Gene Expression Data Mining Reveals the Involvement of GPR55 and Its Endogenous Ligands in Immune Response, Cancer, and Differentiation.

G protein-coupled receptor 55 (GPR55) is a recently deorphanized lipid- and peptide-sensing receptor. Its lipidic endogenous agonists belong to lysoglycerophospholipids, with lysophosphatidylinositol (LPI) being the most studied. Peptide agonists derive from fragmentation of pituitary adenylate cyclase-activating polypeptide (PACAP). Although GPR55 and its ligands were implicated in several physiological and pathological conditions, their biological function remains unclear. Thus, the aim of the study was to conduct a large-scale re-analysis of publicly available gene expression datasets to identify physiological and pathological conditions affecting the expression of GPR55 and the production of its ligands. The study revealed that regulation of GPR55 occurs predominantly in the context of immune activation pointing towards the role of the receptor in response to pathogens and in immune cell lineage determination. Additionally, it was revealed that there is almost no overlap between the experimental conditions affecting the expression of GPR55 and those modulating agonist production. The capacity to synthesize LPI was enhanced in various types of tumors, indicating that cancer cells can hijack the motility-related activity of GPR55 to increase aggressiveness. Conditions favoring accumulation of PACAP-derived peptides were different than those for LPI and were mainly related to differentiation. This indicates a different function of the two agonist classes and possibly the existence of a signaling bias.

PACAP modulates the transcription of TLR-1/TLR-5/MyD88 pathway genes and boosts antimicrobial defenses in Clarias gariepinus.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a multifunctional neuropeptide that belongs to the secretin/glucagon/GHRH/VIP superfamily. Some of these molecules have antimicrobial activity and they are capable of stimulating the immune system. The present work studied the antibacterial and immunostimulatory activity of PACAP-38 from African catfish Clarias gariepinus against the Gram-negative bacterium Pseudomonas aeruginosa in an in vivo test. PACAP-38 improved antimicrobial activity of skin mucus molecules against P. aeruginosa. The peptide modulates the gene expression profile of TLR-1, TLR-5, MyD88, IL-1ß, TNF-ɑ, IL-8, pardaxin, hepcidin and G/C-type lysozymes in skin, spleen and head kidney. The influenced exerted depended on the time after infection and tissue analyzed. This study provides the first evidence of a link between PACAP and antimicrobial peptides hepcidin and pardaxin. Our results suggest further use of PACAP as antimicrobial agent that could potentially be used to control disease in aquaculture.

Infusion of Pituitary Adenylate Cyclase-Activating Polypeptide-38 in Patients with Rosacea Induces Flushing and Facial Edema that Can Be Attenuated by Sumatriptan.

BACKGROUND: The pathogenesis of rosacea is incompletely understood. Signaling neuropeptides, including PACAP, a regulator of vasodilation and edema, are upregulated in rosacea skin. Here, we evaluated PACAP38-induced rosacea features and examined whether a 5-HT1B/1D receptor agonist could reduce these features. METHODS: A total of 35 patients with erythematotelangiectatic rosacea received an intravenous infusion of 10 pmol/kg/minute of PACAP38 followed by an intravenous infusion of 4 mg sumatriptan or placebo (saline) on two study days in a double-blind, randomized, placebo-controlled, and cross-over trial. RESULTS: PACAP38 increased facial skin blood flow by 90%, dilated the superficial temporal artery by 56%, and induced prolonged flushing and facial edema. Compared with placebo, sumatriptan reduced PACAP38-induced facial skin blood flow for 50 minutes (P = 0.023), constricted the superficial temporal artery for 80 minutes (P = 0.010), and reduced duration of flushing (P = 0.001) and facial edema (P < 0.001). CONCLUSIONS: We established a clinical experimental model of rosacea features and showed that sumatriptan was able to attenuate PACAP38-induced rosacea flushing and edema. Findings support a key role of PACAP38 in rosacea flushing pathogenesis. It remains unknown whether PACAP38 inhibition can improve rosacea. TRIAL REGISTER: The trial was registered at ClinicalTrials.govNCT03878784 in March 2019.

