Vasoactive Intestinal Peptide Production Research Articles

VIP and PACAP are autocrine factors that protect the androgen-independent prostate cancer cell line PC-3 from apoptosis induced by serum withdrawal.

1. In the present study, we describe the expression of the neuropeptides vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP) as well as their receptors in PC-3 cells, a human prostate cancer cell line. In addition, we have investigated their role in apoptosis induced by serum starvation. 2. By RT-PCR and immunocytochemistry assays, we have demonstrated the production of VIP and PACAP in PC-3 cells. 3. We have demonstrated by RT-PCR and binding assays the expression of common PACAP/VIP (VPAC(1) and VPAC(2)) receptors, but not PACAP-specific (PAC(1)) receptors. The pharmacological profile of [(125)I]-VIP binding assays was as follows: VPAC(1) antagonist=VPAC(1) agonist>VIP>VPAC(2) agonist (IC(50)=1.2, 1.5, 2.3 and 30 nM, respectively). In addition, both receptor subtypes are functional since VIP, PACAP-27 or VPAC(1) and VPAC(2) agonists all increased the intracellular levels of cAMP. 4. The expression of both peptides and their receptors is similar in serum-cultured and serum-deprived PC-3 cells. The treatment of serum-deprived PC-3 cells with exogenous VIP or PACAP-27 increases cell number and viability in a dose-dependent manner, as demonstrated by cellular counting and MTT assays. The increased cell survival is exerted through the VPAC(1) receptor, since a VPAC(1), but not VPAC(2), receptor agonist, mimics the effects and a VPAC(1) receptor antagonist blocks it. Moreover, VIP and PACAP-27 inhibit genomic DNA fragmentation in PC-3 cells triggered by serum starvation, and increase the immunoreactivity of the antiapoptotic protein bcl-2. 5. Our results suggest that VIP and PACAP are autocrine/paracrine factors that protect PC-3 cells from apoptosis through VPAC1 receptors.

IL-4 inhibits vasoactive intestinal peptide production by macrophages.

In schistosomiasis, eggs induce granulomas that have a vasoactive intestinal peptide (VIP) immunoregulatory circuit. This study explored the regulation of VIP production at sites of inflammation. Splenocytes from uninfected C57BL/6 mice expressed VIP mRNA and protein, which stopped following egg deposition. Eggs induce a Th2 response, suggesting that Th2 cytokines like interleukin (IL)-4 can regulate VIP. To address this issue, splenocytes from uninfected mice were incubated for 4 h with or without recombinant IL-4. IL-4 inhibited VIP mRNA expression. F4/80+ macrophages were the source of constitutively expressed VIP, subject to IL-4 regulation. In IL-4 knockout mice, splenic VIP production did not downmodulate during schistosome infection, suggesting that IL-4 is a critical cytokine regulating VIP production in wild-type mouse spleen. IL-4-producing granulomas in schistosomiasis made VIP. Experiments showed that granuloma VIP derived from F4/80- (nonmacrophage) cell populations, explaining this paradox. Granuloma F4/80+ cells from IL-4 knockout mice expressed VIP. Thus macrophages can make VIP, which is subject to IL-4 regulation. However, in the Th2 granulomas, other cell types produce VIP, which compensates for loss of macrophages as a source of this molecule.

Regulation of VIP production and secretion by murine lymphocytes

Vasoactive intestinal peptide (VIP) is a neuropeptide present in the lymphoid microenvironment with a multiplicity of actions. Two sources for VIP have been described in the immune system, the terminals present in central and peripheral lymphoid organs and the immune cells. Although VIP is synthesized by lymphocytes, there is no evidence demonstrating that VIP is released, and which stimuli are able to induce VIP production and secretion. In this study, we demonstrated for the first time, that agents that mediate important immune functions, such as proliferation and antigenic stimulation (Con A, LPS, and anti-TCR antibody), inflammation (LPS, TNFα, IL-6 and IL-1β) or apoptosis (dexamethasone) induce the production and release of VIP to the lymphoid microenvironment. We conclude that VIP is produced and secreted by lymphocytes and propose that during an immune response, the timely release of VIP within the lymphoid organs and peritoneum should influence the differentiation and/or downregulation of the ongoing response.

