Presence Of Vasoactive Intestinal Peptide Research Articles

Targeting first trimester trophoblast cell metabolism modulates its susceptibility to Zika virus infection.

In the last 15 years Zika virus (ZIKV) caused several outbreaks of increasing scale in Micronesia, South Pacific islands, and more recently in the Caribbean and South America. The severity of the clinical presentation in neonates from pregnant women infected with ZIKV during the last outbreak supports the relevance of unraveling the mechanism of infection and viral persistence in the placenta with local viral isolates. Here, we investigated the relevance of trophoblast metabolic rewiring for viral multiplication and the role of the vasoactive intestinal peptide (VIP) as an endogenous factor associated with placental restriction to ZIKV infection at early pregnancy. Our in vitro model demonstrated that ZIKV triggers metabolic rewiring in first trimester cytotrophoblast-derived cells by increasing glucose utilization as fuel to sustain its replication, decreasing long-chain polyunsaturated fatty acid uptake, and promoting lipid droplets accumulation to favor its multiplication. Of note, variations in nutrient availability modulated viral spread in trophoblast cultures. The presence of VIP during trophoblast infection impaired ZIKV infective particle production and viral replication, restoring cell migration and metabolism. Moreover, the blockade of endogenous VIP signaling increased viral particle production and the viral entry receptor AXL expression. These results highlight the potential role of VIP as an endogenous antiviral factor related to trophoblast cell permissiveness to ZIKV infection at early pregnancy.

Enterochromaffin Cell-Enriched Monolayer Platform for Assaying Serotonin Release from Human Primary Intestinal Cells.

Enteroendocrine (EE) cells within the intestinal epithelium produce a range of hormones that have key roles in modulating satiety and feeding behavior in humans. The regulation of hormone release from EE cells as a potential therapeutic strategy to treat metabolic disorders is highly sought after by the pharmaceutical industry. However, functional studies are limited by the scarcity of EE cells (or surrogates) in both in vivo and in vitro systems. Enterochromaffin (EC) cells are a subtype of EE cells that produce serotonin (5HT). Here, we explored simple strategies to enrich EC cells in in vitro monolayer systems derived from human primary intestinal stem cells. During differentiation of the monolayers, the EC cell lineage was significantly altered by both the culture method [air-liquid interface (ALI) vs submerged] and the presence of vasoactive intestinal peptide (VIP). Compared with traditional submerged cultures without VIP, VIP-assisted ALI culture significantly boosted the number of EC cells and their 5HT secretion by up to 430 and 390%, respectively. The method also increased the numbers of other subtypes of EE cells such as L cells. Additionally, this method generated monolayers with enhanced barrier integrity, so that directional (basal or apical) 5HT secretion was measurable. For all donor tissues, the enriched EC cells improved the signal-to-background ratio and reliability of 5HT release assays. The enhancement in the 5HT secretion behavior was consistent over time from a single donor, but significant variation in the amount of secreted 5HT was present among tissues derived from five different donors. To demonstrate the utility of the EC-enriched monolayer system, 13 types of pungent food ingredients were screened for their ability to stimulate 5HT secretion. Curcumin found in the spice turmeric derived from the Curcuma longa plant was found to be the most potent secretagogue. This EC-enriched cell monolayer platform can provide a valuable analytical tool for the high-throughput screening of nutrients and gut microbial components that alter the secretion of 5HT.

Vasoactive intestinal peptide is required in the maintenance of immune regulatory competency of immune regulatory monocytes.

