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Published in final edited form as: Peptides. 2010 Oct 13;32(1):131–137. doi: 10.1016/j.peptides.2010.09.022

The absence of VPAC2 leads to aberrant antibody production in Aspergillus fumigatus sensitized and challenged mice

Amali E Samarasinghe a, Scott A Hoselton a, Jane M Schuh a
PMCID: PMC3299058  NIHMSID: NIHMS249459  PMID: 20923692

Abstract

Vasoactive intestinal peptide (VIP) facilitates a “pro-allergy” phenotype when signaling through its G protein-coupled receptor, VPAC2. We have shown that VPAC2 knock-out (KO) mice developed an allergic phenotype marked by eosinophilia and elevated serum IgE. Therefore, we hypothesized that the humoral response to allergen challenge in these mice was TH2 dominant similar to wild-type (WT) C57BL/6 mice. Antibody responses in WT and KO mice were measured after Aspergillus fumigatus conidia inhalation. In contrast to previous reports, basal levels of serum IgG2a and IgA were significantly higher in naïve VPAC2 KO animals. Antibody availability in the serum as well as the bronchoalveolar lavage fluid after fungal challenge was dominated by the pro-inflammatory isotype IgG2a and the mucosal isotype, IgA. IgA localizing cells dominated in the peribronchovascular areas of allergic KO mice while IgE immune complexes were found in WT allergic lungs. This research shows for the first time that VPAC2 has a significant effect on antibody regulation, in the context of allergy.

Keywords: VIP, IgG2a, IgA, Asthma

1. Introduction

Allergic asthma is a debilitating TH2 mediated pulmonary syndrome marked by acute exacerbations of reversible airway constriction and inflammation triggered by the inhalation of a sensitizing agent, which may result in permanent airflow dysfunction due to chronic airway wall remodeling. A distinct neurological component, including the attendant inflammation [35], cough [22], airway hyperresponsiveness [24, 25, 29], and mucus production [26], is associated with asthma symptoms. Indeed, dysfunction of pulmonary nervous responses has been implicated as a causative or an aggravating factor in the immunomodulation of allergic asthma [32, 37]. Neuropeptides, such as vasoactive intestinal peptide (VIP), are supplied to peripheral organs like the lungs by an abundant supply of synapsing nerves [2]. Additionally, macrophages and lymphocytes contribute to the local production of “neuropeptides” in lymphoid and other organs and respond to these mediators in an autocrine/paracrine manner. While the availability of neuropeptides in the lung is indisputable, their role in the communication between the nervous and immune networks is complicated and has not been fully elucidated.

Originally identified in 1970 [33], VIP has emerged as an important neuropeptide for its anti-inflammatory effects on immune cells. Acting through at least two G-protein coupled receptors in the lung, VPAC1 [36] and VPAC2 [28], VIP regulates leukocytes by hindering pro-inflammatory mediator release. VIP augments the TH2 phenotype by: (i) acting directly and indirectly on antigen presenting cells to preferentially recruit TH2 cells, (ii) promoting the generation of TH2 cells, and (iii) enhancing the survival and proliferation of existing TH2 cells [31]. The VIP/VPAC2 axis on CD4+ T cells promotes TH2 cytokine production inhibiting the production of classic TH1 cytokines [38]. The strength and duration of the TH2 immune phenotype is enhanced by the auto-regulatory function of VIP produced by TH2 cells [8]. While VPAC2 transgenic mice exhibit increased IgE, eosinophilia, and decreased delayed-type hypersensititvity (DTH) [38]; VPAC2 KO mice have an inflammatory phenotype with increased DTH and decreased immediate-type hypersensitivity (ITH) responses [15].

VIP is a potent bronchodilator and chemically modified VIP has been actively pursued as a potential therapeutic for use in asthma, yet the physiologic role of VIP and its receptors in the development and maintenance of allergic asthma has not been fully explored. Our recent studies using VPAC2 KO mice showed that the absence of this receptor was not protective in the development of fungal allergy and that IgE was produced in the absence of VPAC2 signaling [34]. This was interesting, since other investigators have reported that ITH is significantly decreased in VPAC2 KO mice [15]. In this study, using an Aspergillus fumigatus induced murine model system of fungal asthma, we show that VPAC2 KO mice have aberrant humoral responses to systemic and local allergen provocation.

