neuropeptide-y and Pheochromocytoma

Neuropeptide-Y (NPY) is a 36-amino acid peptide, which belongs, along with peptide YY (PYY), to the pancreatic polypeptide (PP) family. The members of this family of peptides act via G protein-coupled receptors (Rs), six subtypes of which (from Y1- to Y6-R) have been identified. NPY and PYY preferentially bind the Y1-R, Y2-R and Y5-R, while PP mainly acts via the Y4-R. Evidence has been provided that the Y3-R is selective for NPY. NPY and Y-Rs are expressed in the adrenal gland (preferentially adrenal medulla) and pheochromocytomas, where they exert various autocrine-paracrine regulatory functions. Findings indicate that NPY is co-released with catecholamines under a variety of stimuli, including splanchnic nerve and cholinergic- and nicotinic-receptor activation. NPY, mainly acting via the Y1-R, Y2-R and Y3-R, either inhibits catecholamine secretion from bovine adrenal chromaffin cells or stimulates catecholamine secretion from adrenomedullary cells of humans and rats. NPY inhibits aldosterone secretion from dispersed zona glomerulosa (ZG) cells, but this effect has probably to be considered non-specific and toxic in nature, since it is obtained only using micromolar concentrations of the peptide. In contrast, NPY appears to modulate the secretory response of dispersed rat ZG cells to their main agonists (ACTH, angiotensin-II and potassium). However, there is indication that the main effect of NPY on the ZG in rats is indirect and involves the local release of catecholamines, which in turn, acting via beta-adrenoceptors, enhance the secretion of aldosterone. The prolonged treatment with NPY is also able to enhance the growth of the rat ZG. In contrast, the effects of NPY on glucocorticoid secretion from zona fasciculata-reticularis cells are negligible and doutbful. The physiological relevance of the effects of NPY on adrenal medulla and ZG remains to be addressed by future experimental studies employing more selective and potent Y-R antagonists. In contrast, indirect evidence is available that endogenous NPY system may play an important role in the modulation of adrenal functions under paraphysiological conditions (e.g. it seems to dampen exceedingly high responses to stresses). Moreover, it has been also suggested that endogenous NPY may be involved in the regulation of blood pressure and in the pathophysiology of pheochromocytomas.

Topics: Adrenal Glands; Animals; Autocrine Communication; Humans; Neuropeptide Y; Paracrine Communication; Pheochromocytoma; Receptors, Neuropeptide Y

Topics: Adrenal Gland Neoplasms; Alzheimer Disease; Anorexia Nervosa; Autonomic Nervous System Diseases; Biomarkers; Humans; Hypertension; Immunoradiometric Assay; Neuroblastoma; Neuropeptide Y; Pheochromocytoma; Radioimmunoassay; Reference Values

Pheochromocytoma and paragangliomas are rare tumors of chromaffin tissue that secrete catecholamines either intermittently or continuously, producing hypertension with a constellation of symptoms and signs that can be frightening to the patient and that continue to provide perplexing problems for clinicians. With surgical treatment, symptoms will be relieved and hypertension normalized or ameliorated for patients who do not have malignant tumors. Appropriate antihypertensive drugs are used to manage hypertension, to control associated cardiovascular symptoms, and to prepare patients for operation. The question debated most often regarding medical therapy of pheochromocytoma is whether antihypertensive treatment regimens other than nonspecific alpha-blockade are just as effective and safe. Understanding the pathophysiologic mechanisms that sustain the hypertension and the pharmacology of antihypertensive agents allows better selection of antihypertensive therapy.

Topics: Adrenal Gland Neoplasms; Adrenergic alpha-Antagonists; Adrenergic beta-Antagonists; Antihypertensive Agents; Blood Pressure; Calcium Channel Blockers; Catecholamines; Clonidine; Hemodynamics; Humans; Hypertension; Neuropeptide Y; Phenoxybenzamine; Pheochromocytoma

Topics: Humans; Neuropeptide Y; Pheochromocytoma

Topics: Adrenal Gland Neoplasms; Animals; Glioma; Humans; Neuropeptide Y; Pheochromocytoma; Tissue Distribution

This study investigates the release of Neuropeptide Y from eight human pheochromocytomas. Profil immunoreactive Neuropeptide Y (Ir-NPY) levels during the management of surgery were compared with these of norepinephrine (NE) while hemodynamics were monitored. Plasma IrNPY and NE levels increased during tumor manipulation and returned to near normal one hour after operation. However, Ir-NPY levels remained high just after tumor resection while NE levels were significantly decreased. At tumor manipulation and just after tumor resection, plasma Ir-NPY levels were correlated with the systemic vascular resistances (SVR) (r = 0.74; P<0.04 and r = 0.86; P<0.006 respectively). No correlation was found either between plasma Ir-NPY and NE levels or between plasma NE levels and SVR. The release of Ir-NPY from tumor tissue, studied by a superfusion method, exhibited a significant correlation with the plasma Ir-NPY concentrations at the time of corresponding tumor resection (r = 0.95; P<0.007). Chromatographic analysis showed that Ir-NPY in plasma and outflow migrate as human NPY (1-36). These results confirmed that in pheochromocytoma, plasma NPY mainly originates from the tumor and argue for an important role of NPY in pheochromocytoma hypertension as indicated by the correlation between the Ir-NPY levels and the SVR.

Topics: Adrenal Gland Neoplasms; Adult; Analysis of Variance; Female; Hemodynamics; Humans; Hypertension; Male; Middle Aged; Neuropeptide Y; Norepinephrine; Pheochromocytoma; Vascular Resistance

Investigated group consisted of 28 patients (21 female and 7 male) with pheochromocytoma in mean age of 39.9 +/- 13 years, while control group comprised 25 healthy volunteers in mean age 36.0 +/- 7.2 years. Following parameters were determined: plasma and blood viscosity, erythrocyte and platelets aggregation, erythrocyte deformability, hematocrit, beta-thromboglobulin plasma concentration, plasma fibrinogen concentration, lipids concentration, noradrenaline, adrenaline, serotonin, neuropeptide Y plasma concentrations. Plasma fibrinogen and beta-thromboglobulin were increased in pheochromocytoma before surgery after pharmacological treatment and also after excision of the tumor. Whereas altered erythrocyte deformability before any treatment, only. Plasma neuropeptide Y concentration was higher in pheochromocytoma before surgery and after pharmacological treatment. There was no correlation between plasma catecholamines, plasma fibrinogen, beta-thromboglobulin, and erythrocyte deformability. Our results suggest that rheological abnormalities observed in patients with pheochromocytoma may contribute to vascular complications in this subjects. Sympathetic nervous system seems to influence indirectly observed rheological abnormalities.

Topics: Adult; beta-Thromboglobulin; Erythrocyte Deformability; Female; Fibrinogen; Humans; Male; Middle Aged; Neuropeptide Y; Pheochromocytoma; Rheology; Sympathetic Nervous System

The observation that phaeochromocytoma possess specific somatostatin binding sites led us to test the hypothesis that octreotide may have antisecretory potential in patients with phaeochromocytoma. We therefore compared the effects of octreotide and placebo on blood pressure and plasma catecholamines and neuropeptide Y.. Ten consecutive patients referred to a tertiary care centre for the diagnosis and treatment of a phaeochromocytoma.. We performed a crossover comparison of either three 100 micrograms subcutaneous injections of octreotide over one day or 3 injections of octreotide vehicle over another. Blood pressure was measured over 24 hours on each test day using an automatic ambulatory recorder. Blood samples were collected before (at 0800 and 0900 h) and after (at 1000, 1100, 1200, 1300 and 1500 h) placebo or octreotide injection. Plasma catecholamines were assayed by high-performance liquid chromatography and neuropeptide Y was determined using a two-site amplified enzyme immunoassay. All patients then underwent surgery and tumoral somatostatin binding site density was determined by quantitative autoradiography.. Compared to placebo, octreotide did not alter mean 24-hour ambulatory blood pressure or plasma neuropeptide Y, or plasma or urinary catecholamine, levels. Although a moderate reduction in plasma noradrenaline was found in the two patients with the highest tumoral somatostatin binding site densities, overall octreotide-induced variations in plasma noradrenaline did not correlate with somatostatin binding site density. Blood glucose increased from 5.4 +/- 0.3 on placebo to 7.8 +/- 0.5 mmol/l on octreotide (P < 0.01).. In the present controlled conditions, short-term administration of octreotide had no antisecretory effect in patients with phaeochromocytoma.

Topics: Adrenal Gland Neoplasms; Adult; Blood Pressure; Blood Pressure Monitoring, Ambulatory; Catecholamines; Cross-Over Studies; Female; Humans; Male; Middle Aged; Neuropeptide Y; Octreotide; Pheochromocytoma; Receptors, Somatostatin; Single-Blind Method

Topics: Adrenal Gland Neoplasms; Adult; Aged; Biomarkers, Tumor; Epinephrine; Female; Humans; Male; Middle Aged; Neuropeptide Y; Norepinephrine; Pheochromocytoma

Neuropeptide Y (NPY1-36) is a vasoconstrictor peptide co-secreted with catecholamines by sympathetic nerves, the adrenal medulla, and neoplasms such as pheochromocytomas and paragangliomas (PPGLs). It is produced by the intracellular cleavage of proNPY and metabolized into multiple fragments with distinct biological activities. NPY immunoassays for PPGL have a diagnostic sensitivity ranging from 33 to 100%, depending on the antibody used. We have validated a multiplex micro-UHPLC-MS/MS assay for the specific and sensitive quantification of proNPY, NPY1-39, NPY1-37, NPY1-36, NPY2-36, NPY3-36, NPY1-35, NPY3-35, and the C-flanking peptide of NPY (CPON) (collectively termed NPYs), and determined the NPYs reference intervals and concentrations in 32 PPGL patients before, during, and after surgery. Depending on the peptide measured, NPYs were above the upper reference limit (URL) in 20% to 67% of patients, whereas plasma free metanephrine and normetanephrine, the gold standard for PPGL, were above the URL in 40% and 87% of patients, respectively. Age, sex, tachycardia, and tumor localization were not correlated with NPYs. Plasma free metanephrines performed better than NPYs in the detection of PPGL, but NPYs may be a substitute for an early diagnosis of PPGL for patients that suffer from severe kidney impairment or receiving treatments that interfere with catecholamine reuptake.

