA-Binding Protein EWS; Sarcoma, Ewing

We detected distinct plasma concentration profiles of S100B, neuropeptide Y, and secretagogin in 3 of 191 patients enrolled in a previous study investigating brain-tissue-related markers in the blood of patients with atrial fibrillation. Intriguingly, 2 of these 3 patients, both of whom were without neurological symptoms at the time of blood sampling, were diagnosed with malignant glioma (MG) approximately 1 year later. To our knowledge, this is the first report indicating that distinct blood biomarker profiles may be detected long before clinical manifestation of MG.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Atrial Fibrillation; Biomarkers, Tumor; Brain Neoplasms; Calcium-Binding Proteins; Female; Glioma; Humans; Male; Middle Aged; Nerve Growth Factors; Neuropeptide Y; S100 Calcium Binding Protein beta Subunit; S100 Proteins; Secretagogins; Young Adult

Mammalian cells often receive multiple extracellular stimuli under physiological conditions, and the various signaling inputs have to be integrated for the processing of complex biological responses. G protein-coupled receptors (GPCRs) are critical players in converting extracellular stimuli into intracellular signals. In this report, we examined the integration of different GPCR signals by mitogen-activated protein kinases (MAPKs) using the SK-N-MC human brain neuroepithelioma cells as a neuronal model. Stimulation of the Gi-coupled neuropeptide Y1 and Gq-coupled muscarinic M1 acetylcholine receptors, but not the Gs-coupled dopamine D1 receptor, led to the activation of extracellular signal-regulated kinase (ERK). All three receptors were also capable of stimulating c-Jun NH2-terminal kinases (JNK) and p38 MAPK. The Gi-mediated ERK activation was completely suppressed upon inhibition of Src tyrosine kinases by PP1, while the Gq-induced response was suppressed by both PP1 and the Ca2+ chelator, BAPTA-AM. In contrast, activations of JNK and p38 by Gs-, Gi-, and Gq-coupled receptors were sensitive to PP1 and BAPTA-AM pretreatments. Simultaneous stimulation of Gi- and Gq-coupled receptors resulted in the synergistic activation of ERK, but not JNK or p38 MAPK. The Gi/Gq-induced synergistic ERK activation was PTX-sensitive, and appeared to be a co-operative effect between Ca2+ and Src family tyrosine kinases. Enhanced ERK activation was associated with an increase in CREB phosphorylation, while the JNK and p38-responsive transcription factor ATF-2 was weakly enhanced upon Gi/Gq-induction. This report provides evidence that G protein signals can be integrated at the level of MAPK, resulting in differential effects on ERK, JNK and p38 MAPK in SK-N-MC cells.

Topics: Adenylyl Cyclases; Brain Neoplasms; Calcium Signaling; Carbachol; Cyclic AMP Response Element-Binding Protein; Drug Combinations; Drug Synergism; Enzyme Activation; Extracellular Signal-Regulated MAP Kinases; GTP-Binding Proteins; Humans; JNK Mitogen-Activated Protein Kinases; MAP Kinase Kinase 1; MAP Kinase Kinase 2; Neuroectodermal Tumors, Primitive, Peripheral; Neurons; Neuropeptide Y; p38 Mitogen-Activated Protein Kinases; Receptor, Muscarinic M1; Receptors, G-Protein-Coupled; Receptors, Neuropeptide Y; Signal Transduction; src-Family Kinases; Transcription Factors; Tumor Cells, Cultured; Type C Phospholipases

Neuropeptide Y (NPY) regulates neurotransmitter release through activation of the Y2 receptor subtype. We have recently characterized a human glioblastoma cell line, LN319, that expresses exclusively NPY Y2 receptors and have demonstrated that NPY triggers transient decreases in cAMP and increases in intracellular calcium responses. The present study was designed to further characterize calcium signalling by NPY and bradykinin (BK) in LN319 cells. Both agonists elevated free intracellular calcium ([Ca(2+)](i)) without soliciting calcium influx. NPY appeared to activate two distinct signalling cascades that liberate calcium from thapsigargin- and ryanodine-insensitive compartments. One pathway proceeded through phospholipase C (PLC)-dependent phosphatidylinositol turnover, while the other triggered calcium release through a so far unidentified mediator. Part of the response was sensitive to pertussis toxin (PTX) under conditions where the toxin totally abolished the NPY-mediated effects on cAMP. The calcium release induced by BK on the other hand was largely PTX-insensitive, PLC-dependent, and from both thapsigargin- and ryanodine-sensitive stores. Following stimulation with NPY, subsequent [Ca(2+)](i) responses to NPY were strongly depressed. Partial heterologous desensitization occurred, when BK was used as the first agonist, whereas NPY had no effect on a subsequent stimulation with BK. These data suggest that NPY-induced calcium mobilization in LN319 cells involves two different G proteins and signalling mediators, and a hitherto unidentified calcium compartment. Homologous desensitization of NPY signalling might be explained by receptor-G protein uncoupling, while heterologous desensitization by BK could be the result of either transient depletion or inhibition of a mediator in the calcium signalling cascades activated by NPY.