2. Materials and Methods

2.1. Model of allergic airways disease

C57BL/6 mice (WT, Jackson Laboratories, Bar Harbor, ME) and VPAC2 KO mice on a C57BL/6 background (heterozygous animals, a kind gift from Dr. Anthony Harmar, University of Edinburgh, UK, were bred to produce homozygous KO) were used for the study. All animals were housed in a specific pathogen-free facility in microisolator cages throughout the study and bedded on paper bedding, which harbors extremely low microbial contamination. As part of the IACUC-approved animal care protocol, sentinel animals that share bedding with the WT and KO animals were routinely tested for viral and bacterial profiles. None were detected. The study described was performed in accordance with IACUC and Institutional Biosafety Committee guidelines of North Dakota State University.

Animals were sensitized as previously described [34]. Briefly, 10 μg of A. fumigatus antigen (Greer Laboratories, Inc., Lenoir, N.C.) adsorbed in 0.2 ml of Alum was injected subcutaneously (0.1 ml) and intraperitoneally (0.1 ml). Two weeks later, mice received the first of three weekly intranasal sensitizations with 20 μg of A. fumigatus antigen in 20 μl of normal saline. Sensitized mice were challenged as previously described [34]. Briefly, one week after the final sensitization, anesthetized mice were subjected to two 10-min, nose-only exposure to live fungal conidia two weeks apart. Naïve animals were neither sensitized nor challenged. Mice were separated into groups of five and euthanized at predetermined time points (days 3, 7, 14, and 28 after the second allergen exposure). Figure 1 provides a schematic representation of the allergen sensitization and challenge protocol.

Figure 1.

Figure 1

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Schematic representation of the Aspergillus fumigatus induced murine model system of allergic asthma. Mice were sensitized against antigen from A. fumigatus over the course of five weeks. Sensitized mice were exposed to un-manipulated conidia via normal inhalation for 10 minutes two weeks apart and analyzed at days 3, 7, 14, and 28 after the second inhalation challenge.

2.2. Sample collection and determination of antibody concentration in sera and BAL fluid by ELISA

Approximately 500 μl of blood was collected by ocular bleed, and a bronchoalveolar lavage (BAL) was performed on each mouse with 1 ml of sterile saline. Left lungs were harvested and fixed in formalin for histological analysis. Blood harvested from each mouse was centrifuged at 15,000 × g for 10 min to obtain sera which was frozen at −20°C until use. BAL contents were centrifuged at 600 × g for 10 min to separate the cell ular component from the fluid. The BAL fluid was stored at −20°C until used.

To assess the affect of pro-inflammatory (TH1-type) skewing on Ab production in the absence of VIP’s “pro-allergy” receptor VPAC2, and to determine baseline levels for allergic sensitization, we measured the concentrations of IgG2a (pro-inflammatory), and IgA (mucosal), and IgE (pro-allergy) in sera and BAL fluid. IgE, IgG2a (BD OptEIA, San Diego, CA), and IgA (Bethyl Labs, Montgomery, TX) were quantified with ELISA per manufacturer’s protocols.

2.3. Immunohistochemical analysis of VIP, IgE, and IgA

Left lungs were inflated with 10% neutral buffered formalin and longitudinal 5-μm sections were cut across the coronal plane and used for immunohistochemical (IHC) analyses. Goat anti-rabbit VIP antibody (Abcam, Cambridge, MA) diluted at 1:20 was used with the anti-rabbit HRP-AEC tissue staining kit from R&D Systems (Minneapolis, MN) for VIP immunostaining. Goat anti-mouse IgA antibody and goat anti-mouse IgE antibody (Southern Biotech, Birmingham, AL) at 1:50 dilution with the anti-goat HRP-AEC tissue staining kit from R&D stained IgA and IgE red against the Gill III hematoxylin (Surgipath, Richmond, IL) blue counterstain.

