vasoactive-intestinal-peptide and Glioma

Emerging evidence have suggested that calorie restriction (CR) is a reliable method to decrease cancer development since it produces changes in tumor microenvironment that interfere with cell proliferation, tissue invasion, and formation of metastases. Studies on the role of pituitary adenylate cyclase-activating polypeptide (PACAP) and vasoactive intestinal peptide (VIP) in cancer cells indicate that their influence on cell growth is either cell type specific or dependent on culture conditions. Evidence showing the effect of PACAP and VIP in glioma cells grown under conditions mimicking CR are currently unavailable. Therefore, we explored the effects of both PACAP and VIP in C6 glioma cells either grown in a normal growth medium or exposed to serum starvation, to resemble an acute condition of CR. Cell viability, expression of proteins related to cell proliferation (cyclin D1), apoptosis (Bcl2, p53, and cleaved caspase-3), and cell malignancy (GFAP and nestin) were assessed by MTT assay, immunoblot, and immunolocalization, respectively. Results demonstrated that CR significantly decreased cell proliferation, reduced levels of cyclin D1 and Bcl2, and increased the expression of p53 and cleaved caspase-3. Surprisingly, all of these CR-driven effects were further exacerbated by PACAP or VIP treatment. We also found that PACAP or VIP prevented GFAP decrease caused by CR and further reduced the expression of nestin, a prognostic marker of malignancy. In conclusion, these data demonstrate that PACAP and VIP possess antiproliferative properties against glioma cells that depend on the specific culture settings, further supporting the idea that CR might offer new avenues to improve peptide-oriented glioma cancer treatment.

Topics: Animals; Apoptosis Regulatory Proteins; Cell Line, Tumor; Cell Proliferation; Culture Media, Serum-Free; Cyclin D1; Glial Fibrillary Acidic Protein; Glioma; Nestin; Pituitary Adenylate Cyclase-Activating Polypeptide; Rats; Transcription, Genetic; Tumor Suppressor Protein p53; Vasoactive Intestinal Peptide

Pituitary adenylate cyclase-activating polypeptide (PACAP) and vasoactive intestinal peptide (VIP) exert their actions via common receptors, VPAC1 and VPAC2, which are equally sensitive to both peptides, while PACAP stimulates its specific PAC1-type receptors. Both peptides potently stimulate cAMP production in different biological systems. In the present article, we examined the effects of PACAP and VIP on cAMP formation in C6 rat glioma cells used between passages 12-28 (early) and 120-136 (late). In the presence of the PDE inhibitor IBMX (0.1 mM), PACAP (0.1 microM) and VIP (1 microM) strongly stimulated cAMP synthesis in C6 cells in early passages, but not in C6 cells in late passages. In contrast, forskolin (10 microM), a direct activator of adenylyl cyclase, and isoprenaline (10 microM), a beta-adrenergic receptor agonist, strongly stimulated cAMP production in both early and late C6 cell passages. Concentration-dependent studies carried out in early passages with PACAP-38, PACAP-27, mammalian and chicken VIPs, and PHI/PHM peptides (1-5 microM) revealed that both forms of PACAP produced strong cAMP accumulation, VIP peptides were less effective than PACAP, while the cAMP effects of PHI/PHM peptides were noticeable only at the highest doses tested. These results suggest that C6 glioma cells in early passages possess functional PAC1 and possibly VPAC-type receptors, but either the density of PACAP/VIP receptors progressively declines or the PACAP/VIP receptor-Gs protein coupling becomes less effective through culture passages.