Dysfunction of the immune regulatory system plays an important role in the pathogenesis of rheumatoid arthritis (RA). Vasoactive intestinal peptide (VIP) has multiple bioactivities. This study aims to investigate the role of VIP in the maintenance of the immune regulatory capacity of monocytes (Mos). Human peripheral blood samples were collected from RA patients and healthy control (HC) subjects. Mos and CD14+ CD71- CD73+ CD25+ regulatory Mos (RegMos) were isolated from the blood samples and characterized by flow cytometry. A rat RA model was developed to test the role of VIP in the maintenance of the immune regulatory function of Mos. The results showed that RegMos of HC subjects had immune suppressive functions. RegMos of RA patients expressed less interleukin (IL)-10 and showed an incompetent immune regulatory capacity. Serum levels of VIP were lower in RA patients, which were positively correlated with the expression of IL-10 in RegMos. In-vitro experiments showed that the IL-10 mRNA decayed spontaneously in RegMos, which could be prevented by the presence of VIP in the culture. VIP suppressed the effects of tristetraprolin (TTP) on inducing IL-10 mRNA decay in RegMos. Administration of VIP inhibited experimental RA in rats through restoring the IL-10 expression in RegMos. RegMos have immune suppressive functions. VIP is required in maintaining IL-10 expression in RegMos. The data suggest that VIP has translational potential in the treatment of immune disorders such as RA.

Vasoactive intestinal peptide maintains the nonpathogenic profile of human th17-polarized cells.

The cytokine microenvironment modulates CD4 T cell differentiation causing the shift of naïve CD4 T cells into different cell subsets. This process is also regulated by modulators such as vasoactive intestinal peptide (VIP), a neuropeptide with known immunomodulatory properties on CD4 T cells that exert this action through specific receptors, vasoactive intestinal peptide receptor (VPAC)1 and VPAC2. Our results show that the pattern of VIP receptors expression ratio is modified during Th17 differentiation. In this report, we evaluate the capacity of VIP to modulate naïve human cells into Th17 cells in vitro by analyzing their functional phenotype. The presence of VIP maintains the nonpathogenic profile of Th17-polarized cells, increases the proliferation rate, and decreases their Th1 potential. VIP induces the upregulation of the STAT3 gene interaction with the VPAC1 receptor during the onset of Th17 differentiation. Moreover, RAR-related orphan receptor C (RORC), RAR-related orphan receptor A (RORA), and interleukin (IL)-17A genes are upregulated in the presence of VIP through interaction with VPAC1 and VPAC2 receptors. Interestingly, VIP induces the expression of the IL-23R gene through interaction with the VPAC2 receptor during the expansion phase. This is the first report that describes the differentiation of naïve human T cells to Th17-polarized cells in the presence of VIP and demonstrates how this differentiation regulates the expression of the VIP receptors.

Immunohistochemical evidence for the presence of a Vasoactive Intestinal Peptide, Neuropeptide Y, and Substance P, in rat adrenal cortex after acute heat stress

Immunohistochemistry revealed the presence of Vasoactive Intestinal Peptide (VIP), Neuropeptide Y (NPY), and the absence of Substance P (SP) immunoreactivity in the rat adrenal cortex. VIP- and NPY-immunoreactivity were detected in nerve fibers around the small blood vessels projecting into the capsule and cortical zones surrounding blood vessels and cortical cells. After acute heat stress, VIP- and NPY-immunoreactivities in the nerve fibers were reduced, probably as a result of the release of these peptides.

Expression of corticotropin releasing factor (CRF) in the enteric nervous system of the jejunum of sheep