Topics: Adrenal Gland Neoplasms; Healthy Volunteers; Humans; Metanephrine; Neuropeptide Y; Paraganglioma; Pheochromocytoma; Protein Precursors; Tandem Mass Spectrometry

Pheochromocytomas are catecholamine-producing tumors which are generally benign, but which can also present as or develop into malignancy. Molecular pathways of malignant transformation remain poorly understood. Pheochromocytomas express various trophic peptides which may influence tumoral cell behavior. Here, we investigated the expression of trophic amidated peptides, including pituitary adenylate cyclase-activating polypeptide (PACAP), neuropeptide Y (NPY), and adrenomedullin (AM), and their receptors in benign and malignant pheochromocytomas in order to assess their potential role in chromaffin cell tumorigenesis and malignant transformation. PACAP, NPY, and AM are expressed in the majority of pheochromocytomas studied; NPY exhibiting the highest mRNA levels relative to reference genes. Although median gene expression or peptide levels were systematically lower in malignant compared to benign tumors, no statistically significant difference was found. Among all the receptors of these peptides that were analyzed, only the AM receptor RDC1 displayed a differential expression between benign and malignant pheochromocytomas. This receptor exhibited a fourfold higher expression in malignant than in benign tumors. AM and stromal cell-derived factor 1, which has also been described as a ligand for RDC1, increased the number of human pheochromocytoma cells in primary culture and exerted anti-apoptotic activity on rat pheochromocytoma PC12 cells. In addition, RDC1 gene silencing decreased the number of viable PC12 cells. This study shows the expression of several trophic peptides and their receptors in benign and malignant pheochromocytomas, and suggests that AM and its RDC1 receptor could be involved in chromaffin cell tumorigenesis through pro-survival effects. Therefore, AM and RDC1 may represent valuable targets for the treatment of malignant pheochromocytomas.

Topics: Adrenal Gland Neoplasms; Adrenomedullin; Animals; Blotting, Western; Cell Survival; Formazans; Gene Expression Regulation, Neoplastic; Humans; Immunohistochemistry; Neuropeptide Y; PC12 Cells; Pheochromocytoma; Pituitary Adenylate Cyclase-Activating Polypeptide; Rats; Receptors, Adrenomedullin; Receptors, Neuropeptide Y; Receptors, Peptide; Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; RNA, Small Interfering; Tetrazolium Salts

In rat pheochromocytoma (PC12) cells the dopamine D(2) receptor agonists apomorphine (APO) and n-propylnorapomorphine (NPA) produced a concentration dependent inhibition of K(+)-evoked neuropeptide Y release (NPY-ir). The effect of APO was blocked by the dopamine D(2)-receptor antagonist, eticlopride, but not the D(1)/D(3) or the D(4)/D(2) antagonists, SCH23390 or clozapine, respectively. The D(1)/D(5) receptor agonist, SKF38393 or the D(3) agonists PD128907 and 7-OH DPAT had no effect. Selective N and L-type voltage gated Ca(2+) channel blockers, omega-conotoxin GVIa (Ctx-GVIa) and nifedipine, respectively, produced a concentration dependent inhibition of NPY-ir release but were not additive with APO. The Ca(2+)/calmodulin-dependent protein kinase (CaM kinase) II inhibitor KN-62 produced a concentration-dependent inhibition of NPY-ir release but the combination of KN-62 and APO produced no further inhibition. PMA-mediated protein kinase C stimulation significantly increased both basal and K(+)-evoked release of NPY-ir, and in the presence of PMA APO had no inhibitory effect. The PKC antagonist, chelerythrine, inhibited K(+)-evoked NPY-ir release but was not additive with APO. Neither forskolin-mediated adenylate cyclase activation and the active cAMP analog Sp-cAMPS, nor the adenylate cyclase inhibitor SQ 22536, and the competitive inhibitor of cAMP-dependent protein kinases Rp-cAMPS, had any significant effect on K(+)-evoked NPY-ir release. This suggests the inhibitory effect of APO on K(+)-evoked release of NPY-ir from PC12 cells is most likely mediated through activation of dopamine D(2) receptors leading to direct inhibition of N and L-type voltage gated Ca(2+) channels, or indirect inhibition of PKC, both of which would reduce [Ca(2+)](i) and inactivate CaM kinase.

Topics: Adenylyl Cyclases; Adrenal Gland Neoplasms; Animals; Apomorphine; Calcium Channels; Calcium-Calmodulin-Dependent Protein Kinase Type 1; Calcium-Calmodulin-Dependent Protein Kinases; Cattle; Cyclic AMP-Dependent Protein Kinases; Dopamine; Dopamine Antagonists; Drug Interactions; Neuropeptide Y; PC12 Cells; Pertussis Toxin; Pheochromocytoma; Protein Kinase C; Rabbits; Rats

Phaeochromocytomas are rare neuroendocrine tumours that produce catecholamines and numerous secretory proteins and peptides, including neuropeptide Y (NPY), a vasoactive peptide with influences on blood pressure. The production of catecholamines and NPY by phaeochromocytomas is highly variable. This study examined influences of hereditary factors and differences in catecholamine production on tumour expression of NPY, as assessed by quantitative PCR, enzyme immunoassay and immunohistochemistry. Phaeochromocytomas included hereditary adrenaline-producing tumours (adrenergic phenotype) in multiple endocrine neoplasia type 2 (MEN 2), predominantly noradrenaline-producing tumours (noradrenergic phenotype) in von Hippel-Lindau (VHL) syndrome, and other adrenergic and noradrenergic tumours where there was no clear hereditary syndrome. NPY levels in phaeochromocytomas from VHL patients were lower (P<0.0001) than in those from MEN 2 patients for both mRNA (84-fold difference) and the peptide (99-fold difference). These findings were supported by immunohistochemistry. NPY levels were also lower in VHL tumours than in those where there was no hereditary syndrome. Relative absence of expression of NPY in phaeochromocytomas from VHL patients when compared with other groups appears to be largely independent of differences in catecholamine production and is consistent with a unique phenotype in VHL syndrome.

Topics: Adrenal Gland Neoplasms; Adult; Aged; Analysis of Variance; Female; Humans; Immunohistochemistry; Linear Models; Male; Middle Aged; Multiple Endocrine Neoplasia Type 2a; Neuropeptide Y; Phenylethanolamine N-Methyltransferase; Pheochromocytoma; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; von Hippel-Lindau Disease

Neuropeptide Y (NPY) is a sympathetic neurotransmitter recently found to be potently angiogenic and growth promoting for endothelial, vascular smooth muscle and neuronal cells. NPY and its cognate receptors, Y1, Y2 and Y5, are expressed in neural crest-derived tumors; however, their role in regulation of growth is unknown. The effect of NPY on the growth and vascularization of neuroendocrine tumors was tested using three types of cells: neuroblastoma, pheochromocytoma, and Ewing's sarcoma family of tumors (ESFT). The tumors varied in expression of NPY receptors, which was linked to differential functions of the peptide. NPY stimulated proliferation of neuroblastoma cells via Y2/Y5Rs and inhibited ESFT cell growth by Y1/Y5-mediated apoptosis. In both tumor types, NPY receptor antagonists altered basal growth levels, indicating a regulatory role of autocrine NPY. In addition, the peptide released from the tumor cells stimulated endothelial cell proliferation, which suggests its paracrine angiogenic effects. In nude mice xenografts, exogenous NPY stimulated growth of neuroblastoma tumors, whereas it increased apoptosis and reduced growth of ESFT. However, in both tumors, NPY treatment led to an increase in tumor vascularization. Taken together, this is the first report of NPY being a growth-regulatory factor for neuroendocrine tumors, acting both by autocrine activation of tumor cell proliferation or apoptosis and by angiogenesis. NPY and its receptors may become targets for novel approaches in the treatment of these diseases, directed against both tumor cell proliferation and angiogenesis.

Topics: Animals; Apoptosis; Cell Cycle; Cell Proliferation; Culture Media, Conditioned; Endothelial Cells; Humans; Mice; Mice, Nude; Mitogen-Activated Protein Kinases; Neovascularization, Pathologic; Neuroblastoma; Neuropeptide Y; Pheochromocytoma; Rats; Receptors, Neuropeptide Y; Transplantation, Heterologous

Left ventricular hypertrophy (LVH) in patients with arterial hypertension is closely related to the levels of blood pressure (BP), catecholamines, angiotensin II and other mitogenic peptides. Pheochromocytoma (pheo) is a type of hypertension caused by excessive production of catecholamines. The aim of this study was to determinate if left ventricular hypertrophy in patients with pheochromocytoma is related to catecholamines and neuropeptide Y (NPY).. 29 patients with pheochromocytoma (22 F, age 40 +/- 13 years), plasma concentration of neuropeptide Y immunoreactivity, noradrenaline (NA), and adrenaline (A) were determined. Twenty-four hour urine collection for determination of noradrenaline and adrenaline were performed. Every patient had echocardiographic examination and 24 h ambulatory blood pressure monitoring.. Left ventricular hypertrophy was diagnosed in 14 patients. No differences in systolic and diastolic blood pressure in patients with and without left ventricular hypertrophy were found. Plasma noradrenaline and adrenaline levels did not differ between both groups, while plasma neuropeptide Y immunoreactivity was higher in patients with left ventricular hypertrophy than in patients without left ventricular hypertrophy (18.46 +/- 13.26 vs. 9.3 +/- 5.9 fmol/ml (p = 0.02)). Left ventricular mass index (LVMI) correlated with plasma neuropeptide Y-immunoreactivity (r = 0.42 p = 0.023), however, no relationship between left ventricular mass index and plasma or urine noradrenaline and adrenaline levels were found.. Our results indicate that mitogenic effect of neuropeptide Y may play a role in pathogenesis of left ventricular hypertrophy in patients with pheochromocytoma.

Topics: Adrenal Gland Neoplasms; Adult; Blood Pressure; Catecholamines; Electrocardiography; Epinephrine; Female; Humans; Hypertrophy, Left Ventricular; Male; Middle Aged; Neuropeptide Y; Norepinephrine; Pheochromocytoma

Exocytosis in pheochromocytoma cells was induced by electric stimulation. To chase the movement of vesicles by electric stimulation, dense-core secretory vesicles were visualized by expression of the fusion protein between neuropeptide Y and enhanced green fluorescent protein (EGFP) in these differentiated PC12 rat pheochromocytoma cells. When the cells were stimulated with constant voltage potential at -300 mV, the movement of dense-core secretory vesicles could be regulated.

Topics: Animals; Cell Differentiation; Electric Stimulation; Exocytosis; Green Fluorescent Proteins; Luminescent Proteins; Neurites; Neuropeptide Y; PC12 Cells; Pheochromocytoma; Potassium Chloride; Rats; Recombinant Fusion Proteins; Secretory Vesicles; Stimulation, Chemical; Synaptic Vesicles; Transfection

Neuropeptide Y (NPY) and noradrenaline (NA) are frequently co-localized and co-released in the sympathetic nervous system. Since bradykinin (BK) is known to stimulate neurotransmitter release as NA in adrenal glands, we therefore hypothesized that BK might also be involved in the release of NPY. The effect of BK(1-9) on immunoreactive NPY (Ir-NPY) release was investigated in superfused human pheochromocytoma tissue. BK(1-9) (10(-7)-10(-5) M) was shown to induce a rapid Ir-NPY release in a concentration-dependent manner. This effect of BK(1-9) (10(-6) M) was mimicked by the B2 agonist [Phe(8)(CH(2)NH)Arg(9)]-bradykinin (10(-5) M) and blocked by the selective B2-receptor antagonist HOE140 (10(-5) M). Increasing Ir-NPY release was probably not mediated by nitric oxide (NO) since the outflow of Ir-NPY was not influenced by the NO synthase inhibitor N-omega-nitro-L-arginine methyl ester (L-NAME) (10(-4) M). In presence of bapta-AM (10(-5) M), a chelator of cytosolic calcium, W7 (10(-5) M), a calmodulin inhibitor, TMB-8 (10(-5) M), a blocker of intracellular calcium mobilization and ryanodine (10(-5) M), a selective inhibitor of the Ca(2+)-induced release mechanism, the NPY release by BK(1-9) was significantly inhibited by 126%, 98%, 91%, and 94%, respectively. These results indicate that BK increased the release of NPY by the tumor acting through the interaction with the BK-B2 receptor and request intracellular calcium mobilization independently of a NO mechanism.