Pituitary adenylate cyclase-activating polypeptide promotes cutaneous dendritic cell functions in contact hypersensitivity.

BACKGROUND: Sensory nerves regulate cutaneous local inflammation indirectly through induction of pruritus and directly by acting on local immune cells. The underlying mechanisms for how sensory nerves influence cutaneous acquired immune responses remain to be clarified. OBJECTIVE: This study aimed to explore the effect of peripheral nerves on cutaneous immune cells in cutaneous acquired immune responses. METHODS: We analyzed contact hypersensitivity (CHS) responses as a murine model of delayed-type hypersensitivity in absence or presence of resiniferatoxin-induced sensory nerve denervation. We conducted ear thickness measurements, flow cytometric analyses, and mRNA expression analyses in CHS. RESULTS: CHS responses were attenuated in mice that were denervated during the sensitization phase of CHS. By screening neuropeptides, we found that pituitary adenylate cyclase-activating polypeptide (PACAP) mRNA expression was decreased in the dorsal root ganglia after denervation. Administration of PACAP restored attenuated CHS response in resiniferatoxin-treated mice, and pharmacological inhibition of PACAP suppressed CHS. Flow cytometric analysis of skin-draining lymph nodes showed that cutaneous dendritic cell migration and maturation were reduced in both denervated mice and PACAP antagonist-treated mice. The expression of chemokine receptors CCR7 and CXCR4 of dendritic cell s was enhanced by addition of PACAP in vitro. CONCLUSION: These findings indicate that a neuropeptide PACAP promotes the development of CHS responses by inducing cutaneous dendritic cell functions during the sensitization phase.

Targeting pituitary adenylate cyclase-activating polypeptide (PACAP) with monoclonal antibodies in migraine prevention: a brief review.

INTRODUCTION: Interest is growing in the role of pituitary adenylate cyclase-activating polypeptide (PACAP) and its specific PAC1 receptor in migraine and in their antagonism as a strategy for migraine prevention. AREAS COVERED: We discuss and critically evaluate (i) the evidence of the role of PACAP in migraine pathophysiology and (ii) the first clinical trials in migraine prophylaxis with monoclonal antibodies AMG 301 and ALD1910 which act against PAC1 and PACAP38 respectively. We examined PubMed, Scopus, and ClinicalTrials.gov electronic databases to examine the relevant material. EXPERT OPINION: There is much proof of the ability of PACAP to cause migraine, but there is limited evidence that blocking PACAP or PAC1 receptor can prevent migraine. However, the potential of anti-PACAP antibodies in migraine prophylaxis is high. Theoretically, if these antibodies block the activation of the trigeminovascular system, they will prevent the onset of migraine attacks. There are still knowledge gaps in the role of PACAP in migraine and the risk/benefit ratio of anti-PACAP antibodies must be carefully studied.

First in vivo evidence of pituitary adenylate cyclase-activating polypeptide antiviral activity in teleost.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a multifunctional neuropeptide belonging to the glucagon/secretin superfamily. In teleost fish, PACAP has been demonstrated to have an immunomodulatory role. Although previous studies have shown that viral/bacterial infections can influence the transcription of PACAP splicing variants and associated receptors in salmonids, the antiviral activity of PACAP has never been studied in teleost. Thus, in the present work, we investigated in vitro the influence of synthetic Clarias gariepinus PACAP-38 on the transcription of genes related to viral immunity using the rainbow trout monocyte/macrophage-like cell line RTS11 as a model. Positive transcriptional modulation of interferon gamma (IFNγ), interferon alpha (FNα1,2), interleukin 8 (IL-8), Mx and Toll-like receptor 3 (TLR3) genes was found in a dose and time dependent manner. We also explored how a pre-treatment with PACAP could enhance antiviral immune response using poly (I:C) as viral mimic. Interferons and IL-8 transcription levels were enhanced when PACAP was added 24 h previous to poly (I:C) exposure. With these evidences, we tested in vivo how PACAP administration by immersion bath affected the survival of rainbow trout fry to a challenge with viral hemorrhagic septicemia virus (VHSV). After challenge, PACAP-treated fish had increased survival compared to non-treated/challenge fish. Furthermore, PACAP was able to decrease the viral load in spleen/kidney and stimulate the transcription of IFNs and Mx when compared to untreated infected fish. Altogether, the results of this work provide valuable insights regarding the role of teleost PACAP in antiviral immunity and point to a potential application of this peptide to reduce the impact of viral infections in aquaculture.