Topics: Animals; Bradykinin; Brain Neoplasms; Calcium; Cyclic AMP; Estrenes; Glioblastoma; GTP-Binding Proteins; Humans; Inositol Phosphates; Neuropeptide Y; Pertussis Toxin; Phosphodiesterase Inhibitors; Protein Binding; Pyrrolidinones; Receptors, Neuropeptide Y; Ryanodine; Signal Transduction; Swine; Thapsigargin; Tumor Cells, Cultured; Type C Phospholipases; Virulence Factors, Bordetella

The aim of this study was to assess human intracranial tumours for their gene expression pattern of the vasoactive peptide adrenomedullin (AM), its receptor (AM-R) and leptin, which exerts multiple biological effects including proliferation and angiogenesis via the leptin receptor (OB-Rb). Gene activity of neuropeptide Y (NPY) was monitored additionally. We investigated whether there was a characteristic gene expression pattern of AM and leptin in different intracranial tumours, depending on their proliferation activity and biological behaviour. We investigated 35 non-functioning pituitary adenomas (including eight null cell, four silent plurihormonal, 23 silent gonadotroph adenomas), seven somatotropinomas, seven prolactinomas, eight meningiomas, five astrocytomas, two glioblastoma multiformes and unaffected temporal lobe (n = 8). Quantitative reverse transcriptase-polymerase chain reaction (TaqMan RT-PCR) was performed. AM mRNA was detectable in all tumour specimens. AM/GAPDH (glyceraldehyde-3-phosphate dehydrogenase) ratio was significantly higher in somatotropinomas, as was AM/CD31 ratio in prolactinomas, compared with inactive adenomas (P < 0.05). AM-R mRNA was found in all tumour subgroups in small quantities but, in general, higher in tumours than in temporal lobe tissue, respectively. AM-R/CD31 ratio was significantly higher in prolactinomas than in inactive adenomas (P < 0.05). Leptin was detectable in very low quantities in each subgroup. OB-Rb gene expression was found in all tumour subgroups, OB-Rb/GAPDH ratio was highest for meningiomas (P < 0.0001, compared with temporal lobe). NPY mRNA was detectable in temporal lobe in higher quantities than in tumours (P < 0.0001), and almost undetectable in prolactinomas and astrocytomas. Our data demonstrate that AM and AM-R, NPY, as well as leptin and OB-Rb, are expressed in various intracranial tumours in humans but their particular function has to be elucidated further. At present, there is no evidence for a cross-talk on transcriptional level between the peptidergic vasodilative system AM and the putative angiogenic and proliferation affecting factor leptin.

Topics: Adenoma; Adrenomedullin; Adult; Aged; Brain Neoplasms; Carrier Proteins; Female; Gene Expression; Hormones; Humans; Leptin; Male; Middle Aged; Neuropeptide Y; Neuropeptides; Peptides; Pituitary Neoplasms; Receptors, Adrenomedullin; Receptors, Cell Surface; Receptors, Leptin; Receptors, Peptide

Topics: Bombesin; Brain Neoplasms; Child; Gastrin-Releasing Peptide; Humans; Neoplasms, Nerve Tissue; Neuroectodermal Tumors; Neuropeptide Y; Neuropeptides; Neurotensin; Radioimmunoassay; Somatostatin; Vasoactive Intestinal Peptide