Topics: Adrenal Gland Neoplasms; Bradykinin; Calcium; Dose-Response Relationship, Drug; Enzyme Inhibitors; Humans; In Vitro Techniques; Neuropeptide Y; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Pheochromocytoma; Receptor, Bradykinin B2; Receptors, Bradykinin; Stimulation, Chemical

Plasma free metanephrines are a more reliable analyte to measure than catecholamines for the biochemical diagnosis of pheochromocytomas. We hypothesized that the long persistence of total (sulfate-conjugated plus free) metanephrines in the blood might have a significant diagnostic value.. We measured plasma concentrations of catecholamines and total metanephrines (sulfate-conjugated plus free forms) by HPLC with amperometric detection, and neuropeptide Y (NPY) by an amplified ELISA in seven patients before and after removal of their pheochromocytomas. The results for catecholamine, total metanephrines, and NPY in each patient were analyzed for up to 120 min, starting from the time of tumor vessel clamping. The persistence of analytes was quantified as the area under the concentration-time curve over 120 min.. On the basis of the upper reference limit for each variable, plasma free norepinephrine (NE) and epinephrine (E) concentrations were increased preoperatively in at least one sample in seven and six patients, respectively. Total normetanephrine (NMN) and metanephrine (MN) were increased in all samples in seven and six patients, respectively. NPY was increased 2- to 465-fold. After removal of the tumor, MN and NMN showed a higher average relative increase above the upper limit of the reference interval than NE and E (P = 0.05), whereas NPY was intermediate. The persistence of increased values was significantly shorter for catecholamines than for metanephrines. The half-life estimated by nonlinear regression was 12.3 +/- 7.8 min for NPY. Significant correlations were observed among NE, E, NMN, MN, and NPY concentrations, but parent markers (E and MN or NE and NMN) did not appear significantly intercorrelated.. A larger increase and a longer persistence of total metanephrines (reflecting predominantly sulfo-conjugated metanephrines) than catecholamines and NPY in plasma may contribute to their greater diagnostic accuracy in pheochromocytoma.

Topics: Catecholamines; Female; Half-Life; Humans; Male; Metanephrine; Neuropeptide Y; Pheochromocytoma

The expression of the neuropeptide Y (NPY) gene varies considerably in human pheochromocytomas, but the mechanisms for this variation have not been clarified. To investigate the regulation pattern of the NPY gene in human pheochromocytomas, we screened 16 pheochromocytomas and 9 normal adrenal tissues with Northern blots. The expression level of NPY mRNA in normal adrenal medulla was low and relatively constant, while the pheochromocytomas showed a very wide variation in NPY mRNA levels in both malignant and benign tumors. This indicates that NPY gene expression is not correlated with malignancy in pheochromocytomas. In primary cultures of human pheochromocytoma cells, nerve growth factor treatment (causing neuronal differentiation) increased NPY mRNA accumulation 2- to 5-fold (P < 0.05). NPY mRNA levels were also induced by protein kinase modulators (Bu)(2)cAMP and staurosporine in the cultures (P < 0.05). In contrast, treatment with dexamethasone and IGF-II (causing or linked with chromaffin differentiation) reduced NPY mRNA accumulation (P < 0.05). These data show that the regulation pattern of NPY mRNA expression in cultured human pheochromocytoma cells is different from that previously described in rat pheochromocytoma PC12 cells. Regulation of NPY mRNA expression in primary cultures by these differentiating factors suggests that the expression of NPY mRNA in pheochromocytoma tissues may be associated with the neuronal differentiation of the tumor cells affected by multiple factors.

Topics: Adrenal Gland Neoplasms; Adrenal Medulla; Gene Expression Regulation, Neoplastic; Humans; Neuropeptide Y; Pheochromocytoma; RNA, Messenger; Tumor Cells, Cultured

Employing clonal cell lines derived from rat embryonic hippocampal cells, we detected neuropeptide Y (NPY) mRNA in three progenitor subcloned cell lines. These cell lines upon differentiation express markers indicative of commitment to either neuronal (H19-7; NF +, GFAP -), glial (H19-5; GFAP +, NF -), or bipotential (H583-5; NF +, GFAP + ) lineages. Induction of differentiation was associated with the persistence of the NPY mRNA, however, in the differentiated H19-7 cells a 20-fold increase in NPY mRNA levels was observed (P<0.05). NPY immunoreactivity was observed only in cells with a differentiated neuronal phenotype. The cellular radioimmunoassayable NPY peptide levels increased twelve-fold without a change in extracellular NPY peptide levels by multi-factorially induced neuronal or glial cell differentiation. The differentiated H19-5 cells expressed lower levels of NPY that could not be immunocytochemically detected. The peripheral sympathetic PC-12 neuronal cells examined in the undifferentiated and nerve growth factor-driven differentiated states expressed NPY only upon differentiation. We conclude that NPY is expressed by the cultured undifferentiated and differentiated rat hippocampal clonal cell lines, while the peripheral sympathetic PC-12 neuronal cell line only expresses the NPY gene upon differentiation. These immortalized embryonic neural cell line(s) will provide a hippocampal cell line(s) to conduct future in-vitro investigations targeted at determining the cellular and molecular mechanisms governing NPY gene expression.

Topics: Animals; Cell Differentiation; Cell Line, Transformed; Hippocampus; Immunohistochemistry; Neuropeptide Y; PC12 Cells; Pheochromocytoma; Rats; RNA, Messenger

Neuropeptide Y, an abundant neurohormone present with catecholamines in the adrenal medulla, is a potent non-adrenergic vasoconstrictor and a vascular growth factor.. To determine the mechanism of the release from, and possible role of neuropeptide Y in, pheochromocytomas, compared with those of catecholamines.. Plasma and tumour levels of neuropeptide Y-immunoreactivity (by, radioimmunoassay) and of noradrenaline and adrenaline (by a radioenzymatic method) in 29 patients (19 women and 10 men, aged 22-68 years) were measured during surgical removal of the tumour, during alpha-adrenergic and beta-adrenergic blockade. Arterial systemic blood samples were withdrawn before the ligation of the vessels supplying the tumour, during its surgical manipulations and after its removal, while haemodynamics was monitored.. Plasma neuropeptide Y levels in 17 patients (58.6%, group I) significantly increased during manipulations of the pheochromocytoma and returned completely to normal after its removal. This response was independent of the plasma neuropeptide Y immunoreactivity manipulation and was correlated to increases in plasma noradrenaline (r = 0.638, P < 0.02) but not adrenaline levels. Manipulation-induced increases in plasma neuropeptide Y-immunoreactivity were associated with greater neuropeptide Y content in tumours (r = 0.508, P < 0.05) but neither plasma nor tumour levels of neuropeptide Y immunoreactivity were correlated to tumour mass. Plasma levels of neuropeptide Y immunoreactivity in the remaining 12 patients (41.4%, group II) remained unchanged throughout the experimental period, while levels of circulating catecholamine rose. In all, in spite of our attempt at complete adrenergic blockade, tumour manipulation elevated arterial blood pressure and these changes were significantly correlated to increases in levels of catecholamines in patients in both groups but also to plasma neuropeptide Y immunoreactivity in patients in group I.. Pheochromocytomas exhibit different patterns of secretion. For about half of the patients either the secretion of neuropeptide Y is uncoupled from that of catecholamines or its secretion could be obscured by an increase in degradation of neuropeptide Y to inactive fragments undetectable by radioimmunoassay.

Topics: Adult; Aged; Catecholamines; Female; Humans; Male; Middle Aged; Neuropeptide Y; Pheochromocytoma; Radioimmunoassay

Specific assays for measurements of circulating chromogranin (Cg) A, CgB, CgC and pancreastatin (Ps) have recently been developed. The aim of the present study was to investigate the usefulness of these markers in diagnosing and following the effects of treatment of patients with phaeochromocytoma, and to compare the results with those concerning other biochemical markers. CgA was elevated in 19/21 (90%), CgB in 17/21 (81%), Ps in 9/21 (43%) and neuropeptide Y in 9/21 (43%) of the patients. Urinary noradrenaline was increased in 19/21 (90%) and urinary adrenaline in 17/19 (89%) of the patients. All patients had increased levels of either urinary catecholamines or plasma chromogranins. In one patient levels of CgA, CgB and Ps were measured at frequent intervals before, during and after surgery. The CgA level fell to normal shortly after the tumour was removed, whereas the CgB level decreased towards normal over the course of several days. Significant correlation was observed between the contents of CgA and CgB in the tumour tissue and the plasma levels of CgA and CgB respectively. We conclude that CgA and CgB are sensitive circulating markers for phaeochromocytoma and that measurements of both urinary catecholamines and plasma chromogranins improve the diagnostic sensitivity. Furthermore, measurements of CgA may be useful in assessing the radicality of surgery in the early postoperative period.

Topics: Adolescent; Adrenal Gland Neoplasms; Adult; Aged; Biomarkers, Tumor; Catecholamines; Chromogranin A; Chromogranin B; Chromogranins; Creatinine; Female; Humans; Immunohistochemistry; Male; Middle Aged; Neuropeptide Y; Pancreatic Hormones; Paraganglioma; Pheochromocytoma; Prognosis; Proteins

Neuropeptide Y and nitric oxide (NO) synthase are colocalized in nervous tissues. We tested the hypothesis whether or not NO might be involved in the release of neuropeptide Y. Neuropeptide Y concentration in the supernatant of PC12 rat pheochromocytoma cells, shown to express NO synthase I by immunohistochemistry, rose threefold in a time- and dose-dependent manner following sodiumnitroprusside and 3-morpholinosydnonimine (SIN-1) incubation. Neuropeptide Y mRNA expression was induced by NO-donors as a function of incubation-time. Neuropeptide Y production rose fivefold with zaprinast, an inhibitor of the phosphodiesterase V and threefold with nerve growth factor (NGF). Combined application of zaprinast and NGF did not further increase neuropeptide Y production while combination of zaprinast and sodiumnitroprusside potentiated the NO effect on neuropeptide Y release. The data suggest that NO regulates neuropeptide Y secretion of PC12 pheochromocytoma cells on the mRNA level.