PACAP38 improves airway epithelial barrier destruction induced by house dust mites allergen.

PURPOSE: This study aimed to investigate the mechanism of PACAP38 on house dust mite (HDM)-induced asthmatic airway epithelial barrier destruction. METHODS: The HDM-induced asthma mice model and 16HBE cell model was established respectively. The enzyme linked immunosorbent assay (ELSIA), cell count and immunohistochemical assay were performed on mice in control group, HDM group and PACAP38 + HDM group.The cAMP/PKA activity, p-CREB and total CREB expression, TEER and the FITC-DX were investigated on cells in control-16HBE group, HDM-16HBE group and PACAP38 + HDM-16HBE group. RESULTS: The levels of IL-4 and IL-5 in the HDM group were significantly higher than those in the control group (P < 0.05), while the above indexes in the PACAP38 + HDM group were lower than those in the HDM group (P < 0.05). E-cadherin, ß-catenin, ZO-1 and occludin in the control group were highly immunoreactive in airway epithelial cells, whereas connexin staining was attenuated after HDM induction. The TEER level, cAMP levels and PKA activity were decreased, while FITC-DX transmittance was increased in HDM-16HBE group (P < 0.05) compared with the control-16HBE group. CONCLUSION: PACAP38 could reduce the airway inflammation, weaken the AJC protein heterotopia and activate cAMP/PKA signaling pathway in HDM-induced asthma, which indicate that PACAP38 may be an important contributor in HDM-induced asthma.

Immunomodulatory Effects of the Neuropeptide Pituitary Adenylate Cyclase-Activating Polypeptide in Acute Toxoplasmosis.

Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP) is an endogenous neuropeptide with distinct functions including the regulation of inflammatory processes. PACAP is able to modify the immune response by directly regulating macrophages and monocytes inhibiting the production of inflammatory cytokines, chemokines and free radicals. Here, we analyzed the effect of exogenous PACAP on peripheral immune cell subsets upon acute infection with the parasite Toxoplasma gondii (T. gondii). PACAP administration was followed by diminished innate immune cell recruitment to the peritoneal cavity of T. gondii-infected mice. PACAP did not directly interfere with parasite replication, instead, indirectly reduced parasite burden in mononuclear cell populations by enhancing their phagocytic capacity. Although proinflammatory cytokine levels were attenuated in the periphery upon PACAP treatment, interleukin (IL)-10 and Transforming growth factor beta (TGF-ß) remained stable. While PACAP modulated VPAC1 and VPAC2 receptors in immune cells upon binding, it also increased their expression of brain-derived neurotrophic factor (BDNF). In addition, the expression of p75 neurotrophin receptor (p75NTR) on Ly6Chi inflammatory monocytes was diminished upon PACAP administration. Our findings highlight the immunomodulatory effect of PACAP on peripheral immune cell subsets during acute Toxoplasmosis, providing new insights about host-pathogen interaction and the effects of neuropeptides during inflammation.