 Corticotropin releasing factor (CRF), a 41-amino acid neuropeptide widely distributed in the mammalian central nervous system, has been shown to influence several gastrointestinal functions. Recent studies show that CRF released locally from enteric nerves may also underlie alterations in gut function. In this study, immunohistochemisty was applied to demonstrate the presence of CRF in the jejunum of sheep. Using double immunohistochemical staining the co-localization of CRF with vasoactive intestinal peptide (VIP), galanin, tyrosine hydroxylase (TH), neuropeptide Y (NPY) and substance P (SP) was evaluated. The presence of CRF was detected in myenteric neurons (3.6 ± 0.9%) as well as in submucous neurons (10.5 ± 1.2%). In the ovine jejunum different numbers of CRF-expressing nerve fibres were detected in myenteric ganglia, submucous ganglia, circular smooth muscle layer, lamina muscularis mucosae and between mucosal glands. None of the CRF-positive enteric neurons and CRF-positive nerve fibres exhibited the presence of TH. CRF-immunoreactive (IR) myenteric neurons widely co-expressed VIP and/or NPY. A minor population of CRF-IR myenteric neurons additionally co-stored SP. Galanin was not present in CRF-IR myenteric neurons. The presence of VIP was observed in the vast majority of CRF-positive submucous neurons. Moderate numbers of CRF-IR sumbucous neurons co-expressing galanin or NPY were also found. The presence of SP in CRF-positive submucous neurons was noted only incidentally. In the circular smooth muscle layer CRF-IR/VIP-IR, CRF-IR/NPY-IR as well as CRF-IR/SP-IR nerve fibres were present. In the mucosal layer of the ovine jejunum CRF-IR nerve fibres co-stored additionally VIP, galanin, NPY or SP. This present study provides for the first time evidence that CRF present in different subclasses of enteric neurons may influence certain activities of the ovine jejunum. Co-localization studies indicate that VIP, galanin, SP and NPY functionally co-operate with CRF in the jejunum of the sheep.  

Neurotransmitter Influence on Human Meibomian Gland Epithelial Cells

A striking characteristic of the human meibomian gland is its rich sensory, sympathetic, and parasympathetic innervation, yet the functional relevance of these nerve fibers remains unknown. Acting on the hypothesis that neurotransmitters are released in the vicinity of the gland, act on glandular receptors, and influence the production, secretion, and/or delivery of meibomian gland secretions to the ocular surface, the goal in this study was to begin to determine whether neurotransmitters influence the meibomian gland. Immortalized human meibomian gland epithelial (SLHMG) cells were examined for the presence of vasoactive intestinal peptide (VIP) and muscarinic acetylcholine (mACh) receptor transcripts and proteins. Cells were also exposed to VIP, carbachol, forskolin, and/or 3-isobutyl-1-methylxanthine (IBMX) to determine whether these agents, alone or in combination, modulate the adenylyl cyclase pathway, the accumulation of intracellular free calcium ([Ca2+]i), or cell proliferation. Results demonstrate that SLHMG cells transcribe and translate VIP and mACh receptors; VIP, with either IBMX or forskolin, activates the adenylyl cyclase pathway, and the effect of VIP and forskolin together is synergistic; both VIP and carbachol increase intracellular [Ca2+] in SLHMG cells; and VIP with forskolin stimulates SLHMG cell proliferation. This study shows that parasympathetic neurotransmitters and their agonists influence the function of human meibomian gland epithelial cells. It remains to be determined whether this action alters the production, secretion, and/or delivery of meibum to the ocular surface.

Presence of VIP, CGRP, SP neuropeptides and PAF-AH enzyme in porcine colostrum

The aim of this study was to examine levels of vasoactive intestinal peptide (VIP), calcitonin gene-related peptide (CGRP), substance P (SP) and platelet activating factor-acetylhydrolase (PAF-AH) in porcine serum and colostrum. Samples of porcine blood were obtained from six multiparous sows during farrowing and post-lactation. Porcine colostrum was obtained from lactating sows at parturition. Neuropeptides were measured by competitive enzyme immunoassay using streptavidin-conjugated horseradish peroxidase (VIP) and acetyl cholinesterase-peptide conjugates (SP and CGRP) as tracers. PAF-AH activity was measured by a continuous kinetic assay using 2-thio PAF as substrate. The presence of VIP, CGRP, SP and PAF-AH activity in both porcine serum and colostrum was confirmed. The concentrations of VIP, CGRP and PAF-AH in serum and colostrum were comparable, suggesting passive rather than active secretion in the colostral milk. A significant higher concentration of SP was found in porcine colostrum. This could be explained by active secretion into colostrum or production of SP in the mammary gland. The presence of these factors in porcine colostrum may be important for the maturation of the gastrointestinal system in neonatal piglets. However, further research is necessary in order to elucidate the role of these factors in porcine colostrum.