Topics: Animals; Blotting, Northern; Enzyme Inhibitors; Immunohistochemistry; Molsidomine; Nerve Growth Factors; Neuropeptide Y; Nitric Oxide; Nitric Oxide Synthase; Nitroprusside; PC12 Cells; Pheochromocytoma; Phosphodiesterase Inhibitors; Purinones; Radioimmunoassay; Rats; RNA; Sodium-Potassium-Exchanging ATPase

Topics: Adenylyl Cyclases; Adrenal Gland Neoplasms; Animals; Chloramphenicol O-Acetyltransferase; Colforsin; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Enzyme Activation; Gene Expression; Isoquinolines; Kinetics; Naphthalenes; Neuropeptide Y; PC12 Cells; Pheochromocytoma; Polycyclic Compounds; Promoter Regions, Genetic; Protein Kinase C; Rats; Sulfonamides; Transcription, Genetic

Neuropeptide Y (NPY) levels are increased in plasma and tumors of patients with pheochromocytoma. The present study was designed to evaluate plasma and tissue NPY levels simultaneously as well as to study its release and expression in patients with either adrenal or extraadrenal pheochromocytomas. Plasma NPY levels were higher (P < 0.01) in patients with adrenal tumors than in matched normal subjects and patients with extraadrenal tumors. NPY levels were also higher (P < 0.05) in adrenal than in extraadrenal tumors. Bioactive NPY (1-36) was the predominant form in plasma and tumors of patients with adrenal pheochromocytomas. In contrast, patients with extraadrenal pheochromocytomas had an abundance of NPY fragments. NPY mRNA was abundant in 11 of 13 adrenal tumors but in only 1 of 6 extraadrenal tumors. Moreover, NPY was coreleased with NE with manipulation of adrenal but not extraadrenal tumors. These findings indicate that increased NPY gene expression in adrenal pheochromocytomas accounts for the greater biosynthesis and storage of NPY in these tumors and that increased release of NPY results in elevated plasma NPY. Factors regulating NPY gene expression in pheochromocytoma and the role of NPY in the clinical manifestations of the disease remain to be elucidated.

Topics: Adolescent; Adrenal Gland Neoplasms; Adult; Aged; Catecholamines; Female; Humans; Male; Middle Aged; Neuropeptide Y; Pheochromocytoma

Pheochromocytoma is a unique type of hypertension caused by excessive production of catecholamines by the chromaffin tumor. Pheochromocytoma, a potentially life-threatening disease, is a rare cause of hypertension. The incidence varies from 0.1 to 0.8% of hypertensive population. The author's experience is based on 138 patients treated in one institution from 1956 to 1995. Hormonal activity of pheochromocytoma varies considerably, influencing the pattern, of blood pressure and the clinical symptoms. It is emphasized that different other humoral mechanisms may play a role in the pathophysiology of this type of endocrine hypertension. Biochemical tests and non-invasive localizing methods are essential for the definite diagnosis of pheochromocytoma. A great progress has been made in this respect during the last three decades. Surgical removal of the tumor is the only definite therapy with low morbidity and mortality.

Topics: Atrial Natriuretic Factor; Catecholamines; Humans; Hypertension; Neuropeptide Y; Pheochromocytoma; Renin

In PC12 rat pheochromocytoma cells differentiated with nerve growth factor (NGF), neuropeptide Y inhibited depolarization-stimulated catecholamine synthesis as determined by in situ measurement of 3,4-dihydroxyphenylalanine (DOPA) production in the presence of the decarboxylase inhibitor m-hydroxybenzylhydrazine (NSD-1015). The inhibition by neuropeptide Y was concentration-dependent and was prevented by pretreatment with pertussis toxin, suggesting the involvement of a GTP-binding protein of the Gi or Go subtype. The neuropeptide Y analog [Leu31,Pro34]neuropeptide Y also caused inhibition of DOPA production, but was less potent than neuropeptide Y itself, while peptide YY and neuropeptide Y-(13-36) had no significant effect. This pattern is most consistent with the involvement of the neuropeptide Y Y3 receptor subtype. In PC12 cells differentiated with dexamethasone, neuropeptide Y also caused a concentration-dependent inhibition of DOPA production, while peptide YY was again without effect. Neuropeptide Y had no effect on DOPA production in undifferentiated PC12 cells. These results indicate that neuropeptide Y can modulate catecholamine synthesis in addition to its modulatory effects on catecholamine release.

Topics: Adrenal Gland Neoplasms; Analysis of Variance; Animals; Anti-Inflammatory Agents, Non-Steroidal; Aromatic Amino Acid Decarboxylase Inhibitors; Cell Differentiation; Dexamethasone; Dihydroxyphenylalanine; Dose-Response Relationship, Drug; Gastrointestinal Hormones; GTP-Binding Proteins; Hydrazines; Nerve Growth Factors; Neuropeptide Y; PC12 Cells; Peptide Fragments; Peptide YY; Peptides; Pertussis Toxin; Pheochromocytoma; Rats; Structure-Activity Relationship; Virulence Factors, Bordetella

We produced and characterized three anti-C-flanking peptides of neuropeptide Y (CPON) monoclonal antibodies. The Ka for these antibodies ranged from 0.4 to 0.8 x 10(8) l/mol with an IC50 for CPON(1-30) at about 20 nM as determined by ELISA. All these antibodies are IgG1 and recognize the 16-30 part of CPON. These antibodies and a specific anti-NPY monoclonal antibody were used to study the localization of CPON and NPY in the human kidney. The avidin-biotin technique was employed. NPY and CPON immunoreactivities were present in large amount in the renal tubules of the human kidney but not in the glomeruli. No labeling was found within the renal arterioles and veins, but some immunoreactivity was evidenced in the perivascular area. Because no specific receptor for CPON has been described to date, the presence of this peptide in the tubules may be due to a tubular reabsorption or perhaps to a local synthesis of pro-NPY.

Topics: Adrenal Gland Neoplasms; Antibodies, Monoclonal; Antibody Specificity; Biopsy; Cross Reactions; Enzyme-Linked Immunosorbent Assay; Graft Rejection; Humans; Immunoglobulin G; Immunohistochemistry; Kidney; Kidney Diseases; Kidney Glomerulus; Kidney Transplantation; Kidney Tubules; Neuropeptide Y; Peptide Fragments; Pheochromocytoma; Sensitivity and Specificity

A case of a renin-producing paraganglioma of adrenal origin with metastases to the retroperitoneal area, paravaginal area, and the ovary is reported with immunohistochemical findings indicating expression of multiple neuropeptide immunoreactivities. The patient was 23 years old at the time of diagnosis, and died from metastatic spread of the tumor 7 years later.. Tumor tissue was examined by light microscopy, indirect immunohistochemistry, and electron microscopy.. The tumor tissue investigated contained several cells exhibiting opioid peptide-like immunoreactivities (i.e., enkephalin and dynorphin-like immunoreactivity [LI]). A lower number of cells displayed neuropeptide Y-, galanin-, somatostatin-, neurotensin-, substance P-, peptide histidine-isoleucine-, cholecystokinin-, renin-, and calbindin-LI.. To the authors' knowledge, galanin, dynorphin, peptide histidine-isoleucine, cholecystokinin, and calbindin have not been reported previously to occur in paraganglioma, and renin has been reported to occur very rarely. A review of recent literature suggest that enkephalin-, neuropeptide Y-, and somatostatin-like immunoreactivities may be useful as diagnostic markers for paragangliomas.

Topics: Adrenal Gland Neoplasms; Adult; Biomarkers, Tumor; Catecholamines; Enkephalins; Female; Humans; Immunohistochemistry; Neoplasm Metastasis; Neuropeptide Y; Neuropeptides; Pheochromocytoma; Renin; Somatostatin

1. Neuropeptide Y (NPY) gene expression in human phaeochromocytomas was investigated by measuring the levels of NPY mRNA and NPY-immunoreactivity (NPY-IR) in human phaeochromocytoma tissues in comparison with those in normal human adrenal tissues. 2. The amounts of NPY mRNA and NPY-IR in human phaeochromocytomas were 18 and 93 times higher, respectively, than those in normal adrenal glands. In contrast, beta-actin gene expression was similar in human phaeochromocytomas to that in normal adrenal glands. 3. The amount of NPY mRNA relative to total cellular RNA was 6-fold higher in phaeochromocytoma tissues than in normal human adrenal medulla, suggesting increased NPY gene expression in the tumour cells. 4. Induction of differentiation of PC12 rat phaeochromocytoma cells by compounds, such as dexamethasone and nerve growth factor, resulted in a marked increase in the NPY mRNA level. 5. These findings suggest that NPY gene expression is increased in well-differentiated human phaeochromocytoma cells. Its high level of expression could be responsible for the marked overproduction of NPY by this tumour.

Topics: Actins; Adrenal Gland Neoplasms; Adrenal Glands; Amino Acid Sequence; Animals; Blotting, Northern; Cell Differentiation; Dexamethasone; Gene Expression Regulation, Neoplastic; Humans; Molecular Sequence Data; Nerve Growth Factors; Neuropeptide Y; Pheochromocytoma; Radioimmunoassay; Rats; RNA, Messenger; Tumor Cells, Cultured

The potential role of angiotensin-II in mediating catecholamine and neuropeptide-Y release in a human pheochromocytoma has been investigated. Angiotensin-II type I receptors are transcribed and translated into functional proteins in a surgically removed pheochromocytoma. Primary cell culture of the tumor has been studied in a perfused system. Angiotensin-II increased the release of norepinephrine and neuropeptide-Y by the pheochromocytes. Activation of the angiotensin-II type I receptors by angiotensin-II was associated with a rise in cytosolic free calcium. The renin-angiotensin system may, therefore, contribute to the secretion of catecholamines and NPY occurring in patients with pheochromocytoma and when stimulated trigger hypertensive crisis.

Topics: Adrenal Gland Neoplasms; Adult; Angiotensin II; Calcium; Gene Expression; Humans; Male; Neuropeptide Y; Norepinephrine; Perfusion; Pheochromocytoma; Receptors, Angiotensin; Renin-Angiotensin System; Tumor Cells, Cultured

Topics: Adrenal Gland Neoplasms; Catecholamines; Humans; Neuropeptide Y; Pheochromocytoma

Undifferentiated rat pheochromocytoma PC12 cells resemble immature adrenal chromaffin cells, express neuropeptide-Y (NPY) receptors of the Y1 subtype, and synthesize catecholamines as well as NPY. In the present study, we examined how phenotypic alteration of PC12 cells by nerve growth factor (NGF) or glucocorticoid affected cellular responsiveness to NPY and related agonists, especially with regard to modulation of catecholamine overflow. Unlike undifferentiated PC12 cells, cells differentiated to a sympathetic neuronal phenotype with NGF were responsive to the Y2 receptor-selective agonist, NPY 13-36. NPY 13-36 1) inhibited binding of [125I]NPY 1-36, 2) inhibited accumulation of evoked cAMP, and 3) inhibited evoked catecholamine overflow. NGF-differentiated cells were also responsive to the Y1 receptor-selective agonist [Leu31,Pro34]NPY (LP-NPY). Like NPY-(13-36), LP-NPY inhibited binding of [125I]NPY-(1-36); however, LP-NPY and NPY-(13-36) exerted their effects through heterogeneous receptors, as LP-NPY enhanced while NPY 13-36 inhibited evoked catecholamine overflow in NGF-differentiated cells, despite the fact that both agonists inhibited the evoked cAMP. In contrast to NGF-differentiated cells, cells differentiated to a mature chromaffin phenotype with dexamethasone were unresponsive to NPY-(13-36), nor did the Y2 agonist inhibit binding of [125I]NPY-(1-36). Dexamethasone-differentiated PC12 cells were, however, responsive to LP-NPY, as this agonist enhanced evoked catecholamine overflow and inhibited binding of [125I]NPY-(1-36). Peptide-YY also enhanced catecholamine overflow, but only significantly at 100 nM. The data suggest differential expression of NPY receptor subtypes on neuronal and endocrine cells where catecholamine overflow is a key feature. These studies further demonstrate inhibitory or excitatory modulation of catecholamine transmission by NPY via distinct receptor subtypes in homogeneous sympathoadrenomedullary models resembling sympathetic neurons and chromaffin cells.