PACAP Is Lethal to Flavobacterium psychrophilum Through Either Direct Membrane Permeabilization or Indirectly, by Priming the Immune Response in Rainbow Trout Macrophages.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a multifunctional neuropeptide that is widely distributed in mammals and is capable of performing roles as a neurotransmitter, neuromodulator, and vasodilator. This polypeptide belongs to the glucagon/secretin superfamily, of which some members have been shown to act as antimicrobial peptides in both mammalian and aquatic organisms. In teleosts, PACAP has been demonstrated to have direct antimicrobial activity against several aquatic pathogens, yet this phenomenon has never been studied throughout a live bacterial challenge. The present study focuses on the influence of synthetic Clarias gariepinus 38 amino acid PACAP on the rainbow trout monocyte/macrophage-like cell line, RTS11, when exposed to the coldwater bacterial pathogen Flavobacterium psychrophilum. PACAP was shown to have direct antimicrobial activity on F. psychrophilum when grown in both cytophaga broth and cell culture media (L-15). Further, the ability of teleostean PACAP to permeabilize the membrane of an aquatic pathogen, F. psychrophilum, was demonstrated for the first time. The viability of RTS11 when exposed to PACAP was also observed using a trypan blue exclusion assay to determine optimal experimental doses of the antimicrobial peptide. This displayed that only concentrations higher than 0.1 µM negatively impacted RTS11 survival. Interestingly, when RTS11 was pre-treated with PACAP for 24 h before experiencing infection with live F. psychrophilum, growth of the pathogen was severely inhibited in a dose-dependent manner when compared to cells receiving no pre-treatment with the polypeptide. Relative expression of pro-inflammatory cytokines (IL-1ß, TNFα, and IL-6) and PACAP receptors (VPAC1 and PAC1) was also analyzed in RTS11 following PACAP exposure alone and in conjunction with live F. psychrophilum challenge. These qRT-PCR findings revealed that PACAP may have a synergistic effect on RTS11 immune function. The results of this study provide evidence that PACAP has immunostimulatory activity on rainbow trout immune cells as well as antimicrobial activity against aquatic bacterial pathogens such as F. psychrophilum. As there are numerous pathogens that plague the aquaculture industry, PACAP may stimulate the teleost immune system while also providing an efficacious alternative to antibiotic use.

Pituitary Adenylate Cyclase-Activating Polypeptide-A Neuropeptide as Novel Treatment Option for Subacute Ileitis in Mice Harboring a Human Gut Microbiota.

The neuropeptide Pituitary adenylate cyclase-activating polypeptide (PACAP) is well-known for its important functions in immunity and inflammation. Data regarding anti-inflammatory properties of PACAP in the intestinal tract are limited, however. In our present preclinical intervention study we addressed whether PACAP treatment could alleviate experimental subacute ileitis mimicking human gut microbiota conditions. Therefore, secondary abioitic mice were subjected to human fecal microbiota transplantation (FMT) and perorally infected with low-dose Toxoplasma gondii to induce subacute ileitis on day 0. From day 3 until day 8 post-infection, mice were either treated with synthetic PACAP38 or placebo. At day 9 post-infection, placebo, but not PACAP treated mice exhibited overt macroscopic sequelae of intestinal immunopathology. PACAP treatment further resulted in less distinct apoptotic responses in ileal and colonic epithelia that were accompanied by lower T cell numbers in the mucosa and lamina propria and less secretion of pro-inflammatory cytokines in intestinal ex vivo biopsies. Notably, ileitis-associated gut microbiota shifts were less distinct in PACAP as compared to placebo treated mice. Inflammation-ameliorating effects of PACAP were not restricted to the intestines, but could also be observed in extra-intestinal including systemic compartments as indicated by lower apoptotic cell counts and less pro-inflammatory cytokine secretion in liver and lungs taken from PACAP treated as compared to placebo control mice, which also held true for markedly lower serum TNF and IL-6 concentrations in the former as compared to the latter. Our preclinical intervention study provides strong evidence that synthetic PACAP alleviates subacute ileitis and extra-intestinal including systemic sequelae of T cell-driven immunopathology. These findings further support PACAP as a novel treatment option for intestinal inflammation including inflammatory bowel diseases (IBD).

Pharmacologic Characterization of ALD1910, a Potent Humanized Monoclonal Antibody against the Pituitary Adenylate Cyclase-Activating Peptide.