2.4. Statistical analysis of data

All results are expressed as mean ± S.E.M. WT and KO mice at each time point were compared with Prism GraphPad software (San Diego, CA) using an unpaired Student’s two tailed t-test with Welch’s correction to determine statistical significance at predetermined time points after allergen challenge. p <0.05 was considered statistically significant.

3. Results

Both groups of mice were demonstrated to develop an allergic phenotype against A. fumigatus conidia previously [34]. Based on this evidence, we hypothesized that VPAC2 null mice would have similar humoral responses against allergen challenge compared to WT controls. In contrast to previous reports that demonstrated VPAC2 null mice to have “normal basic immune characteristics” [15], we show that these mice have aberrant antibody responses in naïve and allergic states.

3.1. VIP localization in the naïve and allergic lungs was similar in WT and KO mice

Since VIP can serve as a regulator of leukocyte migration [4, 10, 17], and we have shown differences in VIP mRNA expression after allergen challenge between WT and KO lungs [34], we used immunohistochemistry to identify the cellular localization of VIP in the naïve and allergic WT and KO lungs. Naïve mice of both groups exhibited a similar pattern of VIP localization with the predominant site being the columnar epithelia (Fig 2A & C). While naïve WT columnar epithelia showed homogenous VIP localization (Fig 2A), the naïve VPAC2 KO lungs had prominent VIP localization in the apical ends of the columnar epithelia (Fig 2C). After allergen challenge, there was a reduction in VIP in the columnar epithelia of both groups (Fig 2B & D).

Figure 2.

Figure 2

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Immunohistochemical staining for VIP localization in naïve and day 7 wild-type (WT) and VPAC2 knock-out (KO) lungs. The columnar epithelia and Type II pneumocytes were the predominant areas of VIP localization in both the WT (A) and KO (C) naïve lungs. VIP localization was decreased in these areas after allergen challenge, but equivalent localization occurred in WT (B) and KO (D) day 7 lungs. Scale bars = 100 μm

3.2. VPAC2 KO mice had elevated IgG2a and IgA in naïve and disease states

We have previously demonstrated that VPAC2 KO mice produced IgE equivalent to that of WT animals after allergen challenge and that they sustained elevated levels of IgE in the sera for longer than their WT counterparts [34]. Naïve KO mice demonstrated significant overexpression of IgG2a in serum when compared to WT mice (Fig 3A) which was in stark contrast with a previous report [15]. Serum levels of IgG2a in naïve KO mice were approximately 175× that of naïve WT mice (Fig 3A). A continual increase in IgG2a occurred in the KO mice after allergen challenge, while a reduction was noticed in allergic WT mice (Fig 3A). Serum IgG2a was maintained at a much higher level in the KO at all time points after the final conidia inhalation (Fig 3A).

Figure 3.

Figure 3

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Antibodies quantified in sera of naïve and allergic wild-type (WT) and VPAC2 knockout (KO) mice via ELISA. Naïve KO mice contained more IgG2a (A) and IgA (B) in the sera compared to naïve WT mice. Allergen challenge led to a decreasing trend in serum IgG2a in the WT, while KO mice had increased IgG2a production as a result of fungal inhalation (A). The amount of IgA in the sera did not change in either group during the early time points after allergen challenge (B). However, at day 28, WT IgA availability decreased while KO levels remained elevated (B). All values expressed as the mean ± S.E.M. n=5 mice/group, *p<0.05

The IgA content in the sera in the naïve KO and WT groups were equivalent (Fig 3B). Although serum IgA levels were significantly different between WT and KO after allergen challenge, there was no difference in IgA availability in each group compared to their respective naïve controls (Fig 3B). While WT IgA in the sera decreased by day 28, KO levels remained elevated (Fig 3B).

3.3. VPAC2 KO mice had an aberrant local antibody response to fungal challenge

Since fungal allergen challenge predominantly impacts mucosal immunity in the lung, we assessed the local availability of immunoglobulins in the BAL fluid of the lung. While both groups of mice increased the availability of IgE in the lung in response to fungal challenge, KO mice had significantly more IgE at days 7 and 14 compared to that in WT mice (Fig 4A). Both groups maintained IgE in the BAL fluid at day 28 reiterating their allergic phenotype (Fig 4A). However, IgE was the least abundant of the isotypes in the BAL after allergen challenge (Fig 4).