Topics: Adrenal Gland Neoplasms; Animals; Binding, Competitive; Cell Differentiation; Cell Division; Colforsin; Cyclic AMP; Dose-Response Relationship, Drug; Kinetics; Nerve Growth Factors; Neurons; Neuropeptide Y; Nicotine; PC12 Cells; Peptide Fragments; Pheochromocytoma; Potassium Chloride; Rats; Receptors, Neuropeptide Y

Pheochromocytoma is often associated with paroxysmal hypertension. We report a 49-year-old woman with pheochromocytoma whose blood pressure (BP) was elevated regularly only at night. Plasma norepinephrine (NE) and neuropeptide Y (NPY) concentrations increased in parallel with the elevation of BP. After resection of the adrenal tumor, these circadian changes disappeared. Plasma NE and NPY, especially the former, from the tumor were considered to be the cause of this unusual fluctuation in BP.

Topics: Adrenal Gland Neoplasms; Blood Pressure; Circadian Rhythm; Female; Humans; Hypertension; Middle Aged; Neuropeptide Y; Norepinephrine; Pheochromocytoma

The nature of NPY-like immunoreactivity (NPY-LI) was investigated in plasma and tumour tissue of 17 pheochromocytoma patients by HPLC, gel filtration and isoelectric focusing using two radioimmunoassays (RIAs) directed against the C- and N-terminals of NPY respectively. The two RIAs gave similar results in pheochromocytomas: 86% of cases had higher NPY-LI concentrations than those found in normal adrenal glands and NPY-LI behaved like authentic human NPY during gel filtration and HPLC. Assessed by isoelectric focusing, NPY was found to be amidated in seven of nine tumours. Contrary to the findings obtained in tumours, the results of the two RIAs in plasma samples were not always concordant: compared to controls, elevated concentrations of NPY-LI were found in 86% of cases of pheochromocytomas using the C-terminally directed RIA and in 76% of cases using the N-terminally directed RIA. The results of HPLC and gel filtration of NPY-LI in plasma suggested that circulating C- and N-terminal NPY fragments account for the discrepancy between the results of the two RIAs. In conclusion, most pheochromocytomas contain large amounts of NPY-LI that behaved like authentic NPY by chromatographic analysis. On the contrary, circulating NPY-LI in some pheochromocytoma patients is heterogeneous with cleaved products which influence differently the power of the C- and N-terminally directed RIAs for the diagnosis of pheochromocytoma.

Topics: Adrenal Gland Neoplasms; Adrenal Glands; Adult; Chromatography, Gel; Chromatography, High Pressure Liquid; Female; Humans; Isoelectric Focusing; Male; Middle Aged; Neuropeptide Y; Oxidation-Reduction; Pheochromocytoma; Radioimmunoassay

1. We investigated the usefulness of neuropeptide Y as a plasma marker for phaeochromocytoma, ganglioneuroblastoma and neuroblastoma using a simple and highly sensitive r.i.a. for human neuropeptide Y. 2. Plasma immunoreactive neuropeptide Y concentrations were measured without extraction in plasma samples (100 microliters) from patients with various diseases. 3. The plasma immunoreactive neuropeptide Y concentration in patients with phaeochromocytoma (172.3 +/- 132.4 pmol/l, mean +/- SD, n = 23) was significantly higher than that in healthy adult subjects (40.1 +/- 10.1 pmol/l, n = 40, P < 0.0001). The plasma immunoreactive neuropeptide Y concentrations in patients with ganglioneuroblastoma (590.7 +/- 563.6 pmol/l, n = 6) and patients with neuroblastoma (566.9 +/- 524.4 pmol/l, n = 15) were significantly higher than those in control children (1-9 years old, 82.2 +/- 39.9 pmol/l, n = 72, P < 0.0001). 4. The plasma immunoreactive neuropeptide Y concentration in patients with essential hypertension (34.0 +/- 3.7 pmol/l, n = 18) was within the normal range, but in patients with chronic renal failure undergoing maintenance haemodialysis (192.1 +/- 68.0 pmol/l, n = 25) and in non-dialysed patients with chronic renal failure (85.1 +/- 23.1 pmol/l, n = 7) it was significantly higher than that in healthy adult subjects (P < 0.0001). 5. Eighty-seven per cent of the patients with phaeochromocytoma, 67% of the patients with ganglioneuroblastoma and 80% of the patients with neuroblastoma showed plasma immunoreactive neuropeptide Y concentrations higher than the upper limits in the control subjects [62 pmol/l (adult) and 160 pmol/l (children)].(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adolescent; Adrenal Gland Neoplasms; Adult; Aged; Biomarkers, Tumor; Child; Child, Preschool; Female; Ganglioneuroma; Humans; Hypertension; Infant; Kidney Failure, Chronic; Male; Middle Aged; Neuroblastoma; Neuropeptide Y; Pheochromocytoma; Radioimmunoassay

The mechanisms of hypertension during primary hyperaldosteronism and Cushing's syndrome are not completely understood. An enhanced vascular sensitivity to noradrenaline has been described in both situations. Neuropeptide Y (NPY) induces direct vasoconstriction and potentiates the action of noradrenaline. Sodium retention and dexamethasone have been shown to increase circulating NPY levels in animals and the expression of NPY in neuroendocrine cells. In order to determine if NPY could be involved in the enhanced vascular sensitivity to noradrenaline associated with adrenocortical hyperactivity, we measured plasma NPY in patients with Cushing's syndrome (n = 26) and primary hyperaldosteronism (n = 15) and compared it with that of hypertensive patients with pheochromocytomas (n = 13) or essential hypertension (n = 51) and with normotensive controls (n = 47). The concentration of NPY-Like immunoreactivity (NPY-Li) (mean +/- S.E.) in controls was 39.6 +/- 3.0 pg/ml. Elevated concentrations were found in 77% of the samples collected from pheochromocytoma patients (1180.4 +/- 394.0 pg/ml). NPY-Li levels in patients with essential hypertension (35.0 +/- 2.6 pg/ml), primary hyperaldosteronism (31.3 +/- 3.9 pg/ml) and Cushing's syndrome (33.1 +/- 4.8 pg/ml) were not different from that of controls. NPY-Li levels in hypertensive and normotensive patients with Cushing's syndrome were similar (38.5 +/- 7.5 vs 24.2 +/- 3.7 pg/ml). No correlation was found between the NPY-Li level and the mean blood pressure at the time of sampling. Our results suggest that NPY is unlikely to be involved in the pathogenesis of hypertension associated with primary hyperaldosteronism and Cushing's syndrome.

Topics: Adolescent; Adrenal Gland Neoplasms; Adult; Aged; Blood Pressure; Cushing Syndrome; Female; Humans; Hyperaldosteronism; Hypertension; Male; Middle Aged; Neuropeptide Y; Pheochromocytoma

Twenty patients, 16 males and 4 females, aged 11-76 yr, were treated for a metastatic pheochromocytoma at our institution between 1985 and 1990. A neurofibromatosis was associated in 4. Thirteen patients had a unilateral adrenal tumor, 3 had an extraadrenal retroperitoneal tumor, 2 had a bilateral adrenal pheochromocytoma, one had a unilateral tumor with a contralateral medullary hyperplasia and one an adrenal and an extraadrenal pheochromocytoma. Metastases occurred in all patients, at presentation in 11, 10 to 30 months later in 7, and 9 and 28 yr later, respectively in two. Histology did not afford conclusive evidence for malignancy. Catecholamine hyperproduction was present in all, predominantly affecting norepinephrine. Neuron Specific Enolase level was elevated in 11, Neuro-Peptide Y level in 9 and procalcitonin level in 11/18. High dopamine, methoxytyramine and homovanillic acid excretion levels seemed to correlate with large tumors or terminal stage. MIBG uptake was found in 16 after a diagnostic dose and in 1 only after a therapeutic dose. Surgery was performed on primary tumor in 18 and on distant metastase in 10. Iodine-131 MIBG therapy was performed in 11, among whom 9 were evaluable. Cumulative activity ranged from 100 to 711 mCi, in 1 to 6 courses. Symptomatic improvement occurred in 5 patients, stabilization was observed in 3 and tumor partial response in two, which lasted for 28 and 9 months, respectively terminating in a rapidly progressing disease with bone marrow involvement. Moderate myelosuppression occurred in 4 patients. Chemotherapy gave no response in 7 evaluable patients. Fourteen patients died with a median survival of 16 months from diagnosis of metastases (range 3-60). Response to therapy was poor and warrants further cooperative trials.

Topics: 3-Iodobenzylguanidine; Adolescent; Adrenal Gland Neoplasms; Adult; Aged; Antineoplastic Agents; Biomarkers, Tumor; Calcitonin; Calcitonin Gene-Related Peptide; Catecholamines; Child; Chromogranin A; Chromogranins; Female; Follow-Up Studies; Humans; Iodine Radioisotopes; Iodobenzenes; Male; Middle Aged; Neoplasm Metastasis; Neuropeptide Y; Pheochromocytoma; Phosphopyruvate Hydratase; Protein Precursors; S100 Proteins; Tomography, Emission-Computed; Treatment Outcome

Topics: Adrenal Gland Neoplasms; Adrenal Glands; Catecholamines; Choristoma; Diagnosis, Differential; Humans; Metanephrine; Neuropeptide Y; Pheochromocytoma

Plasma levels of chromogranin A + B, neuropeptide Y and catecholamines were analysed before, during and after surgery in seven patients with pheochromocytoma. The aim of the study was to determine the diagnostic sensitivity of these plasma amines and peptides, and to investigate their peroperative fluctuations. Chromogranin A + B in plasma was increased preoperatively in all patients, showed no significant increase during surgery, and normalized postoperatively. Neuropeptide Y, which alone can induce hypertension, was present in high levels in plasma from three patients preoperatively, increased further in four patients during surgery, and was postoperatively low in all patients. Fractionated plasma catecholamines were increased in five patients before surgery, increased in all patients during tumour dissection, and normalized postoperatively. It may be concluded that plasma chromogranin A + B exhibited as high a sensitivity for pheochromocytoma as fractionated urinary catecholamines in the patients studied.