Migraine is a debilitating disease that affects almost 15% of the population worldwide and is the first cause of disability in people under 50 years of age, yet its etiology and pathophysiology remain incompletely understood. Recently, small molecules and therapeutic antibodies that block the calcitonin gene-related peptide (CGRP) signaling pathway have reduced migraine occurrence and aborted acute attacks of migraine in clinical trials and provided prevention in patients with episodic and chronic migraine. Heterogeneity is present within each diagnosis and patient's response to treatment, suggesting migraine as a final common pathway potentially activated by multiple mechanisms, e.g., not all migraine attacks respond to or are prevented by anti-CGRP pharmacological interventions. Consequently, other unique mechanisms central to migraine pathogenesis may present new targets for drug development. Pituitary adenylate cyclase-activating peptide (PACAP) is an attractive novel target for treatment of migraines. We generated a specific, high-affinity, neutralizing monoclonal antibody (ALD1910) with reactivity to both PACAP38 and PACAP27. In vitro, ALD1910 effectively antagonizes PACAP38 signaling through the pituitary adenylate cyclase-activating peptide type I receptor, vasoactive intestinal peptide receptor 1, and vasoactive intestinal peptide receptor 2. ALD1910 recognizes a nonlinear epitope within PACAP and blocks its binding to the cell surface. To test ALD1910 antagonistic properties directed against endogenous PACAP, we developed an umbellulone-induced rat model of neurogenic vasodilation and parasympathetic lacrimation. In vivo, this model demonstrates that the antagonistic activity of ALD1910 is dose-dependent, retaining efficacy at doses as low as 0.3 mg/kg. These results indicate that ALD1910 represents a potential therapeutic antibody to address PACAP-mediated migraine.

Immunomodulatory Roles of PACAP and VIP: Lessons from Knockout Mice.

A bidirectional cross-talk is established between the nervous and immune systems through common mediators including neuropeptides, neurotransmitters, and cytokines. Among these, PACAP and VIP are two highly related neuropeptides widely distributed in the organism with purported immunomodulatory actions. Due to their well-known anti-inflammatory properties, administration of these peptides has proven to be beneficial in models of acute and chronic inflammatory diseases. Nevertheless, the relevance of the endogenous source of these peptides in the modulation of immune responses remains to be elucidated. The development of transgenic mice with specific deletions in the genes coding for these neuropeptides (Vip and Adcyap1) or for their G-protein-coupled receptors VPAC1, VPAC2, and PAC1 (Vipr1, Vipr2, Adcyap1r1) has allowed to address this question, underscoring the complexity of the immunoregulatory properties of PACAP and VIP. The goal of this review is to integrate the existing information on the immune phenotypes of mice deficient for PACAP, VIP, or their receptors, to provide a global view on the roles of these endogenous neuropeptides during immunological health and disease.

Real Time and Label-Free Research on the Detection of Pituitary Adenylate Cyclase-Activating Polypeptide Based on Surface Plasmon Resonance Technique.

BACKGROUND: The self-developed portable surface plasmon resonance (SPR) biosensor was used in the quantitative detection and kinetic study of pituitary adenylate cyclase-activating polypeptide (PACAP), as the existing detection methods were complicated, with a long detection period, high-cost instrument, and sample needing to be labeled. METHODS: After preparing SPR biochip, the direct detection proceeded in an immune reaction detection between the PACAP samples with concentrations of 0.5 mg/L, 1 mg/L, 2 mg/L, 5 mg/L, 8 mg/L, 10 mg/L, and PACAP type 1 receptor (PAC1R). The standard curve of PACAP direct detection was established. According to the 1:1 Langumair model, the immune responses dynamic characteristic parameters of PAC1R with PACAP-38, and their reconstructive PN37R, PK38W were calculated, respectively. RESULTS: The direct detection limit of PACAP could be 0.5 mg/L. The absolute deviation and relative deviation of the detected value and the true value are both low. The magnitude orders of kinetic parameters of immune response between PAC1R and PACAP-38, PK38W, PN37R are basically the same. However, the specific values of the binding rate constant and dissociation rate constant of PK38W and PN37R are slightly larger than that of PACAP-38. CONCLUSIONS: The experimental results show that the SPR biochip detection system can be used for the effective quantitative detection of PACAP and can be used for kinetic study. The developed device could provide a labelfree, simple, quick, and low-cost method for the concentration detection of PACAP in the samples and the immune response study between PACAP and its receptors. It could promote PACAP to play an important role in the pharmaceutical industry, clinical treatment, and other industries.