Cyclic Nucleotides and Vasoactive Intestinal Peptide Production in a Rabbit Model of Escherichia Coli Septicemia

Nitric oxide and vasoactive intestinal peptide (VIP) are potent vasodilators and postulated as inducers of hypotension. These mediators activate guanylate cyclase and adenylate cyclase, respectively, with subsequent biosynthesis of cyclic guanosine monophosphate (cGMP) and cyclic adenosine monophosphate (cAMP) producing vascular smooth muscle relaxation and vasodilatation. Cyclic nucleotides and VIP were evaluated during Escherichia coli septicemia in two groups of rabbits; 1) sepsis alone and 2) sepsis and a competitive inhibitor of nitric oxide synthase, NG-monomethyl-L-arginine. Arterial blood was obtained for determination of bacteremia, lactic acidemia, nucleotides, nitrites, and VIP levels. Significant bacteremia, endotoxemia, tachycardia, lactic acidosis, and hypotension occurred in all animals (P < 0.005). Circulating blood levels of cGMP, nitrites, cAMP, and VIP (P < 0.005) increased with development of shock. The NG-monomethyl-L-arginine treated animals had less cGMP, nitrites, cAMP, and VIP produced (P < 0.01). Plasma cGMP levels remained stable, suggesting that stimulated phagocytes in whole blood were responsible for increased cGMP levels. Infusion of VIP produced profound hypotension and lactic acidemia. Results of these experiments provide definitive evidence that nitric oxide and VIP are mediators during septic shock and their messengers are cGMP and cAMP, respectively. In addition, phagocytic stimulation with increased production of cGMP may initiate shock, with these mediators acting synergistically to prolong hypotension.

Cyclic Nucleotides and Vasoactive Intestinal Peptide Production in a Rabbit Model of Escherichia Coli Septicemia

Cyclic Nucleotides and Vasoactive Intestinal Peptide Production in a Rabbit Model of Escherichia Coli Septicemia

Induction of vasoactive intestinal peptide gene expression and prolactin secretion by insulin-like growth factor I in rat pituitary cells: evidence for an autoparacrine regulatory system.

The effects of recombinant human insulin-like growth factor I (IGF-I) on both vasoactive intestinal peptide (VIP) and PRL production and gene expression were studied using rat anterior pituitary cell cultures grown in serum-free defined medium. We also examined whether pituitary VIP could be involved in the PRL response to IGF-I and hence in a paracrine regulatory system. Exposure of cultured anterior pituitary cells to IGF-I (2.6 nM) for 3 h caused a significant decrease in both VIP content and media PRL. Treatment with IGF-I (from 0.65-5.2 nM) for 48 h increased VIP production and VIP messenger RNA (mRNA) accumulation, whereas only an increase in media and intracellular PRL content without changes in mRNA was observed. In all these experiments, IGF-I led to a decrease in both GH secretion and expression. Immunoglobulins G purified from VIP antiserum inhibited IGF-I-induced PRL release without affecting intracellular and mRNA levels. The inhibition of both GH secretion and gene expression induced by IGF-I was not blocked by VIP antiserum. In conclusion, these results indicate that IGF-I induces VIP gene expression, and its secretion and also increases PRL secretion. The effect of IGF-I on PRL release is specifically mediated by VIP through a paracrine or autocrine mechanism.

Induction of vasoactive intestinal peptide gene expression and prolactin secretion by insulin-like growth factor I in rat pituitary cells: evidence for an autoparacrine regulatory system

The effects of recombinant human insulin-like growth factor I (IGF-I) on both vasoactive intestinal peptide (VIP) and PRL production and gene expression were studied using rat anterior pituitary cell cultures grown in serum-free defined medium. We also examined whether pituitary VIP could be involved in the PRL response to IGF-I and hence in a paracrine regulatory system. Exposure of cultured anterior pituitary cells to IGF-I (2.6 nM) for 3 h caused a significant decrease in both VIP content and media PRL. Treatment with IGF-I (from 0.65-5.2 nM) for 48 h increased VIP production and VIP messenger RNA (mRNA) accumulation, whereas only an increase in media and intracellular PRL content without changes in mRNA was observed. In all these experiments, IGF-I led to a decrease in both GH secretion and expression. Immunoglobulins G purified from VIP antiserum inhibited IGF-I-induced PRL release without affecting intracellular and mRNA levels. The inhibition of both GH secretion and gene expression induced by IGF-I was not blocked by VIP antiserum. In conclusion, these results indicate that IGF-I induces VIP gene expression, and its secretion and also increases PRL secretion. The effect of IGF-I on PRL release is specifically mediated by VIP through a paracrine or autocrine mechanism.

Serotonin depletion by p-chlorophenylalanine decreases VIP mRNA in the suprachiasmatic nucleus.

The influence of serotonergic inputs on the levels of vasoactive intestinal peptide (VIP) mRNA in the rat suprachiasmatic nucleus (SCN) was examined by in situ hybridization combined with morphometrical analysis. Depletion of serotonin by p-chlorophenylalanine methyl ester (300 mg/kg i.p. daily for 3 days) caused a marked decrease in VIP mRNA signal levels in the ventrolateral part of the SCN. This finding suggests that serotonergic input controls VIP production at the mRNA level.