Topics: Adrenal Gland Neoplasms; Adult; Aged; Catecholamines; Chromogranin A; Chromogranins; Epinephrine; Female; Humans; Intraoperative Period; Male; Middle Aged; Nerve Tissue Proteins; Neuropeptide Y; Norepinephrine; Pheochromocytoma

The regulation of the preproneuropeptide Y gene (NPY gene) by nerve growth factor (NGF) and second messenger systems in PC12 rat pheochromocytoma cells was studied by means of steady state NPY mRNA and nuclear run-on transcription analyses. Treatment of cells with 2.5S NGF increased the NPY mRNA abundance up to 100-fold over 1-6 days. Glucocorticoids (e.g. dexamethasone) potentiated by up to 3-fold the stimulation by NGF at early times (less than or equal to 7 h), but strongly suppressed it at later times (greater than or equal to 25 h). The response to NGF was blocked by cycloheximide, indicating a requirement for ongoing protein synthesis. Treatment of cells for 24-48 h with combinations of NGF, forskolin to elevate cAMP levels, and phorbol-12-myristate-13-acetate (PMA) to activate protein kinase C synergistically elevated NPY mRNA levels. The rate of NPY gene transcription in PC12 nuclei was increased by NGF, forskolin plus PMA, or NGF plus forskolin plus PMA, indicating that these regulators act at least in part at a transcriptional level. beta-Actin gene transcription also was elevated synergistically by forskolin and PMA. In summary, NPY gene transcription and NPY mRNA levels are controlled by multiple, potentially interacting regulatory systems. The striking antagonism between NGF and glucocorticoids may reflect the hormonal control of phenotypic choice during neural crest differentiation.

Topics: Adenosine Monophosphate; Adenylyl Cyclases; Adrenal Gland Neoplasms; Animals; Colforsin; Cyclic AMP; Glucocorticoids; Nerve Growth Factors; Neuropeptide Y; Pheochromocytoma; Phorbol Esters; Rats; RNA, Messenger; Tetradecanoylphorbol Acetate; Transcription, Genetic; Tumor Cells, Cultured

The presence of functional receptors for human atrial natriuretic hormone in human pheochromocytomas was recently reported. The present study reports the binding of hANH as measured by Scatchard analysis in 4 human adrenal glands and in 5 human pheochromocytomas. Binding assays using [3H]ANH revealed a single class of high-affinity binding sites for hANH in both tissues. Human pheochromocytomas present a lower number of binding sites than normal human adrenal gland (Bmax of 7.1 +/- 2.1 vs 33.6 +/- 6.9 fmol/mg protein, respectively). However, the decreased number of ANH receptors was not paralleled by modifications of tissular cyclic GMP (cGMP). Moreover, plasma hANH concentrations in 7 patients with pheochromocytomas (20.2 +/- 2.7 pmol/l) were statistically higher than those obtained in 25 normal control humans (8.1 +/- 0.6 pmol/l, p less than 0.001). We also demonstrated the presence of immunoreactive ANH in the tumour itself.

Topics: Adrenal Gland Neoplasms; Adrenal Glands; Aldosterone; Angiotensin II; Atrial Natriuretic Factor; Binding, Competitive; Chromatography, Ion Exchange; Cyclic GMP; Desoxycorticosterone; Dopamine; Humans; Neuropeptide Y; Pheochromocytoma; Receptors, Atrial Natriuretic Factor; Receptors, Cell Surface; Vasopressins

Neuron-specific enolase (NSE) is the isoform of enolase, a glycolytic enzyme found in the neuroendocrine system. Neuropeptide Y (NPY) is a peptide recently discovered in the peripheral and central nervous systems. Serum NSE and plasma NPY levels have been reported to be increased in some patients with pheochromocytoma. The authors evaluated whether the measurement of these molecules could help to discriminate between benign and malignant forms of pheochromocytoma. The NSE levels were normal in all patients with benign pheochromocytoma (n = 13) and elevated in one half of those with malignant pheochromocytoma (n = 13). Plasma NPY levels were on the average significantly higher in the malignant (177.1 +/- 38.9 pmol/l, n = 16) than in the benign forms of the disease (15.7 +/- 389 pmol/l, n = 24). However, there was no difference in the percentage of patients with elevated NPY levels. These results show that determination of serum NSE may be useful for distinguishing between malignant and benign pheochromocytoma; the measurement of plasma NPY is not useful for differentiating the two kinds of tumors.

Topics: Adrenal Gland Neoplasms; Adult; Aged; Biomarkers, Tumor; Diagnosis, Differential; Female; Humans; Male; Middle Aged; Neuropeptide Y; Pheochromocytoma; Phosphopyruvate Hydratase

To elucidate how the neuropeptide Y (NPY) gene is regulated by physiological/pharmacological changes in neural functions, the expression and regulation of the NPY gene were studied by measuring changes in the abundances of NPY and NPY mRNA in the adrenal gland and brain regions of rats in vivo and in PC12 rat pheochromocytoma cells after reserpine treatment. Long term treatment with reserpine in vivo, which causes hypotension and increased splanchnic nerve activity, induced prolonged increases in the abundance of NPY mRNA and putative NPY pre-mRNA, with concomitant increases in NPY, in the adrenal gland in a tissue-dependent manner but caused no changes in the abundance of beta-actin mRNA. Transection of the splanchnic nerves almost completely (76%) prevented the reserpine-induced increases in the abundance of NPY mRNA and NPY pre-mRNA, but denervation alone did not affect their steady state levels. These results suggested that increased activity of the splanchnic nerves regulates NPY gene expression positively in the adrenal gland, probably at the level of transcription. In PC12 cells, reserpine decreased the abundance of NPY mRNA directly, but nicotinic receptor activation increased its abundance transiently and the persistent membrane depolarization increased its abundance markedly. Thus, NPY gene expression is positively regulated by membrane depolarization via increased transsynaptic activation with reserpine.

Topics: Actins; Adrenal Gland Neoplasms; Adrenal Glands; Animals; Blotting, Northern; Cerebral Cortex; Chromatography, High Pressure Liquid; Denervation; Gene Expression Regulation; Male; Neuropeptide Y; Pheochromocytoma; Protein Precursors; Radioimmunoassay; Rats; Rats, Inbred Strains; Reserpine; RNA, Messenger; Tumor Cells, Cultured

The role of the clonidine suppression test and determination of the plasma concentration of noradrenaline and of neuropeptide Y (NPY) for the diagnosis and monitoring of bilateral pheochromocytoma was studied in a ten year old girl. Prior to tumor removal plasma concentration of noradrenaline (25.9 nmol/l) and NPY (14 pmol/l) were markedly elevated. Inhibition of central sympathetic activity by clonidine (150 micrograms orally) did not lower the plasma concentration of noradrenaline (70.4 nmol/l) and NPY (52.6 pmol/l). However in an adolescent boy with renal hypertension plasma concentration of noradrenaline and NPY fell following clonidine application (noradrenaline from 1.9 to 1.3 nmol/l, NPY from 3.6 to 2.4 pmol/l). After complete removal of the pheochromocytoma noradrenaline and NPY fell back to normal.

Topics: Adrenal Gland Neoplasms; Child; Clonidine; Female; Humans; Neoplasms, Multiple Primary; Neuropeptide Y; Norepinephrine; Pheochromocytoma

To investigate the release of neuropeptide Y (NPY) from the pheochromocytomas, we studied the relationship between the plasma and tumor tissue immunoreactive (IR) NPY concentrations in 13 patients with pheochromocytoma and measured the IR-NPY concentration in plasma samples obtained by catheter from several veins (jugular veins, superior vena cava, renal veins, adrenal veins and inferior vena cava) in 2 patients with pheochromocytoma. The plasma IR-NPY concentration in 13 patients with pheochromocytoma ranged from 118 to 1460 pg/ml and the concentration in 10 of 13 patients with pheochromocytoma was above 290 pg/ml (the upper limit of normal range). The tumor tissue IR-NPY ranged from 0.025 to 95.3 micrograms/g wet tissue. Plasma IR-NPY was parallel with tumor tissue IR-NPY in 13 cases of pheochromocytoma (r = 0.76, P less than 0.01). The highest concentration of IR-NPY was found in plasma obtained from the drainage vein from a tumor among the plasma samples obtained from several veins in 2 cases of pheochromocytoma. These findings indicate that in patients with pheochromocytoma, NPY is in most cases excessively released from the tumors into the systemic circulation and plasma IR-NPY in the periphery is increased.

Topics: Adrenal Gland Neoplasms; Chromatography, Gel; Chromatography, High Pressure Liquid; Humans; Neuropeptide Y; Pheochromocytoma; Radioimmunoassay

In order to develop an immunoradiometric assay for human neuropeptide (hNPY), a recently discovered and potent vasoconstrictor 36-amino-acid peptide, we used hNPY and some of its subpeptides to prepare monoclonal anti-NPY antibodies. Two monoclonal antibodies with high affinity for hNPY, that is affinity constants in the range of 10(10) mol/L-1, which respectively, reacted with the 9-18 portion and the 32-36 portion of hNPY were used in the immunoradiometric system. The assay was highly specific, NPY-related peptides such as pancreatic polypeptide and peptide YY not being detected. The lower limit of sensitivity was 0.5 pmol/L. In 303 normal subjects, plasma NPY concentrations were less than 0.5 pmol/L in 67%, 0.5 to 5.0 pmol/L in 25% and 5.1 to 30 pmol/L in the remaining 8%. A value of 7.5 pmol/L (95th percentile value in the normal group) was considered as the upper limit of normal. Among 111 patients with various neuroendocrine tumors, elevated plasma NPY concentrations were found in patients with pheochromocytomas and neuroblastomas, the highest plasma levels being found in patients with malignant pheochromocytomas. We conclude that patients with neuroendocrine tumors, especially secreting and or malignant tumors of the sympathochromaffin system, often have elevated plasma NPY concentrations.

Topics: Adolescent; Adult; Aged; Antibodies, Monoclonal; Biomarkers, Tumor; Carcinoid Tumor; Carcinoma; Child; Child, Preschool; Epitopes; Female; Humans; Infant; Infant, Newborn; Middle Aged; Neuroblastoma; Neuropeptide Y; Pheochromocytoma; Radioimmunoassay

By using monoiodinated radioligands of both intact neuropeptide Y (NPY) and of a long C-terminal fragment, NPY13-36, two subtypes of binding sites, which differ in affinity and specificity, have been characterized. The Y1 type of binding site, characterized on a human neuroblastoma cell line, MC-IXC, and a rat pheochromocytoma cell line, PC-12, binds NPY with a dissociation constant (Kd) of a few nanomolar but does not bind NPY13-36. The Y2 type of binding site, characterized on porcine hippocampal membranes and on another human neuroblastoma cell line, SMS-MSN, is of higher affinity and binds both NPY and NPY13-36. None of the binding sites distinguish between NPY and the homologous peptide YY (PYY). It is concluded that NPY/PYY-binding sites occur in two subtypes which may represent two types of physiological receptors.