Migraine, Neurogenic Inflammation, Drug Development - Pharmacochemical Aspects.

BACKGROUND: Migraine is a primary headache disorder. Despite numerous studies conducted with the aim to understand the pathophysiology of migraine, several aspects are still unclear. The trigeminovascular system plays a key role. Neurogenic inflammation is presumed to be an important factor in migraine pathophysiology, mediated by the activation of primary neurons, leading to the release of various pro-inflammatory neuropeptides and neurotransmitters such as Calcitonin Gene-Related Peptide (CGRP), substance P (SP), and vasoactive intestinal peptide (VIP). Nitric oxide (NO), Pituitary adenylate cyclase-activating polypeptide (PACAP) and Glutamate (Glu) also play an important role in the modulation of inflammatory mechanisms. OBJECTIVE: To review the literature focusing on novel therapeutic targets in migraine, related to neurogenic inflammation. METHOD: A systematic literature search in the database of PUBMED was conducted regarding therapeutic strategies in migraine, focusing on substances and cytokines released during neurogenic inflammation, published until January 2017. RESULTS: Ongoing phase III clinical studies with monoclonal antibodies against CGRP and CGRP receptors offer promising novel aspects for migraine treatment. Preclinical and clinical studies targeting SP and nitric oxide synthase (NOS) were all terminated with no significant results compared to placebo. New promising therapeutic goal could be PACAP and its receptor (PAC1), and kynurenic acid (KYNA) analogues. CONCLUSION: Current migraine treatment offers pain relief only for a small proportion of migraine patients and might not be adequate for patients with cardiovascular comorbidity due to side effects. Better understanding of migraine pathophysiology might, therefore, lead to novel therapeutic lines both in migraine attack treatment and prophylaxis.

Chemical Coding of Sensory Neurons Supplying the Hip Joint Capsule in the Sheep.

Immunohistochemical properties of nerve fibres supplying the joint capsule were previously described in many mammalian species, but the localization of sensory neurons supplying this structure was studied only in laboratory animals, the rat and rabbit. However, there is no comprehensive data on the chemical coding of sensory neurons projecting to the hip joint capsule (HJC). The aim of this study was to establish immunohistochemical properties of sensory neurons supplying HJC in the sheep. The study was carried out on 10 sheep, weighing about 30-40 kg. The animals were injected with a retrograde neural tracer Fast Blue (FB) into HJC. Sections of the spinal ganglia (SpG) with FB-positive (FB+) neurons were stained using antibodies against calcitonin gene-related peptide (CGRP) substance P (SP), pituitary adenylate cyclase-activating peptide (PACAP), nitric oxide synthase (n-NOS), neuropeptide Y (NPY), vasoactive intestinal polypeptide (VIP), Leu-5-enkephalin (Leu-Enk), galanin (GAL) and vesicular acetylcholine transporter (VACHT). The vast majority of FB+ neurons supplying HJC was found in the ganglia from the 5th lumbar to the 2nd sacral. Immunohistochemistry revealed that most of these neurons were immunoreactive to CGRP or SP (80.7 ± 8.0% or 56.4 ± 4.8%, respectively) and many of them stained for PACAP or GAL (52.9 ± 2.9% or 50.6 ± 19.7%, respectively). Other populations of FB+ neurons were those immunoreactive to n-NOS (37.8 ± 9.7%), NPY (34.6 ± 6.7%), VIP (28.7 ± 4.8%), Leu-Enk (27.1 ± 14.6) and VACHT (16.7 ± 9.6).

Pituitary adenylate cyclase-activating peptide-27 (PACAP-27) is co-stored with galanin, substance P and corticotropin releasing factor (CRF) in intrapancreatic ganglia of the sheep.