Acquired megacolon is associated with alteration of vasoactive intestinal peptide levels and acetylcholinesterase activity

Based upon previous morphologic studies, we hypothesized that the development of acquired megacolon was associated with abnormalities of enteric neurotransmitter concentrations and enzymatic activities. Specimens were obtained at surgery from patients with normal descending-sigmoid colon ( n = 13) and patients with sigmoid megacolon ( n = 6; defined by radiologic measurement). Radioimmunoassays were used to measure the non-adrenergic, non-cholinergic inhibitory neuropeptide, vasoactive intestinal peptide, and the non-adrenergic, non-cholinergic excitatory neuropeptide, substance P, while spectrophotometric assays were used to quantitate acetylcholinesterase activity and choline acetyltransferase activity. There were significantly decreased concentrations of vasoactive intestinal peptide and decreased acetylcholinesterase activity in muscularis externa from patients with acquired megacolon. In megacolon, vasoactive intestinal peptide-containing nerve fibers appeared to be diminished in circular and longitudinal smooth muscle, and immunostaining of nerve cell bodies in the plexus submucosus externus appeared diminished. These results suggest the hypothesis that production of vasoactive intestinal peptide is altered allowing secondary colonic hypertrophy to develop from prolonged cholinergic nerve-mediated contractions of circular smooth muscle. As a corollary to this hypothesis, colonic dilatation might result from prolonged contraction of longitudinal smooth muscle.

Vasoactive intestinal peptide (VIP), a putative neurotransmitter of nonadrenergic, noncholinergic (NANC) inhibitory innervation and its relevance to therapy

Nonadrenergic, noncholinergic (NANC) innervation, the third division of the autonomic nervous system, has both inhibitory and excitatory parts. The excitatory part received only limited attention. Substance P has been suggested to be the neurotransmitter of the excitatory part. The NANC-inhibitory innervation has recently been studied in detail. Although the neurotransmitter has not been conclusively identified, a substantial body of evidence exists to support vasoactive intestinal peptide (VIP) as the neurotransmitter. VIP is widely distributed in the body. Reports show that this innervation in animals and man plays a significant role in both health and disease. Pathological conditions could result from either an increase or decrease in VIP production. An absence of VIP-producing neurons has been identified to be responsible for Hirschsprung's disease in the alimentary system and hyperactive airways in the respiratory system. An increase in VIP production is associated with chronic water diarrhea syndrome in humans. Taking these factors into consideration, various therapeutic measures are suggested with the use of VIP or its antagonists.

Studies toward the biosynthesis of vasoactive intestinal peptide (VIP)

In view of the potential biological importance of VIP, we have begun to examine the regulation of its biosynthesis. For this purpose we have, as a first step, searched for an enriched source of VIP biosynthesis. By a combination of chromatographic procedures and radioimmunoassays we discovered an as yet unknown source for VIP production, namely a human buccal tumor, containing 0.67±0.05 ng VIP/μg protein which is greater than the richest source in brain (the cerebral cortex). Thus, we decided to use the tumor tissue for VIP-mRNA purification and characterization. To identify VIP-mRNA we are using as hybridization probes, synthetic oligodeoxynucleotides with relatively unambiguous nucleotide sequence complementary to the predicted VIP-mRNA sequence. These probes are synthesized, using the deoxynucleoside phosphoramidite approach, to a length of 17 bases each, and contain all the possible DNA sequences according to the genetic code. These specific probes are then radioactively labelled using the reaction catalyzed by the enzyme polynucleotide kinase and afterwards hybridized to mRNA, which had been resolved on denaturing agarose gels. Employing this approach, we identified a single putative VIP-mRNA band which was then partially purified by sucrose gradient centrifugation. Upon in vitro translation in a rabbit reticulocyte lysate cell free system, this mRNA was found to code for VIP immunoreactive proteins. In conclusion, our studies suggest the existence of high molecular weight precursors to VIP cross-reactive with anti-VIP antibodies, that are coded for by a partially purified mRNA containing VIP sequences.

Cellular localization of a vasoactive intestinal peptide in the mammalian and avian gastrointestinal tract.

Immunohistochemical studies using an antiserum to a pure porcine vasoactive intestinal peptide, possessing no cross reactivity against the related hormones glucagon, secretin, and gastrin-inhibitory peptide, revealed a wide distribution of vasoactive intestinal peptide cells throughout the entire length of the mammalian and avian gut. The highest numbers of cells were present in the small intestine and more particularly in the large intestine in all species investigated. Three types of endocrine cell in the mammalian gut are sufficiently widely distributed to be considered as the sites for production of vasoactive intestinal peptide. In the avian gut there are only two identifiable cell types. Sequential immunofluorescence and silver staining showed, in the bird, that the enterochromaffin (EC) cell was not responsible. This procedure could not be used in our mammalian gut samples but here serial section immunofluorescence for enteroglucagon and vasoactive intestinal peptide indicated that the two cells were not identical and that each was differently localized in the mucosa. These results leave the D cell of the Wiesbaden classification as the most likely site for the production of vasoactive intestinal peptide. The final identification must come from successful immune electron cytochemistry but this has not yet been achieved.