Figure 4.

Figure 4

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Antibodies quantified in the bronchoalveolar lavage (BAL) fluid of naïve and allergic wild-type (WT) and VPAC2 knock-out (KO) mice. Naïve animals in both groups had minimal antibody production in the lungs (A-C). IgE in the BAL fluid increased after allergen challenge in both groups although KO mice had a marked increase in local IgE production at days 7 and 14 compared to WT controls (A). WT mice had negligible levels of IgG2a in the BAL fluid after allergen challenge while a massive increase in IgG2a was observed in the KO BAL fluid (B). The mucosal antibody, IgA, was increased as a result of fungal inhalation in both groups, although KO had far more IgA after day 3 compared to WT (C). All values expressed as the mean ± S.E.M. n=5 mice/group, *p<0.05

In WT mice, IgG2a was not a major Ab component of the naïve BAL, nor was it induced in the lung after allergen challenge (Fig 4B). After allergen challenge, IgG2a concentration was increased in the BAL fluid of VPAC2 KO animals and remained significantly elevated through the day-28 time point as compared to naïve levels and that produced in allergic WT animals (Fig 4B). IgA was observed at very low levels in naïve WT and KO (Fig 4C). In contrast to the pattern seen in the serum, IgA was induced in the BAL fluid following allergen challenge in both groups (Fig 4C). While WT mice maintained these same level of antibodies at all subsequent time points, KO mice had significantly more of IgA at later time points (Fig 4C). The trend in IgA in the BAL followed that of IgE for both groups (Fig 4).

3.4. Immune complexes with IgE were prominent in WT allergic lungs while allergic VPAC2 KO lungs had more IgA localizing cells in the peribronchovascular areas

IgE is important in the allergic response because of its ability to cross link antigen to elicit mast cell degranulation and activation of the TH2 immune cascade. After allergen challenge, we observed that serum and BAL IgE was significantly increased in both WT and KO animals. The source of the BAL IgE was not readily apparent, since either increased serum leak into the airways or increased local production of IgE would result in similar findings. We analyzed IgE’s local production by immunohistochemistry in the allergic lungs 7 days after allergen challenge, a time point at which both WT and KO animals had peak production of this immunoglobulin. Immunohistochemical analysis showed that naïve controls of WT (Fig 5A) and KO (Fig 5C) groups had very few IgE-producing cells. While IgE staining was observed in the peribronchovascular areas in the WT lungs at day 7 (Fig 5B), IgE-stained cells were localized to the perivascular areas in the KO allergic lung (Fig 5D). Contrasting with the ELISA data, IgE staining was much more prominent in WT lungs (Fig 5B) than in KO lungs (Fig 5D), an observation that also supports the extrapulmonary production of the majority of IgE in the KO animals.

Figure 5.

Figure 5

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Immunohistochemical staining for IgE (A-D) and IgA (E-H) localization in naïve and day 7 wild-type (WT) and VPAC2 knock-out (KO) lungs. Cells that localized IgE in naïve WT (A) and KO (C) mice were scarce. The pattern of IgE staining in the WT at day 7 showed the presence of immune complexes (arrows, B) rather than cellular localization. In the KO lungs at day 7, a few IgE localizing cells were found around the perivascular areas (D). Naïve lungs in both groups had very few cells that localized IgA (E & G). Cells that localized IgA was identified in the peribronchovascular areas in the WT lungs at day 7 (F). Compared to WT, the day 7 KO lungs contained substantial numbers of IgA localizing cells around the airways and blood vessels (H). Scale bars = 200 μm

Mucosal protection is largely mediated by IgA that is actively pumped across the mucus membrane. While naïve lungs of both the WT (Fig 5E) and KO (Fig 5G) groups showed little evidence of active IgA production, WT allergic lungs contained IgA-producing cells in the peribronchovascular areas (Fig 5F). IgA-positive cells which were obvious in peribronchovascular areas in VPAC2 KO allergic lungs, were more abundant than in the WT lung (Fig 5H). These cells may contribute to the local IgA in the BAL fluid quantified by ELISA.