Topics: Animals; Cell Line, Tumor; Cellular Senescence; Colforsin; Cyclic AMP; Glioma; Isoproterenol; Pituitary Adenylate Cyclase-Activating Polypeptide; Rats; Time Factors; Vasoactive Intestinal Peptide

Glia cell line-derived neurotrophic factor (GDNF), a recently cloned member of the transforming growth factor-beta (TGF-beta) superfamily, has been implicated in the survival, morphological and functional differentiation of midbrain dopaminergic neurons and motoneurons in vitro and in vivo. The factor may thus have utility in the treatment of various human neurodegenerative disorders. Mechanisms regulating expression of GDNF in normal and diseased brain as a possible means to increase the local availability of GDNF are only beginning to be explored. We have established and employed a competitive reverse transcriptase-polymerase chain reaction (RT-PCR) to study and compare levels of expression of GDNF mRNA in several cell types and to investigate its regulation. GDNF expression was clearly evident in primary cultured astrocytes, the glioma B49 and C6 cell, but less pronounced in the Schwannoma RN22 cell lines. Little or no signal could be observed in neuroblastoma cell lines (IMR32, LAN-1) or the pheochromocytoma cell line PC12, emphasizing the glial character of this factor. Using the C6 cell line we found that fibroblast growth factor-2 (FGF-2; bFGF) can increase GDNF mRNA levels, whereas FGF-1, platelet-derived growth factor (PDGF), and vasoactive intestinal polypeptide (VIP) are apparently ineffective. Several other factors (forskolin, kainic acid, triiodothyronine dexamethasone, GDNF, TGF-beta 1, and interleukin-6) appear to have slightly negative effects on GDNF mRNA levels at the concentrations tested. To further explore the relationship between FGF-2 and GDNF, we also addressed the question whether GDNF, like FGF-2, may have an effect on C6 cell proliferation. We conclude that (1) glial and glial tumor cells, rather than neuronal cell lines, express GDNF, (2) that FGF-2 has a prominent inductive effect on GDNF expression and (3) that GDNF stimulates C6 cell proliferation. Finally, these data suggest that neurotrophic actions of FGF-2 in mixed glial-neuronal cell cultures might be mediated in part by GDNF.

Topics: Animals; Astrocytes; Brain Neoplasms; Cell Division; Colforsin; Dexamethasone; Fibroblast Growth Factor 1; Fibroblast Growth Factor 2; Gene Expression Regulation, Neoplastic; Glial Cell Line-Derived Neurotrophic Factor; Glioma; Humans; Interleukin-6; Kainic Acid; Neoplasm Proteins; Nerve Growth Factors; Nerve Tissue Proteins; Neuroblastoma; Neurons; Organ Specificity; Pheochromocytoma; Platelet-Derived Growth Factor; Polymerase Chain Reaction; Rats; Recombinant Proteins; RNA, Messenger; RNA, Neoplasm; Triiodothyronine; Tumor Cells, Cultured; Vasoactive Intestinal Peptide

Vasoactive intestinal peptide (VIP) is a potent mediator of gastrointestinal, nervous, pulmonary, vascular and immune functions. A cDNA was obtained from human HT29 intestinal epithelial cells and found to encode a 457 amino acid, 52 kDa VIP receptor. Transfection of the cDNA into COS-7 cells and 293 cells resulted in expression of specific saturable binding of VIP with a Kd of 0.8 nM, and induction of increases in intracellular cAMP by VIP with an EC50 of 1 nM. The human VIP receptor is homologous to other G protein-coupled receptors of the secretin-parathyroid hormone receptor family. A 2.8 kb transcript was detected in human lung, HT29 cells and Raji B-lymphoblasts with weaker expression in human brain, heart, kidney, liver and placenta.