Role of VPAC1 and VPAC2 in VIP mediated inhibition of rat pulmonary artery and aortic smooth muscle cell proliferation

Recent studies have suggested the potential use of vasoactive intestinal peptide (VIP) in the treatment of pulmonary arterial hypertension (PAH). An understanding of the mechanism of action of VIP is important for the development of new therapies for PAH. The biological effects of VIP are mediated by two type II guanine nucleotide binding protein (G-protein)-coupled receptors VIP/PACAP (pituitary adenylate cyclase activating peptide) receptor type1 (VPAC1) and VIP/PACAP receptor type 2 (VPAC2). In the present study, the distribution and role of these receptors were investigated and compared in cultured smooth muscle cells from rat aorta and pulmonary artery, as well as in fixed tissue sections of the aorta and pulmonary artery. Western blot analysis, RT-PCR and immunohistochemistry showed the expression of both VIP receptors in tissue sections of the aorta and pulmonary artery as well as in cultured smooth muscle cells from these vessels. The application of a specific antagonist of VPAC1 resulted in a small release from VIP induced inhibition of cell proliferation. In contrast (VIP 6–28; 300 nM) which is an antagonist against both receptors resulted in a significant restoration of proliferation. The expression of cAMP was reduced in the presence of VIP 6–28 and slightly decreased by VPAC1 antagonist. These findings suggest a dual role for VPAC1 and VPAC2 receptors in mediating the antiproliferative effects of VIP with VPAC2 appearing to play a more dominant role.

Vasoactive intestinal peptide and inflammatory cytokines enhance vascular endothelial growth factor production from epidermal keratinocytes

Overexpression of vascular endothelial growth factor (VEGF) in epidermal lesions of psoriasis is well documented; however, its underlying mechanisms are largely unknown. We have recently demonstrated that vasoactive intestinal peptide (VIP) induces the production of cytokines such as interleukin-6 and stem cell factor from keratinocytes, thereby contributing to the development of inflammatory dermatoses such as psoriasis. In this study, we attempted to determine whether VIP could increase the production of VEGF in human keratinocytes. We examined the expression of VEGF using reverse transcription-polymerase chain reaction, immunocytochemistry, enzyme-linked immunosorbent assay and immunoblotting in normal human epidermal keratinocytes and human epidermal keratinocyte cell line DJM-1 cultured in the absence or presence of VIP and/or inflammatory cytokines. We demonstrate that human keratinocytes produced VEGF in a steady state at both mRNA and protein levels. VIP significantly upregulated the production of VEGF in keratinocytes in a dose- and time-dependent manner. The VIP-mediated production of VEGF was further enhanced by inflammatory cytokines such as interferon-gamma, tumour necrosis factor-alpha and interleukin-4, with maximum enhancement being observed with the combination of VIP and interferon-gamma. VIP and other cytokines from nerve endings, mast cells and local inflammatory cells are capable of enhancing VEGF production from epidermal keratinocytes, which may underlie excessive angiogenesis and vasodilation in skin lesions of psoriasis.

Presence of pituitary adenylate cyclase activating polypeptide-38 in human plasma and milk