4. Discussion

VIP is a potent anti-inflammatory cytokine and neuropeptide that has been targeted as a potential bronchodilator for therapeutic use in asthma. However, its production by leukocytes in addition to the nervous system contribution, its ability to signal through multiple receptors, and the widespread availability of these receptors on leukocytes and structural cells of the lung makes VIP a complex peptide to study in health and disease. Using a mouse strain deficient in VPAC2, we show for the first time that VPAC2’s regulation significantly impacts antibody production by B cells. In this study, VPAC2 regulation restrained systemic inflammatory antibody production in the naïve animal, a fact that has not been recognized before. This is in contrast to other reports that failed to identify significant differences in the immune regulation between WT and KO animals [15].

The immunoregulatory role played by VIP is complex and involves cells of the innate and adaptive branches. T lymphocyte differentiation into various subsets is dependent on the signals conveyed by antigen presenting cells. The type and load of antigen, the activation state of the antigen presenting cell and the genetic background of the host with the cytokine microenvironment available, all mediate the T cell bias toward a TH1/TH2 phenotype [8]. Macrophages are capable of producing various cytokines depending on the stimulus. Macrophages express VPAC receptors and those that mature in the presence of VIP reduce IL-12 production [6] and increase IL-10 [5]. Functions of dendritic cells (DCs) depend on the mediators in the environment that they are in at the time of antigen uptake. DCs that mature in the presence of VIP, downregulate CD80 and CXCL10, while upregulating the production of CCL22, thereby preferentially attracting TH2 cells [4]. The decreased VIP in the VPAC2 knock-out lung immediately after fungal challenge [34] removes VIP’s effect on dendritic cells that transport antigen for presentation to T cells in the lymph nodes, thereby limiting the interaction between TH2 cells and allowing TH1 polarization instead. Since VPAC2 elevates the migration of TH2 cells [9] TH2 migration toward antigen presenting dendritic cells may be hindered in the absence of VPAC2 further promoting dendritic cell interaction with TH1 cells (Fig 6).

Figure 6.

Figure 6

Figure 6

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Hypothetical model for VIP’s regulation of antibody production in the presence and absence of the VPAC2 receptor. In the presence of VPAC2, VIP’s effect on dendritic cells (DCs) would promote the preferential recruitment and activation of TH2 cells to produce IL-10 that suppresses macrophage (Mφs) production of IL-12 to negatively impact TH1 activation (A). In the absence of VPAC2, VIP’s effect on the TH2 population would be reduced thereby leading to a reduction in pro-inflammatory Mφ function. The promotion and activation of the TH1 population will therefore promote preferential class switching to the IgG2a and IgA in the VPAC2 KO mice (B). IL = interleukin; CCL = CC chemokines ligand; IFN = interferon; BAFF = B cell activating factor of the TNF family

Analysis in naïve VPAC2 mice has led to the conclusion that there was no immunological difference in these mice compared to their WT counterparts [15]. However, we have shown that naïve VPAC2 KO mice had far greater abundance of serum IgG2a and IgA compared to WT, marking this as the major differentiating characteristic between the two groups. We did not find evidence of enlarged spleens or differences in splenic CD4+ T cells and CD19+ B cells in naïve KO mice (data not shown), confirming the findings by by Goetzl and colleagues [15]. Therefore, the increased production of systemic antibodies in the KO could not result from increased lymphocytes. Naïve B cells require antigen stimulus, CD40 signaling, and cytokines in order to activate switch enzymes that dictate class-switch recombination [39]. Of the various cytokines, IFNγ, IL-4, and TGFβ are the main cytokines known to induce class-switching to IgG, IgE, and IgA respectively [12]. The generation of each isotype analyzed is dependent on T cell help via CD40L and cytokines. T cell migration is promoted by VIP [23]. However, in our model, the equal expression of CD40L mRNA (data not shown) between WT and KO indicates that the massive induction in VIP mRNA in the WT at day 3 did not result in an equally greater T cell recruitment [34]. However, the possibility that it may regulate the number of CD40L expressed on each T cell will need to be investigated in the future.