Topics: Amino Acid Sequence; Animals; Base Sequence; Cell Line; Cloning, Molecular; Cyclic AMP; Gene Expression; Glioma; Humans; Hybrid Cells; Kinetics; Mice; Molecular Sequence Data; Neuroblastoma; Oligodeoxyribonucleotides; Polymerase Chain Reaction; Rats; Receptors, Gastrointestinal Hormone; Receptors, Vasoactive Intestinal Peptide; Recombinant Proteins; RNA, Messenger; Sequence Homology, Amino Acid; Transfection; Vasoactive Intestinal Peptide

The interaction between two neuropeptides, VIP and TRH, was studied. The TRH receptor binding ability was examined using intact GH3 cells and its membrane fraction. The TRH binding ability decreased when intact cells were preincubated with VIP, forskolin or db-cAMP, but not when the membrane fraction was treated with these agents. The binding was reduced only when the membrane fraction was treated with catalytic subunit of cAMP dependent protein kinase (A-kinase). These results indicate that the binding ability of TRH receptors, which are linked to inositol phospholipid metabolism, is suppressed when A-kinase increases due to activation of the adenylate cyclase or its dependent system. This, in turn, suggests the presence of a communication via a cytoplasmic factor (probably A-kinase) between the two principal second messenger systems in the cell.

Topics: Animals; Bucladesine; Cell Line; Colforsin; Glioma; Kinetics; Phorbol 12,13-Dibutyrate; Receptors, Neurotransmitter; Receptors, Thyrotropin-Releasing Hormone; Thyrotropin-Releasing Hormone; Vasoactive Intestinal Peptide

Topics: Adenosine; Adenosine-5'-(N-ethylcarboxamide); Adenylyl Cyclases; Animals; Calcium; Cell Line; Central Nervous System; Cyclic AMP; Glioma; Guanylyl Imidodiphosphate; Inositol Phosphates; Models, Biological; Neurotransmitter Agents; Phenylisopropyladenosine; Signal Transduction; Vasoactive Intestinal Peptide

Vasoactive intestinal peptide (VIP) and secretin are two related peptides that activate adenylate cyclase on membranes of striatal neurons and glial cells from embryonic mouse brain grown in primary culture. On the two cell types, the maximal activation that could be induced by secretin was only 40% above basal activity, which represented less than 15% of the maximal effect obtainable with VIP. From competition experiments performed on glial cells and the neuroblastoma X glioma hybrid, NG 108-15, a cell line known to possess both VIP and secretin sensitive-adenylate cyclase, we demonstrate that secretin does not activate VIP receptors. Furthermore, secretin has an apparent high affinity (EC50 10(-8) M) for its receptors on striatal neurons and NG 108-15 whereas an apparent low affinity (EC50 7 X 10(-6) M) was found on striatal glial cells. This suggests the existence of either two distinct secretin receptors or a desensitized form.

Topics: Adenylyl Cyclases; Animals; Cell Line; Corpus Striatum; Cyclic AMP; Embryo, Mammalian; Enzyme Activation; Glioma; Hybrid Cells; Mice; Neuroblastoma; Neuroglia; Neurons; Receptors, Cell Surface; Receptors, G-Protein-Coupled; Receptors, Gastrointestinal Hormone; Receptors, Vasoactive Intestinal Peptide; Secretin; Vasoactive Intestinal Peptide

Extracts of neuroblastoma X glioma hybrid cells 108CC15 and their parental lines were investigated for the presence of vasoactive intestinal polypeptide (VIP). With the aid of a radioimmunoassay and a receptor binding assay, VIP activity was found in the hybrid cells, to a lesser extent in neuroblastoma cells, but not in glioma cells. These results suggest a neuronal function of VIP. Since the hybrid cells also contain acetylcholine and opioids, they may be useful in studies of co-release of neurohormones.

Topics: Animals; Cell Line; Cells, Cultured; Gastrointestinal Hormones; Glioma; Hybrid Cells; Mice; Neoplasms, Experimental; Neuroblastoma; Rats; Vasoactive Intestinal Peptide

Topics: Cell Line; Cells, Cultured; Drug Resistance; Floxuridine; Glioma; Humans; Mercaptopurine; Morphine Dependence; Neoplasms, Experimental; Nervous System Diseases; Neuroblastoma; Neurophysiology; Thioguanine; Vasoactive Intestinal Peptide