Pituitary adenylate cyclase activating polypeptide (PACAP) is a pleiotropic and multifunctional neuropeptide widely distributed throughout the body. It is involved in the regulation of various physiological and pathophysiological processes, such as reproduction, thermoregulation, motor activity, brain development, neuronal survival, inflammation and pain. Since little is known about its distribution in humans, our aim was to examine PACAP-38 in human plasma. Furthermore, based on the presence of vasoactive intestinal peptide, structurally the closest to PACAP, in milk and PACAP and its receptors in the mammary gland, our aim was to study PACAP-38 in human milk. The presence of PACAP-38 was determined by mass spectrometry in plasma samples from healthy male and female volunteers (age: 20-40), as well as in plasma and milk samples from lactating women (age: 20-35). PACAP concentration was measured with a specific and sensitive RIA. Our results revealed that PACAP-38 is present in human plasma, its concentration is relatively stable in healthy volunteers and it is not significantly altered by gender, age, food intake or hormonal cycle in females. However, PACAP-38 plasma levels significantly increased in lactating women having 1-6 month-old babies. Moreover, this study is the first which provides evidence for the presence of PACAP-38 in the human milk with levels 5-20-fold greater in the milk whey than in the respective plasma samples. We found PACAP-38 in human plasma and its increase during the first 6 months of the lactation period. A prominent, nearly 10-fold higher concentration of this peptide was detected in human milk. Based on the literature, several important actions of milk-derived PACAP-38 can be suggested such as mammary gland proliferation, nutrient transfer as well as regulation of growth/differentiation of certain tissues of the neonates. The novelty of the present descriptive data provides a basis for further investigations on the mechanism of PACAP-38 secretion in human milk and its functional significance.

Therapeutic Treatment of Experimental Colitis With Regulatory Dendritic Cells Generated With Vasoactive Intestinal Peptide

Crohn's disease is a chronic debilitating disease characterized by severe T helper cell (Th)1-driven inflammation of the colon partially caused by a loss of immune tolerance against mucosal antigens. The use of regulatory dendritic cells (DCs) with the capacity to induce regulatory T cells has been proposed recently for the treatment of Crohn's disease in a strategy to restore immune tolerance. Vasoactive intestinal peptide is an immunomodulatory neuropeptide that induces regulatory DCs. The aim of this study was to investigate the therapeutic effect of vasoactive intestinal peptide-induced regulatory DCs (DC(VIP)) in a murine model of colitis. We examined the therapeutic action of DC(VIP) in the colitis induced by intracolonic administration of trinitrobenzene sulfonic acid, evaluating diverse clinical signs of the disease including weight loss, diarrhea, colitis, and histopathology. We also investigated the mechanisms involved in the potential therapeutic effect of DC(VIP), such as inflammatory cytokines and chemokines, Th1-type response, and the generation of regulatory T cells. DC(VIP) injection significantly ameliorated the clinical and histopathologic severity of colitis, abrogating body weight loss, diarrhea, and inflammation, and increasing survival. The therapeutic effect was associated with down-regulation of both inflammatory and Th1-driven autoimmune response, by regulating a wide spectrum of inflammatory mediators directly through activated macrophages, and by generating interleukin-10-secreting regulatory T cells with suppressive capacity on autoreactive T cells. The possibility to generate/expand ex vivo regulatory DC(VIP) opens new therapeutic perspectives for the treatment of Crohn's disease in human beings, and may minimize the dependence on nonspecific immunosuppressive drugs used currently for autoimmune disorders.

Vasoactive intestinal peptide enhances wound healing and proliferation of human bronchial epithelial cells

In the present study, we investigated the effects of vasoactive intestinal peptide (VIP) on wound healing of bronchial epithelium. Wound healing of the mechanical damaged human bronchial epithelial cells (HBEC) was observed in the absence or presence of VIP. Effects of VIP on chemotactic migration, cell proliferation of HBEC were also tested. HBEC chemotaxis was assessed by the blind well chamber technique, the cell cycle was determined by flow cytometry, and cell proliferation was determined by measuring the expression of proliferating cell nuclear antigen Ki67. Effects of VIP on epithelial E-cadherins protein and mRNA were also measured by immunohistochemistry and RT-PCR. The results showed that VIP accelerated the recovery of wound area of HBEC. VIP increased the migration and proliferation of HBEC, and these effects were blocked by a VPAC1 receptor antagonist. VIP also increased the expression of E-cadherin mRNA and protein in HBEC, suggesting that protective effects of VIP on wound healing may be related to its ability to increase the expression of E-cadherin. In conclusion, VIP has protective effects against human bronchial epithelial cell damage, and the beneficial effects of VIP might be mediated, at least in part, by VPAC1, and associated with increased expression of E-cadherin.