In the absence of VPAC2, VIP may act as a secretogogue through VPAC1 on TH2 cells (activated independent of VIP) to enhance cytokine secretion. The VIP/VPAC2 axis acts on T cells to reduce IL-2 [14] and IFNγ [15], increase IL-4 [15], and acts on macrophages to decrease IL-6 and IL-12 [7] all of which hinders a TH1 response. The diminished regulatory effect of VIP may skew the TH1/TH2 balance in favor of TH1 cells thereby promoting B cell switch recombination toward IgG2a and IgA in the VPAC2 KO mice (Fig 6).

Antibodies induced in the allergic lungs marked by their presence in the BAL fluid may aid in the clearance of the inhaled fungal conidia. Allergic asthma is characterized by increased mediators such as histamine and leukotrienes which result in blood vessel permeability [20]. Therefore, circulating antibodies from vascular leakage probably adds to the antibodies produced in situ in the allergic lungs by recruited and resident B cells. Although IgE is not known to perform a mucosal defense function like IgA, it has been found in the BAL fluid of asthmatics [30] and allergen challenged mice contain immune complexes with IgE in the BAL fluid [40]. Based on our staining pattern, IgE containing immune complexes surrounding the peribronchovascular areas of allergic lungs may serve as a method of fungal clearance in the WT lungs. In the VPAC2 null mice, the IgE-staining was modest although the amount of IgE in the BAL fluid was significantly greater than that of WT suggesting that these immune complexes with IgE may largely be in the airway lumen of KO lungs. The systemic and local availability of IgE in this “TH1-skewed” mouse suggests that VPAC2 is not sufficient to control the generation of TH2 cells and IL-4 production in allergic asthma.

By performing numerous functions such as neutralization, immune exclusion, and antigen clearance in the lumen, IgA is believed to be vital for mucosal defense. B cells produce VIP and express VPAC1 [16], and VIP has been shown to induce class switching in CD40-activated B cells [13]. VIP induces B cell class switching to IgA directly [13, 27] or indirectly through regulation of IgA-inducing protein expression by DCs [11]. Peyer’s patches contain the most amount of IgA producing cells [3], and the gut-associated lymphoid organs are the predominant site of IgA production [21]. Synapsing of VIPergic nerves at these locations perfectly positions VIP to regulate T cells, DCs, and B cells to regulate IgA production. Similarly, we propose based on the close proximity of IgA stained cells and VIP localizing columnar epithelia, that IgA induction and secretion in the lung occurs in situ as part of mucosal defense which may be hindered in the presence of VPAC2. The decrease in VPAC2 compared to an increase in VPAC1 in the WT allergic lungs [34] indicates that VPAC1 may be the predominant VIP receptor in allergic asthma. However, this does not speak to individual cell expression of VIP receptors since we looked at VIP receptor expression in the whole lung rather than cellular subsets. The immediate reduction in VIP in the KO may allow unhindered production of IL-6 by activated macrophages promoting class-switching to IgA [1] (Fig 6).

Decades have passed since the discovery of VIP by Said and Mutt [33]. While our understanding of VIP has expanded, its effects on the respiratory and immune systems in allergic asthma are still unclear. The VPAC2 null mouse has shown that the VIP/VPAC2 axis neither impedes organ development [18, 19] nor immune system development [15]. While Goetzl and colleagues have shown that the VPAC2 null mice have decreased ITH and increased DTH responses [15], we have shown that these mice are capable of developing allergic responses against A. fumigatus allergens [34]. Furthermore, in this report, we show for the first time, that the VIP/VPAC2 axis plays a significant role in the regulation of immunoglobulin production by B cells. The absence of the regulatory receptor VPAC2 promotes the TH1 skewing of the C57BL/6 mouse strain further or fails to shift the balance in favor of TH2. Nonetheless, VPAC2 knock-out mice are still susceptible to a TH2 mediated disease such as allergic asthma indicating the fine balance between TH1 and TH2 immune phenotypes in disease pathogenesis. Taken as a whole, this research supports the increasing body of knowledge that VIP regulates inflammatory responses by signaling through VPAC2 receptors on leukocytes. Future studies analyzing various cell types such as T and B lymphocytes, and antigen presenting cells from allergic WT and VPAC2 KO mice will shed light on the intricate cellular interactions that occur between these cells with/without the VPAC2 receptor.