Topics: Adrenal Gland Neoplasms; Animals; Binding Sites; Cell Membrane; Hippocampus; Humans; Neuroblastoma; Neuropeptide Y; Peptide Fragments; Pheochromocytoma; Rats; Receptors, Neuropeptide Y; Receptors, Neurotransmitter; Swine; Tumor Cells, Cultured

The expression of the potent vasoactive peptide neuropeptide Y (NPY) was studied in 16 clinically and/or histologically diagnosed human pheochromocytomas and 3 human neuroblastoma tumors. All tumors contained NPY in concentrations ranging from 21 pmol/g of tissue, similar to that found in normal adrenal tissue, to 91,000 pmol/g (median, 1,700 pmol/g). Three control tumors of Cushing's type did not contain NPY. An almost total proteolytic processing of pro-NPY to normal NPY was observed in the tumors (median, 93%; range, 72-100%). A positive correlation between the processing efficiency and the NPY content was also observed. The small amount of pro-NPY found in the tumors was characterized by "in vitro conversion" with endoproteinase Lys-C. In the tumor extracts, the majority of the NPY immunoreactivity, corresponding in size to the NPY standard, also behaved like synthetic NPY by high performance liquid chromatography and isoelectric focusing. As assessed by both its elution position in isoelectric focusing and its reaction with an antiserum specific for the COOH-terminal amidated sequence, the peptide produced by the tumors was found to be efficiently amidated, a modification which is essential for the biological activity of NPY. It is concluded that although only a subset of chromaffin cells express NPY, a very high number of pheochromocytomas and neuroblastomas produce correctly amidated and thus biologically active NPY in large amounts, and that this is of potential importance for tumor-related cardiovascular symptoms and for autocrine stimulation of tumor cells.

Topics: Adolescent; Adrenal Cortex; Adrenal Gland Neoplasms; Adult; Aged; Child; Child, Preschool; Chromatography, Gel; Chromatography, High Pressure Liquid; Humans; Isoelectric Focusing; Male; Middle Aged; Neuroblastoma; Neuropeptide Y; Pheochromocytoma; Protein Precursors; Radioimmunoassay

The synthesis and processing of the precursor for neuropeptide Y (NPY) were studied in 16 human and murine neuroendocrine cell lines. Eight of the cell lines, NS-20Y, PC12, LA-N-5, CHP-234, SMS-KCNR, SH-SY5Y, SMS-KCN, and BE(2)-M17, produced sufficient quantities to permit chromatographic characterization of the NPY immunoreactivity. Although the cell lines varied in the amount of NPY they produced, both within and between cell lines, they displayed a relatively constant pattern of posttranslational modifications. In contrast to observations in tumor extracts (M. M. T. O'Hare and T. W. Schwartz, Cancer Res., 49: 7010-7014, 1989), all cell lines studied contained a substantial amount of the intracellular NPY in the form of the unprocessed propeptide, 57% (range, 33-72%) as characterized by both gel filtrations (32 experiments in 8 cell lines) and "in vitro conversion" with endoproteinase Lys-C. In the majority, 4 of 6 cell lines studied, almost all of the NPY, which by size corresponded to the mature 36-amino acid form, was amidated as assessed by isoelectric focusing and by a radioimmunoassay specific for the COOH-terminal amide group of the peptide. Both the propeptide and smaller molecular forms of NPY were secreted from the cell cultures; however, proteolytic degradation in the tissue culture medium prevented a detailed, meaningful characterization of these peptides. It is concluded that many neuroendocrine cell lines, especially those derived from human neuroblastomas, express the NPY gene; the cells display a partly impaired dibasic processing capacity but they generally amidate the products efficiently.

Topics: Adrenal Gland Neoplasms; Animals; Chromatography, Gel; Chromatography, High Pressure Liquid; Humans; Isoelectric Focusing; Mice; Neuroblastoma; Neuropeptide Y; Pheochromocytoma; Protein Precursors; Protein Processing, Post-Translational; Radioimmunoassay; Tumor Cells, Cultured

A second antibody solid phase radioimmunoassay for measuring neuropeptide Y (NPY) in biological fluids is introduced. The sensitivity of the method allows also measurements of sub-normal levels of plasma/serum NPY, necessary for obtaining further information of possible biological actions of circulating NPY. Characteristics of the NPY assay procedure are presented together with clinical experiments forced upon a possible renal dependence for NPY. Our data show that NPY at least partly is eliminated by the kidneys. Whether the highly increased NPY levels in patients with renal failure also are involved in the hypertensive mechanisms in these patient groups has to be further evaluated.

Topics: Adrenal Gland Neoplasms; Adult; Body Fluids; Chromatography, High Pressure Liquid; Humans; Kidney; Kidney Failure, Chronic; Middle Aged; Neuropeptide Y; Pheochromocytoma; Radioimmunoassay; Reference Values

No reliable gross or microscopic features distinguish benign from malignant pheochromocytomas. The diagnosis of malignant pheochromocytoma is based solely on the presence of regional or distant metastases. This study evaluated the expression of neuropeptide Y messenger RNA (mRNA) in nine benign and 11 malignant pheochromocytomas and has found that neuropeptide Y mRNA was expressed in all nine benign tumors but in only four of 11 malignant tumors (P = .0084). These data suggest that the determination of neuropeptide Y expression in the evaluation of patients with pheochromocytoma may have prognostic significance.

Topics: Adrenal Gland Neoplasms; Blotting, Northern; Gene Expression Regulation, Neoplastic; Humans; Immunohistochemistry; Neuropeptide Y; Nucleic Acid Hybridization; Pheochromocytoma; RNA, Messenger

Pancreatic polypeptide (PP) and neuropeptide Y (NPY) belong to a family of regulatory peptides which hold a distinct tertiary structure, the PP-fold, even in dilute aqueous solution. High-affinity receptors, specific for both PP and NPY, are described on the rat phaeochromocytoma cell line, PC-12. The binding of [125I-Tyr36]PP to PC-12 cells was inhibited by concentrations of unlabeled PP which correspond to physiological concentrations of the hormone, 10(-11)-10(-9) mol/l. The affinity of the receptor for the neuropeptide, NPY, was 10(2)-times lower than that of the PP receptor. C-terminal fragments of both PP (PP24-36) and NPY (NPY13-36) were between 10(2)- and 10(3)-times less potent in displacing the radiolabeled 36-amino-acid peptides from their respective receptors. It is concluded that PC-12 cells are suited for structure-function studies of the PP-fold peptides and studies on the cellular events following cellular binding of PP-fold peptides.

Topics: Adrenal Gland Neoplasms; Animals; Binding, Competitive; Iodine Radioisotopes; Kinetics; Monoiodotyrosine; Neuropeptide Y; Pancreatic Polypeptide; Peptide Fragments; Pheochromocytoma; Rats; Receptors, Gastrointestinal Hormone; Receptors, Neuropeptide Y; Receptors, Neurotransmitter; Tumor Cells, Cultured

High plasma concentrations of neuropeptide Y (NPY) were found in a patient with bilateral adrenal phaeochromocytomas and medullary thyroid carcinoma associated with MEN IIa (32 pmol/l, normal less than 3.5 pmol/l). Both adrenal tumours contained and secreted NPY. Manipulation at operation produced a remarkable increase in plasma NPY concentrations (peak = 1631 pmol/l) coinciding with increases in plasma levels of catecholamines and arterial pressure. NPY was also shown to be present in thyroid tumour tissue: the concentration of NPY in tumour was 50-fold higher (0.9 nmol/g vs 0.004 nmol/g) than in adjacent normal thyroid tissue. It is possible that NPY from some phaeochromocytomas may contribute to hypertension during surgery.

Topics: Adrenal Gland Neoplasms; Epinephrine; Female; Humans; Middle Aged; Multiple Endocrine Neoplasia; Neuropeptide Y; Norepinephrine; Pheochromocytoma; Thyroid Neoplasms

To investigate the clinical usefulness of radio-immunoassay of neuropeptide Y (NPY), we measured plasma immunoreactive neuropeptide Y (IR-NPY) concentrations in normal subjects (n = 21), essential hypertensive patients (n = 33), patients with phaeochromocytoma (n = 7), patients with chronic renal disease with serum creatinine levels of less than 1.9 mg/dl (n = 5) and patients with chronic renal failure whose serum creatinine levels were greater than or equal to 1.9 mg/dl (n = 18, eight without haemodialysis and 10 undergoing maintenance haemodialysis), by radio-immunoassay. Plasma IR-NPY concentrations in patients with phaeochromocytoma (577 +/- 256 pg/ml, mean +/- s.d.) were significantly higher (P less than 0.001) than those in normal subjects (151 +/- 28 pg/ml), essential hypertensive patients (177 +/- 49 pg/ml) and patients with chronic renal disease with serum creatinine levels less than 1.9 mg/dl (198 +/- 71 pg/ml). Plasma IR-NPY concentrations in patients with chronic renal failure (without haemodialysis: 330 +/- 63 pg/ml; undergoing maintenance haemodialysis: 374 +/- 80 pg/ml) were also high. These results suggest that NPY is useful as one of the tumour markers of phaeochromocytomas. However, this study revealed that patients with chronic renal failure, without phaeochromocytoma also have increased plasma IR-NPY concentrations.

Topics: Adrenal Gland Neoplasms; Adult; Chromatography, Gel; Creatinine; Female; Humans; Hypertension; Kidney Failure, Chronic; Male; Middle Aged; Neuropeptide Y; Pheochromocytoma; Radioimmunoassay; Renal Dialysis

Neuropeptide Y (NPY) and its flanking peptide (CPON) were measured in extracts of pheochromocytomas (n = 26), ganglioneuroblastomas (n = 8), and other tumors (n = 95) and plasma (n = 38). NPY was present in high concentrations in the majority of the pheochromocytomas, but was absent in 3 tumors. Similar concentrations of NPY were measured using N- and C-terminal-directed antisera, and there was a good correlation between the concentrations of NPY immunoreactivity and CPON immunoreactivity in the same extracts. Gel permeation chromatography revealed that the separate peptides NPY and CPON were the major products. No significant amounts of a large mol wt precursor containing both immunoreactivities was found. Among the other tumors, 6 of 22 carcinoid tumors, 2 of 2 somatostatinomas, and 3 of 18 insulinomas contained detectable amounts of NPY. Plasma NPY concentrations were very low in normal subjects, and extraction and a 10-fold concentration step were necessary to establish the normal range (0.35-1.25 pmol/L). Plasma NPY concentrations were detectable in unextracted plasma (10 pmol/L) in 50% of patients with pheochromocytomas, but no correlation was found between plasma NPY concentrations and either plasma norepinephrine or epinephrine concentrations. These results indicate that 1) NPY and CPON are present in most, but not all, pheochromocytomas and ganglioneuroblastomas; 2) secretion of NPY immunoreactivity may be independent of that of catecholamines; 3) CPON immunoreactivity is present in plasma of patients with pheochromocytoma; 4) the majority of NPY and CPON immunoreactivities appears to be in the form of the separate free peptide, and there is very little precursor containing both immunoreactivities expressed in these tumors; and 5) NPY immunoreactivity is present in a variety of other endocrine tumors.

Topics: Chromatography, Gel; Chromatography, High Pressure Liquid; Endocrine System Diseases; Humans; Neuropeptide Y; Osmolar Concentration; Pheochromocytoma; Radioimmunoassay; Reference Values

The human neuropeptide Y (NPY) gene was isolated from a human genomic DNA library. The transcription unit spans approximately 8 kilobase pairs and is interrupted by three intervening sequences. The first exon contains only nontranslated DNA. The site where transcription initiates was determined by primer extension analysis using a primer derived from a human cDNA, pheochromocytoma RNA and avian myeloblastosis virus reverse transcriptase. A TATA-like sequence and a CAAT-like sequence occur 25 and 70 base pairs 5' to the transcription start site, respectively. The second exon begins with the initiator Met for preproNPY and extends to the Arg (residue 63) which precedes the Tyr-amide of mature NPY. The third exon contains the coding region for 27 amino acids, and the fourth exon codes for the terminal heptapeptide and the 3' nontranslated DNA. Transcriptional control elements were investigated by fusing 581 base pairs of the 5' sequences of the NPY gene to the promoterless structural gene for chloramphenicol acetyltransferase. NPY promoter activity was assayed by transfection of these hybrid constructions into CA-77 and PC12 cells followed by the determination of chloramphenicol acetyltransferase activity in cellular extracts. DNA sequences located within 530 bases of the start of transcription are sufficient for transient expression in the two neuronally derived cell lines examined.