Vasoactive intestinal peptide generates human tolerogenic dendritic cells that induce CD4 and CD8 regulatory T cells

Induction of antigen-specific tolerance is critical for autoimmunity prevention and immune tolerance maintenance. In addition to their classical role as sentinels of the immune response, dendritic cells (DCs) play important roles in maintaining peripheral tolerance through the induction/activation of regulatory T (T(reg)) cells. The possibility of generating tolerogenic DCs opens new therapeutic perspectives in autoimmune/inflammatory diseases. Characterizing endogenous factors that contribute to the development of tolerogenic DCs is highly relevant. We here report that the immunosuppressive neuropeptide vasoactive intestinal peptide (VIP) induces the generation of human tolerogenic DCs with the capacity to generate CD4 and CD8 T(reg) cells from their respective naive subsets. The presence of VIP during the early stages of DC differentiation from blood monocytes generates a population of IL-10-producing DCs unable to fully mature after the effects of inflammatory stimuli. CD4 T(reg) cells generated with VIP-differentiated DCs resemble the previously described Tr1 cells in terms of phenotype and cytokine profile. CD8 T(reg) cells generated with tolerogenic VIP DCs have increased numbers of IL-10-producing CD8(+)CD28(-)-CTLA4(+) T cells. CD4 and CD8 T(reg) cells primarily suppress antigen-specific T(H)1-mediated responses. Therefore, the possibility of generating or expanding ex vivo tolerogenic DC(VIPs) opens new therapeutic perspectives for treating autoimmune diseases and graft-versus-host disease after allogeneic transplantation in humans.

VIP Immunoreactivity in Human Aqueous Humor

Purpose: To demonstrate the presence of vasoactive intestinal peptide (VIP)-immunoreactive molecule in the human aqueous humor collected from eyes undergoing either cataract or glaucoma surgeries and to identify the masses of molecules responsible for the VIP immunoreactivity. Methods: Aqueous humor specimens were collected by parencentesis from nine cataract patients and also from eight patients undergoing glaucoma surgery following the creation of the limbal based conjunctival flap, partial dissection of the scleral flap, and application of mitomycin-C. The aqueous humor specimens were analyzed by radioimmunoassay to determine the level of VIP immunoreactivity. Specimens from 10 other cataract patients were pooled and analyzed for VIP immunoreactivity by Western blot analysis.Results: Levels of VIP immunoreactivity in aqueous humor of cataract and glaucoma patients were significantly different and were 610 ± 160 and 260 ± 64 pg VIP/ml, respectively (p = 0.03), while there was no correlation between the donor age and the level of VIP immunoreactivity. VIP immunoreactivity was detected as a single molecule with a molecular weight of 9000. Conclusions: The disease status and the treatments of the eye that led to surgery and procedures applied to the eye immediately before aqueous humor collection, but not the age of the patients, affected the level of VIP immunoreactivity in the aqueous humor. The relationship between the 9000 Da VIP-immunoreactive molecule and the authentic (3326 Da) VIP remains to be studied.

Effects of vasoactive intestinal peptide and galanin on survival of cultured porcine myenteric neurons

Enteric neuronal plasticity is probably fundamental in order to withstand injury or changes in intestinal activity. The role of the neuropeptides in neuroprotection is still enigmatic. The expression of galanin and vasoactive intestinal peptide (VIP) and the effects of the two peptides on survival of small intestinal porcine myenteric neurons cultured for 6 days were studied. Immunocytochemistry and cell counting were used to evaluate the numbers of surviving neurons and their expression of galanin and VIP. To reflect the in vivo situation, cryostat sections of porcine mid-jejunum were used. A concentration-dependent and marked increase in neuronal survival was noted when neurons were grown in the presence of VIP (10 −8–10 −6 M), whereas addition of galanin (10 −8–10 −6 M) slightly decreased neuronal survival. A dramatic increase in the proportions of myenteric neurons containing VIP or galanin immunoreactivity occurred during culturing. The presence of VIP further increased the number of galanin-expressing neurons. A majority of the galanin-immunoreactive neurons lacked VIP, while all VIP-immunoreactive neurons contained galanin. In conclusion, culturing porcine myenteric neurons in the presence of VIP increases, while the presence of galanin reduces, survival. Culturing significantly increased the proportion of neurons expressing VIP and/or galanin; the presence of VIP further increased the number of galanin-expressing neurons.