1. In this study we have investigated neuropeptide Y (NPY) and somatostatin (SRIF) receptor-mediated elevation of intracellular Ca2+ concentration ([Ca2+]i) in the human neuroblastoma cell line SH-SY5Y. 2. The Ca(2+)-sensitive dye fura 2 was used to measure [Ca2+]i in confluent monolayers of SH-SY5Y cells. Neither NPY (30-100 nM) nor SRIF (100 nM) elevated [Ca2+]i when applied alone. However, when either NPY (300 pM-1 microM) or SRIF (300 pM-1 microM) was applied in the presence of the cholinoceptor agonist carbachol (1 microM or 100 microM) they evoked an elevation of [Ca2+]i above that caused by carbachol alone. 3. The elevation of [Ca2+]i by NPY was independent of the concentration of carbachol. In the presence of 1 microM or 100 microM carbachol NPY elevated [Ca2+]i with a pEC50 of 7.80 and 7.86 respectively. 4. In the presence of 1 microM carbachol the NPY Y2 selective agonist peptide YY(3-36) (PYY(3-36)) elevated [Ca2+]i with a pEC50 of 7.94, the NPY Y1 selective agonist [Leu31, Pro34]-NPY also elevated [Ca2+]i when applied in the presence of carbachol, but only at concentrations > 300 nM. The rank order of potency, PYY(3-36) > or = NPY > > [Leu31, Pro34]-NPY indicates that an NPY Y2-like receptor is involved in the elevation of [Ca2+]i. 5. In the presence of 1 microM carbachol, SRIF elevated [Ca2+]i with a pEC50 of 8.24. The sst2 receptor-preferring analogue BIM-23027 (c[N-Me-Ala-Tyr-D-Trp-Lys-Abu-Phe]) elevated [Ca2+]i with a pEC50 of 8.63, and the sst5-receptor preferring analogue L-362855 (c[Aha-Phe-Trp-D-Trp-Lys-Thr-Phe]) elevated [Ca2+]i with a pEC50 of approximately 6.1. Application of the sst3 receptor-preferring analogue BIM-23056 (D-Phe-Phe-Tyr-D-Trp-Lys-Val-Phe-D-Nal-NH2, 1 microM) to SH-SY5Y cells in the presence of carbachol neither elevated [Ca2+]i nor affected the elevations of [Ca2+]i caused by a subsequent coapplication of SRIF. The rank order of potency, BIM-23026 > or = SRIF > > L-362855 > > > BIM-23026 suggests that an sst2-like receptor is involved in the elevation of [Ca2+]i. 6. Block of carbachol activation of muscarinic receptors with atropine (1 microM) abolished the elevation of [Ca2+]i by the SRIF and NPY. 7. Muscarinic receptor activation, not a rise in [Ca2+]i, was required to reveal the NPY or SRIF response. The Ca2+ channel activator maitotoxin (2 ng ml-1) also elevated [Ca2+]i but subsequent application of either NPY or SRIF in the presence of maitotoxin caused no further changes in [Ca2+]i. 8. The elevations of [Ca2+

Topics: Brain Neoplasms; Calcium; Calcium Channels; Carbachol; Electrophysiology; Humans; Inosine Triphosphate; Muscarinic Agonists; Neuroblastoma; Neuropeptide Y; Pertussis Toxin; Receptors, Gastrointestinal Hormone; Receptors, Opioid, delta; Receptors, Somatostatin; Somatostatin; Tumor Cells, Cultured; Virulence Factors, Bordetella

The distribution of voltage-sensitive elevations of the level of Ca2+ in untreated SH-SY5Y cells and cells that had been induced to differentiate with staurosporine was investigated by monitoring fura-2 fluorescence in cell suspensions, and by using microfluorometry and quantitative fluorescence imaging on cell bodies and on cellular processes. Cell bodies of both types of cells displayed small Ca2+ elevations, which were composed of transient and sustained components. Elevations were partially sensitive to the L- and N-channel blockers nifedipine (1 microM) and omega-conotoxin GVIA (100 nM) respectively. Up to ten times Ca2+ elevations were observed in varicosities of treated cells than in cell bodies of treated and cells. These elevations were insensitive to compounds known to release Ca2+ from intracellular stores. Elevations of Ca2+ were sustained, and they were insensitive to 5 microM nifedipine, 100 nM omega-agatoxin IVA and 100 nM omega-conotoxin GVIA, and partially sensitive to 2 microM omega-conotoxin GVIA, indicating predominance of non-L-type, non-N-type, non-P-type channel activity. The intracellular localization of neuropeptide Y, a marker of differentiation in these cells, was also investigated by fluorescence immunocytochemistry. Varicosities of treated cells displayed marked fluorescence when viewed in a confocal microscope. These findings show that the varicosities of staurosporine-treated cells exhibit some of the functional properties of nerve terminals. The varicosities resemble boutons en passant nerve endings and they seem to express Ca2+ channels different from those in the cell body.

Topics: Brain Neoplasms; Calcium Channel Blockers; Calcium Channels; Cell Differentiation; Culture Media; Electrophysiology; Enzyme Inhibitors; Fluorescent Dyes; Fura-2; Humans; Immunohistochemistry; Ion Channel Gating; Neuroblastoma; Neuropeptide Y; Protein Kinase Inhibitors; Staurosporine

Neuropeptide Y (NPY) was continuously administered to Wistar rats in the normal and glioma bearing conditions intracarotidly via the external carotid artery and local cerebral blood flow (LCBF) was measured by a [14C] iodoantipyrine technique. In the normal rats, the LCBF at the injecting side of NPY markedly reduced in 2 minutes after beginning of drug administration. In glioma-bearing rats, LCBF in the cerebral cortex and caudate nucleus reduced to the same level as normal rats. However reduction of LCBF in the tumor was not significantly different from that of ipsilateral cortex and caudate nucleus. The results of this study suggest that flow moduration by NPY injection increases delivery of intra-carotid administered anticancer drugs to the brain tumor with concomitant decrease of neurotoxicity to the normal brain.