Topics: Adrenal Gland Neoplasms; Amino Acid Sequence; Avian Myeloblastosis Virus; Base Sequence; Cloning, Molecular; DNA; DNA Restriction Enzymes; Gene Expression Regulation; Humans; Nerve Tissue Proteins; Neuropeptide Y; Pheochromocytoma; RNA-Directed DNA Polymerase; Transcription, Genetic

The occurrence of neuropeptide Y (NPY)-like immunoreactivity (LI) in the adrenal gland of several species as well as in tumor tissue and plasma from pheochromocytoma patients was investigated. NPY-LI was present in chromaffin cells of the adrenaline type in all species investigated except in the pig, as demonstrated by a colocalization of NPY-LI and the adrenaline-synthetizing enzyme phenylethanolamine N-methyltransferase (PNMT). NPY-LI in the adrenaline cells of the cat was clearly separated from the neurotensin-LI in the noradrenaline dopamine-beta-hydroxylase-positive, PNMT-negative cells. NPY-LI seems to co-exist with enkephalin-like material in the chromaffin cells. In addition, NPY-LI was present in nerves both within the adrenal cortex and medulla. The highest levels of NPY-LI were found in mouse and cat, while only a very low amount of NPY-LI was present in the pig adrenal. Characterization of the adrenal NPY-LI by reversed-phase high-performance liquid chromatography revealed that the main peak was similar to porcine NPY. In addition, two minor peaks of NPY-LI were present. High levels of NPY-LI were found in plasma and tumors from the pheochromocytoma patients. During manipulation of the tumors upon surgical removal, there was a marked increase in plasma NPY-LI in parallel with the raise in catecholamines and in blood pressure. At least two forms of NPY-LI were present in plasma and tumor extracts from pheochromocytoma patients with the main peak corresponding to porcine NPY. Since NPY exerts vasoconstrictor effects, it may be postulated that NPY contributes to the adrenal cardiovascular response and to the hypertension seen in pheochromocytoma patients.

Topics: Adrenal Gland Neoplasms; Adrenal Medulla; Animals; Cats; Chromatography, High Pressure Liquid; Enkephalins; Epinephrine; Fluorescent Antibody Technique; Guinea Pigs; Humans; Mice; Nerve Tissue Proteins; Neuropeptide Y; Neurotensin; Phenylethanolamine N-Methyltransferase; Pheochromocytoma; Radioimmunoassay; Rats; Species Specificity; Swine

Samples of adrenal medullary phaeochromocytoma from two patients were extracted in acid. The tissue concentrations of neuropeptide Y (NPY)-like immunoreactivity were found to be 0.94 and 8.11 micrograms/g. Plasma from these patients subjected to NPY radioimmunoassay after Sephadex G-75 gel filtration chromatography revealed circulating plasma concentrations of 1889 and 2079 pg/ml, compared to levels in normal plasma of less than 50 pg/ml, treated in the same way. Further characterisation of this plasma NPY-like immunoreactivity by HPLC showed this peptide to possess identical elution behaviour to human NPY standard. It is suggested that circulating NPY may be a contributory factor in the symptoms exhibited by some patients with adrenal medullary phaeochromocytoma.

Topics: Adrenal Gland Neoplasms; Chromatography, Gel; Female; Humans; Male; Middle Aged; Nerve Tissue Proteins; Neuropeptide Y; Pheochromocytoma; Radioimmunoassay

Cells from three human pheochromocytomas grown in monolayer culture for 21 days contained markedly greater amounts of neuropeptide Y (NPY) than freshly dissociated cells from the same tumors. The content of NPY did not consistently correlate with the extent of neurite outgrowth in the cultures and was not further increased in the presence of nerve growth factor. The content of NPY was decreased in the presence of 10(-5) M dexamethasone in two cultures. The findings suggest that human pheochromocytoma cultures may be useful in studies of cellular and molecular mechanisms regulating NPY production, and that these mechanisms may differ somewhat from those regulating production of other regulatory peptides in cultures of the same tumors.

Topics: Adrenal Gland Neoplasms; Cells, Cultured; Humans; Nerve Growth Factors; Nerve Tissue Proteins; Neuropeptide Y; Pheochromocytoma; Specimen Handling

Using a porcine neuropeptide Y directed radioimmunoassay it was shown that acid extracts of human phaeochromocytoma tumour tissue contained a neuropeptide Y-like peptide. Further fractionation and purification of this immunoreactivity showed that this human neuropeptide Y-like immunoreactivity was closely similar in molecular size and separation characteristics to porcine neuropeptide Y. The possible contribution of neuropeptide Y to the hypertension characterizing human phaeochromocytoma is discussed.

Topics: Chromatography, Gel; Cross Reactions; Humans; Nerve Tissue Proteins; Neuropeptide Y; Pancreatic Polypeptide; Peptide YY; Peptides; Pheochromocytoma; Radioimmunoassay; Vasoconstriction

High concentrations of a novel regulatory peptide, neuropeptide Y, have been identified within the PC12 rat phaeochromocytoma cell line by radioimmunoassay and chromatographic analysis. The cellular concentration of this peptide has been demonstrated to be predominantly influenced by the presence of nerve growth factor. After 14 days in culture in the presence of dexamethasone, NPY concentrations were 0.19 +/- 0.03 pmol/mg protein, whereas following similar exposure to nerve growth factor, concentrations were 3.44 +/- 0.19 pmol/mg protein. Addition of nerve growth factor to the dexamethasone-treated cells resulted in a rise in NPY concentration to 0.90 +/- 0.08 pmol/mg over 7 days, whereas addition of dexamethasone to the cells treated with nerve growth factor resulted in no further rise in NPY concentration. The concentrations of NPY far exceed those previously reported for neurotensin in PC12 cells, under the same culture conditions. These two peptides appear also to be regulated by different mechanisms.

Topics: Adrenal Gland Neoplasms; Animals; Cell Line; Chromatography, High Pressure Liquid; Dexamethasone; Kinetics; Nerve Growth Factors; Nerve Tissue Proteins; Neuropeptide Y; Pheochromocytoma; Radioimmunoassay; Rats

Human neuropeptide Y was isolated from acid extracts of adrenal-medullary phaeochromocytoma tissue. After (NH4)2SO4 fractionation, the neuropeptide Y-like immunoreactivity was purified from the resolubilized 80%-saturation-(NH4)2SO4 peptide-rich precipitate, by gel filtration, cation-exchange chromatography and reverse-phase high-pressure liquid chromatography. Amino acid analysis of the peptide revealed a composition almost identical with that of the pig peptide, the exception being the loss of one leucine residue and its replacement with methionine. Tryptic digestion of the peptide and subsequent amino acid analysis of the fragments further confirmed the identity of the peptide. Carboxypeptidase Y digestion of the (1-19)-peptide tryptic fragment has shown the methionine to be located at position 17 in human neuropeptide Y.

Topics: Adrenal Gland Neoplasms; Adrenal Medulla; Amino Acid Sequence; Carboxypeptidases; Chromatography, Gel; Chromatography, High Pressure Liquid; Chromatography, Ion Exchange; Humans; Nerve Tissue Proteins; Neuropeptide Y; Pheochromocytoma; Trypsin

The neuropeptide tyrosine precursor (pre-pro-NPY) messenger RNA (mRNA) has been localised in formaldehyde-fixed human phaeochromocytoma tissue using a sensitive in situ hybridisation procedure and a novel single-stranded cDNA probe. The reaction product was revealed by avidin-biotin-peroxidase complex and streptavidin-gold complex with silver enhancement. This technique may be applied for the determination of biosynthetic activity of endocrine and neuronal cell bodies. This is largely due to its rapidity by comparison with conventional autoradiographic procedures, to the permanence of the reaction product and to the sensitivity of the visualisation steps.

Topics: Adrenal Gland Neoplasms; DNA; DNA, Single-Stranded; Genes; Humans; Nerve Tissue Proteins; Neuropeptide Y; Nucleic Acid Hybridization; Pheochromocytoma; Protein Biosynthesis; Protein Precursors; RNA, Messenger; Transcription, Genetic

In vitro translation of the RNA isolated from a human pheochromocytoma demonstrated that this tumor contained a mRNA encoding a 10.5-kDa protein, which was immunoprecipitated with antiserum raised against porcine neuropeptide Y. Double-stranded cDNA was synthesized from total RNA and inserted into the Pst I site of pUC8. Transformants containing the neuropeptide Y cDNA were identified using the mixed hybridization probe d[A-(A,G)-(A,G)-T-T-(A,G,T)-A-T-(A,G)-T-A-(A,G)-T-G]. The probe sequences were based on the known amino acid sequence, His-Tyr-Ile-Asn-Leu, found in porcine neuropeptide Y. The nucleotide sequence of the cDNA was determined and contained 86 and 174 bases in the 5'- and 3'-untranslated regions, respectively. The coding sequence consisted of 291 bases, suggesting a precursor to neuropeptide Y that was 97 amino acids long (10,839 Da). The deduced amino acid sequence of the precursor suggested that there were at least two sites of proteolytic processing, which would generate three peptides having 28 (signal peptide), 36 (human neuropeptide Y), and 30 (COOH-terminal peptide) amino acid residues. A partial NH2-terminal sequence obtained by Edman degradation of the immunoprecipitated in vitro translation product identified the positions of methionine and leucine in the first 30 residues of the prepropeptide. A highly sensitive single-stranded complementary mRNA hybridization probe specific for neuropeptide Y mRNA was prepared using the bacteriophage SP6 promoter. This probe was used to identify a mRNA corresponding to neuropeptide Y of approximately 800 bases.

Topics: Adrenal Gland Neoplasms; Amino Acid Sequence; Animals; Base Sequence; Cell-Free System; DNA; Humans; Nerve Tissue Proteins; Neuropeptide Y; Nucleic Acid Hybridization; Pheochromocytoma; Protein Precursors; RNA, Messenger; Swine; Transcription, Genetic

Tumour tissue from nineteen patients with phaeochromocytomas and nine with ganglioneuroblastomas contained large numbers of neuropeptide Y (NPY) producing cells and extracts of these tumours had very high concentrations of immunoreactive NPY. Plasma NPY concentrations were also raised, averaging 460 pmol/l in patients with tumours of the sympathetic chain and 55 pmol/l in healthy controls. Since plasma NPY is straightforward to measure and relatively stable, its estimation may prove helpful as a screening tests for phaeochromocytoma.

Topics: Adrenal Gland Neoplasms; Chromatography, Gel; Ganglioneuroma; Humans; Nerve Tissue Proteins; Neuropeptide Y; Pheochromocytoma; Radioimmunoassay