Prevention of Experimental Autoimmune Uveoretinitis by Vasoactive IntestinalPeptide

Vasoactive intestinal peptide (VIP), a neuropeptide that is known to be present in lymphoid tissue microenvironments, shows prominent anti-inflammatory actions. To examine the potential effect of VIP on the development of experimental autoimmune uveoretinitis (EAU). We immunized C57BL/6 mice with human interphotoreceptor retinoid-binding protein peptide 1-20 (h-IRBP peptide). Vasoactive intestinal peptide was administered intraperitoneally on alternate days until day 21 after immunization (entire group). In some cases, VIP was injected at different time points after the induction of immunity with h-IRBP peptide (efferent group). In each experiment, a control group of mice was injected with phosphate-buffered saline instead of VIP. Development of EAU was evaluated by means of histological examination on day 21 after immunization. Furthermore, we determined whether intravenous injection of peritoneal exudate cells cultured with VIP overnight in vitro abrogated EAU. We analyzed delayed hypersensitivity for h-IRBP peptide and the occurrence and severity of EAU using evaluation of histopathological sections for inflammatory ocular disease. Treatment with VIP suppressed the expression of delayed hypersensitivity responses to h-IRBP peptide significantly (positive control vs entire group, P =.02; positive control vs efferent group, P<.001). Mice treated with VIP (n = 10) showed a lower occurrence (40%) and decreased severity of EAU (entire group mean score, 0.3; median score, 0) compared with untreated mice (occurrence, 80%; mean score, 0.85; median score, 0.75), as assessed by histopathological analyses (P =.049). Suppressive effects of VIP on EAU were also observed, even when VIP was administered on days 8 through 20 after immunization (efferent group [n = 9] occurrence, 11%; mean score, 0.1; median score, 0) (P =.003). Moreover, expression of EAU was significantly suppressed when the animals were pretreated with peritoneal exudate cells pulsed with h-IRBP in the presence of VIP (control mean score, 1.2; median score, 1.0; occurrence, 80% [n = 10]) compared with the VIP-treatment group (mean score, 0.3; median score, 0; occurrence, 30% [n = 10]) (P =.004). In addition, VIP-treated peritoneal exudate cells generated regulator T cells in the spleens of recipient mice that were able to interfere with the development of EAU (control group mean score, 0.5; median score, 0.5; occurrence, 63% [n = 8]) compared with the VIP-treatment group (mean score, 0.08; median score, 0; occurrence, 17% [n = 6]) (P =.08). Treatment with VIP is a highly effective therapy to suppress EAU. As a result of its efficacy in preventing EAU, VIP might be considered as a novel therapeutic modality for human uveitis.

Vasoactive intestinal peptide and pituitary adenylate cyclase-activating polypeptide promote in vivo generation of memory Th2 cells.

Functionally active effector T cells are generated through clonal expansion. Most effector T cells are later eliminated, whereas a small number survive and differentiate into memory T cells. The mechanisms by which some effector T cells escape apoptosis and become memory T cells are not understood. Neuropeptides such as the vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP) inhibit antigen-induced apoptosis of CD4 T cells. By using an in vivo long-term experimental model, in which CD4 T cells from TRC-transgenic mice were transferred into hosts, we demonstrate that VIP and PACAP induce the survival and/or generation of antigen-specific CD4 T cells with a memory Th2 phenotype. This was confirmed by the fact that transgenic CD4 T cells were recovered only from mice that received Th2, but not Th1 effector cells, in the presence of VIP or PACAP. In vitro, VIP/PACAP support the survival of Th2, but not Th1, cell lines through an inhibition of antigen-induced apoptosis. The role of neuropeptides in the biased development of Th2 memory cells is particularly relevant in view of the immune deviation existing in immune-privileged sites such as the brain and eye, where Th2, but not Th1, responses occur in nonpathological conditions.