Topics: Animals; Antineoplastic Agents; Brain Neoplasms; Carotid Artery, External; Cerebrovascular Circulation; Drug Delivery Systems; Glioma; Injections, Intra-Arterial; Male; Neuropeptide Y; Rats; Rats, Wistar

Biopsies of human cerebral cortex were fixed by immersion and immunostained for the detection of neuropeptides in neuronal cell bodies and axons. Four neuropeptides (neuropeptide Y, somatostatin, , substance P and cholecystokinin) were visualized in a series of adjacent sections. All populations of immunoreactive neurons had a morphology characteristic of interneurons, with variations in dendritic arborizations and laminar distribution. The cholecystokinin-immunoreactive neurons were most numerous in the supragranular layers, whereas neurons containing the other three peptides occurred mainly in infragranular layers, or even in neurons populating the subcortical white matter. Quantitatively, each population of neuropeptide-containing neurons accounted for 1.4-2.5% of the total neuronal population. The distribution of these neurons varied slightly between cytoarchitectonic divisions, with substance P- and somatostatin-immunoreactive neurons dominating in the temporal lobe and cholecystokinin-immunoreactive neurons in the frontal lobe. Neuropeptide Y-immunoreactive neurons dominated in the gray matter of the frontal half of the hemisphere and in the subcortical white matter of the caudal half of the hemisphere. Furthermore, co-existence of neuropeptide Y or substance P immunoreactivity within somatostatin-immunoreactive neurons could be demonstrated using double labeling immunofluorescence techniques. The axonal plexuses immunoreactive for neuropeptide Y, somatostatin, or substance P were distributed in all layers, with a strong predominance of horizontally oriented fibers in layer I, a moderate plexus of randomly oriented fibers in the supra- and infragranular layers, and a slightly weaker innervation of layer IV. Immunoreactive axons formed, in addition, complex terminal arbors, mostly in older subjects, suggesting that they resulted from an as yet undefined aging process. The present study underlines several aspects of the organization of the neuropeptide-containing neurons of the human cerebral cortex, which are of particular interest in the light of the involvement of these neurons in several neurodegenerative diseases.

Topics: Axons; Brain Neoplasms; Cerebral Cortex; Cholecystokinin; Frontal Lobe; Humans; Immunohistochemistry; Neurons; Neuropeptide Y; Neuropeptides; Occipital Lobe; Parietal Lobe; Somatostatin; Substance P; Temporal Lobe

Neuropeptide Y (NPY) was measured in central and peripheral cerebrospinal fluid (CSF) in patients suffering from various intracranial disorders. The central NPY-like immunoreactivity (LI) level showed a concentration of 129 +/- 19 pmol.l-1 and was significantly increased (p less than 0.05) compared to peripheral CSF (73 +/- 9 pmol.l-1). From five patients with subarachnoid haemorrhage the CSF NPY-LI levels reached 154 +/- 47 pmol.l-1. In five patients peripheral and central CSF was collected at the same occasion and the CSF NPY-LI concentration was 76 +/- 17 pmol.l-1 in peripheral and 142 +/- 23 pmol.l-1 in central CSF (p less than 0.01), respectively. In a reference group of 9 patients, who were examined by lumbar myelography because of suspected intervertebral herniated discs, the peripheral CSF NPY-LI concentration was 59 +/- 5 pmol.l-1 a value which was also significantly lower compared to NPY-LI levels in central CSF. Thus it is obvious that NPY is present in human CSF with a relatively higher concentration in central than in peripheral CSF at least in patients with disorders of the central nervous system, suggesting a central origin of the NPY.

Topics: Brain Diseases; Brain Neoplasms; Humans; Intervertebral Disc Displacement; Neuropeptide Y

Concentrations of seven neuropeptides have been determined in 69 human neurological tumours. The majority of tumours were intrinsic to the central nervous system, being astrocytomas. In general, within the the better differentiated tumours (Grade I/II astrocytomas) higher concentrations of five neuropeptides (neuropeptide Y, somatostatin, substance P, vasoactive intestinal peptide and cholecystokinin) were measured in comparison to the poorly differentiated tumours. Of the metastatic tumours, five were derived from oat cell carcinoma of the bronchus. Very high concentrations of bombesin were identified in these metastases.

Topics: Astrocytoma; Bombesin; Brain Neoplasms; Humans; Meningeal Neoplasms; Nerve Tissue Proteins; Neuropeptide Y; Substance P; Vasoactive Intestinal Peptide