Acknowledgments

The authors wish to thank Dr. Anthony Harmar (University of Edinburgh, UK) for providing mice heterozygous for VPAC2 for the generation of VPAC2 knock-out mice. This project was made possible by NIH/NCRR2P20 RR015566 (JS, including the use of the Core Biology Facility), NIAID/NIH 1R15AI69061 (JS), and through a faculty development grant from the NDSU Advance FORWARD program (NSF HRD-0811239). Additional grant funding was provided by the following grant: P20 RR015566-08577 Centers of Biomedical Research Excellence: Proteases in Disease, Mukund Sibi (PI)

Footnotes

Disclosures

The authors declare no conflict of interest.

Author contributions

A.E.S. and J.M.S. designed the study; A.E.S. and S.A.H. performed the research; A.E.S. analyzed the data; A.E.S. wrote the paper; A.E.S., S.A.H., and J.M.S. edited the paper.

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Contributor Information

Amali E. Samarasinghe, Email: a.samarasinghe@ndsu.edu.

Scott A. Hoselton, Email: Scott.Hoselton@ndsu.edu.

Jane M. Schuh, Email: Jane.Schuh@ndsu.edu.

References

  • 1.Bao S, Beagley KW, Allanson M, Husband AJ. Exogenous IL-6 promotes enhanced intestinal antibody responses in vivo. Immunology and Cell Biology. 1998;76:560–2. doi: 10.1046/j.1440-1711.1998.00785.x. [DOI] [PubMed] [Google Scholar]
  • 2.Bellinger DL, Lorton D, Brouxhon S, Felten S, Felten DL. The significance of vasoactive intestinal polypeptide (VIP) in immunomodulation. Adv Neuroimmunol. 1996;6:5–27. doi: 10.1016/s0960-5428(96)00008-3. [DOI] [PubMed] [Google Scholar]
  • 3.Craig SW, Cebra JJ. Peyer’s patches: an enriched source of precursors for IgA-producing immunocytes in the rabbit. J Exp Med. 1971;134:188–200. doi: 10.1084/jem.134.1.188. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 4.Delgado M, Gonzalez-Rey E, Ganea D. VIP/PACAP preferentially attract Th2 effectors through differential regulation of chemokine production by dendritic cells. FASEB J. 2004;18:1453–5. doi: 10.1096/fj.04-1548fje. [DOI] [PubMed] [Google Scholar]
  • 5.Delgado M, Munoz-Elias EJ, Gomariz RP, Ganea D. Vasoactive intestinal peptide and pituitary adenylate cyclase-activating polypeptide enhance IL-10 production by murine macrophages: in vitro and in vivo studies. J Immunol. 1999;162:1707–16. [PubMed] [Google Scholar]
  • 6.Delgado M, Munoz-Elias EJ, Gomariz RP, Ganea D. VIP and PACAP inhibit IL-12 production in LPS-stimulated macrophages. Subsequent effect on IFNγ synthesis by T cells. Journal of Neuroimmunology. 1999;96:167–81. doi: 10.1016/s0165-5728(99)00023-5. [DOI] [PubMed] [Google Scholar]
  • 7.Delgado M, Munoz-Elias EJ, Martinez C, Gomariz RP, Ganea D. VIP and PACAP38 Modulate Cytokine and Nitric Oxide Production in Peritoneal Macrophages and Macrophage Cell Lines. Annals of the New York Academy of Sciences. 1999;897:401–14. doi: 10.1111/j.1749-6632.1999.tb07909.x. [DOI] [PubMed] [Google Scholar]
  • 8.Delgado M, Pozo D, Ganea D. The significance of vasoactive intestinal peptide in immunomodulation. Pharmacol Rev. 2004;56:249–90. doi: 10.1124/pr.56.2.7. [DOI] [PubMed] [Google Scholar]