Pituitary adenylate cyclase-activating polypeptide modulates plasminogen activator expression in rat granulosa cell.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a bioactive peptide isolated from ovine hypothalamus. It has been demonstrated to be transiently expressed in preovulatory follicles and to positively affect several parameters correlated with the ovulatory process. The aim of the present study was to investigate whether PACAP influences the plasminogen/plasmin system in rat ovary. Plasminogen activators (PAs) are serine proteases, modulated by gonadotropins and several peptides in preovulatory follicles, that appear to be involved in ovulation. Granulosa cells obtained from immature eCG-treated rats were cultured for 24 h in the presence of increasing concentrations of PACAP and vasoactive intestinal peptide (VIP). A significant, dose-dependent increase in tissue-type PA (tPA) activity and decrease in urokinase-type (uPA) PA activity were observed in PACAP-treated cells. These effects were exerted at the mRNA level. The use of cycloheximide, a protein synthesis inhibitor, suggested that PACAP requires an intermediary protein to decrease uPA-mRNA, but not to induce tPA-mRNA. However, no significant modulation of PAs was observed in the presence of VIP. When granulosa cells were stimulated within the intact follicle (i.e., maintaining the three-dimensional structure and in the presence of the theca cell layers), both PACAP and VIP dose-dependently stimulated tPA. These data suggest that, in addition to the PACAP type I receptor present on granulosa cells, different subtypes of PACAP receptors are present in the different ovarian compartments.

Vasoactive intestinal peptide (VIP) stimulates rat prostatic epithelial cell proliferation.

Androgens play a major role in supporting normal growth and functional maintenance in the prostate. However, this gland contains an array of neuroendocrine peptides that can play a regulatory role in its physiopathology. Among these peptides, one of the best studied is vasoactive intestinal peptide (VIP), which is abundant in autonomic nerves surrounding both human and rat prostatic acini. This neuropeptide may act through interaction with two types of high-affinity receptors, named VPAC(1) and VPAC(2) receptors. Another regulatory peptide, the pituitary adenylate cyclase-activating peptide (PACAP), interacts with these receptors with the same affinity as VIP, but binds with higher affinity to PAC(1) receptors. Human prostate tumors and rat prostate show a major presence of VPAC(1) receptors, whereas various findings suggest a role for VIP in prostatic development. Here we studied the effects of VIP on the proliferation of rat prostatic epithelial cells in culture. We studied the [(3)H]-thymidine uptake by rat prostatic epithelial cells in culture, characterized previously by using biomarkers such as cytokeratin and vimentin. In these cells we tested the effect of VIP and PACAP-27 on two different signaling pathways, the cyclic AMP (cAMP) and the inositol phosphate (IPs). The rat prostatic cells in culture were cytokeratin (5,6,8) and vimentin positive, indicating that the culture was predominantly epithelial. The proliferation curves showed that the cells followed different states of growth: a quiescent, an exponential proliferative, and a steady state. Cyclic AMP production, but not inositol phosphate production, was increased in the presence of VIP and PACAP-27, which suggests the expression of VPAC(1) and/or VPAC(2) receptors primarily. VIP significantly increased prostatic cell proliferation in a bimodal manner, as shown for dibutyryl cyclic AMP (dbcAMP), which suggests that the effect of VIP upon prostatic proliferation is cAMP-dependent. Here, we demonstrate that VIP increased [(3)H]thymidine uptake by rat prostatic epithelial cells in culture, conceivably by the activation of the adenylate cyclase.