Pituitary adenylate cyclase activating polypeptide (PACAP) is a neuropeptide existing in two variant forms (of either 27 or 38 residues), widely present in numerous organs and evoking multiple effects both in the central and peripheral nervous systems. The present study was undertaken to evaluate the distribution pattern of PACAP-27 expression in the ovine pancreas. Using double immunohistochemical stainings co-localizations of PACAP-27 with galanin, SP or CRF were studied in intrapancreatic neurons. In intrapancreatic ganglia, immunoreactivty to PACAP-27 was found in 87.6 ± 5.4% of PGP 9.5-positive intrapancreatic neurons but not in intraganglionic nerve fibres. Numerous PACAP-27-immunoreactive nerve terminals were also observed between pancreatic acini and around small arterioles. No immunoreactivity to PACAP-27 was found in the endocrine pancreas. In 42.9 ± 6.2% of PACAP-27-immunoreactive intrapancreatic neurons the expression of galanin was also found. Statistically lower subpopulation (12.4 ± 4.0%) of intrapancreatic neurons exhibited simultaneously the immunoreactivity to PACAP-27 and SP. The expression of CRF was detected in the relatively smallest group (3.2 ± 1.4%) of PACAP-27-positive intrapancreatic neurons. The present results suggest that in the ovine pancreas PACAP-27 may play an important role as mediator of pancreatic functions. In PACAP-related pancreatic activities, a modulatory role of galanin, SP and to a lower extend of CRF is also likely.

Examination of PACAP38-like immunoreactivity in different milk and infant formula samples.

Pituitary adenylate cyclase activating polypeptide (PACAP) is a neuropeptide with special importance in reproductive and developmental processes. PACAP is found in two bioactive forms: PACAP27 and PACAP38. Recently, we have described that PACAP38 is present in high levels in the milk of human and ruminant animals. Breastfeeding is of utmost importance in proper nutrition of the newborn, but artificial nursing with infant formulas is necessary when breastfeeding is not available. Composition of the breast milk varies during the whole period of nursing and it shows differences at the beginning (foremilk) and the end of an actual suckling (hindmilk). The aim of this study was to investigate PACAP38-like immunoreactivity (PACAP38-LI) in different milk and infant formula samples by radioimmunoassay and to prove the presence of PACAP38 in the infant formula by mass spectrometry. We found similar PACAP38-LI in human mature foremilk and hindmilk samples, in the fresh and pasteurized cow milk and also in formulas. However, we found significantly higher PACAP38-LI in the hypoantigenic formula undergoing extensive hydrolysis compared to the non-hypoantigenic ones. Our results suggest that PACAP38 is relatively stable in the milk and it can withstand the manufacturing processes.

Pituitary adenylate cyclase-activating peptide and vasoactive intestinal polypeptide bias Langerhans cell Ag presentation toward Th17 cells.

Epidermal Langerhans cells (LCs) are dendritic APCs that play an important role in cutaneous immune responses. LCs are associated with epidermal nerves and the neuropeptides vasoactive intestinal polypeptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP) inhibit LC Ag presentation for Th1-type immune responses. Here, we examined whether PACAP or VIP modulates LC Ag presentation for induction of IL-17A-producing CD4(+) T cells. Treatment with VIP or PACAP prior to in vitro LC Ag presentation to CD4(+) T cells enhanced IL-17A, IL-6, and IL-4 production, decreased interferon (IFN)-γ and interleukin (IL)-22 release, and increased RORγt and Gata3 mRNA expression while decreasing T-bet expression. The CD4(+) T-cell population was increased in IL-17A- and IL-4-expressing cells and decreased in IFN-γ-expressing cells. Addition of anti-IL-6 mAb blocked the enhanced IL-17A production seen with LC preexposure to VIP or PACAP. Intradermal administration of VIP or PACAP prior to application of a contact sensitizer at the injection site, followed by harvesting of draining lymph node CD4(+) T cells and stimulation with anti-CD3/anti-CD28 mAbs, enhanced IL-17A and IL-4 production but reduced production of IL-22 and IFN-γ. PACAP and VIP are endogenous mediators that likely regulate immunity and immune-mediated diseases within the skin.