  • 9.Dorsam G, Voice J, Kong Y, Goetzl EJ. Vasoactive intestinal peptide mediation of development and functions of T lymphocytes. Ann N Y Acad Sci. 2000;921:79–91. doi: 10.1111/j.1749-6632.2000.tb06953.x. [DOI] [PubMed] [Google Scholar]
  • 10.Dunzendorfer S, Meierhofer C, Wiedermann CJ. Signaling in neuropeptide-induced migration of human eosinophils. J Leukoc Biol. 1998;64:828–34. doi: 10.1002/jlb.64.6.828. [DOI] [PubMed] [Google Scholar]
  • 11.Endsley MA, Njongmeta LM, Shell E, Ryan MW, Indrikovs AJ, Ulualp S, et al. Human IgA-inducing protein from dendritic cells induces IgA production by naive IgD+ B cells. J Immunol. 2009;182:1854–9. doi: 10.4049/jimmunol.0801973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 12.Esser C, Radbruch A. Immunoglobulin Class Switching: Molecular and Cellular Analysis. Annu Rev Immunol. 1990;8:717–35. doi: 10.1146/annurev.iy.08.040190.003441. [DOI] [PubMed] [Google Scholar]
  • 13.Fujieda S, Waschek JA, Zhang K, Saxon A. Vasoactive Intestinal Peptide Induces Sα/Sμ Switch Circular DNA in Human B Cells. J Clin Invest. 1996;98:1527–32. doi: 10.1172/JCI118944. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 14.Ganea D. Regulatory effects of vasoactive intestinal peptide on cytokine production in central and peripheral lymphoid organs. Advances in Neuroimmunology. 1996;6:61–74. doi: 10.1016/s0960-5428(96)00007-1. [DOI] [PubMed] [Google Scholar]
  • 15.Goetzl EJ, Voice JK, Shen S, Dorsam G, Kong Y, West KM, et al. Enhanced delayed-type hypersensitivity and diminished immediate-type hypersensitivity in mice lacking the inducible VPAC(2) receptor for vasoactive intestinal peptide. Proc Natl Acad Sci U S A. 2001;98:13854–9. doi: 10.1073/pnas.241503798. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 16.Gomariz RP, Leceta J, Garrido E, Garrido T, Delgado M. Vasoactive intestinal peptide (VIP) mRNA expression in rat T and B lymphocytes. Regul Pept. 1994;50:177–84. doi: 10.1016/0167-0115(94)90033-7. [DOI] [PubMed] [Google Scholar]
  • 17.Grimm MC, Newman R, Hassim Z, Cuan N, Connor SJ, Le Y, et al. Vasoactive Intestinal Peptide Acts as a Potent Suppressor of Inflammation In Vivo by Trans-Deactivating Chemokine Receptors. The Journal of Immunology. 2003;171:4990–4. doi: 10.4049/jimmunol.171.10.4990. [DOI] [PubMed] [Google Scholar]
  • 18.Harmar AJ, Marston HM, Shen S, Spratt C, West KM, Sheward WJ, et al. The VPAC(2) receptor is essential for circadian function in the mouse suprachiasmatic nuclei. Cell. 2002;109:497–508. doi: 10.1016/s0092-8674(02)00736-5. [DOI] [PubMed] [Google Scholar]
  • 19.Harmar AJ, Sheward WJ, Morrison CF, Waser B, Gugger M, Reubi JC. Distribution of the VPAC2 receptor in peripheral tissues of the mouse. Endocrinology. 2004;145:1203–10. doi: 10.1210/en.2003-1058. [DOI] [PubMed] [Google Scholar]
  • 20.Hedqvist P, Gautam N, Lindbom L. Interactions between Leukotrienes and Other Inflammatory Mediators/Modulators in the Microvasculature. American Journal of Respiratory and Critical Care Medicine. 2000;161:S117–S9. doi: 10.1164/ajrccm.161.supplement_1.ltta-23. [DOI] [PubMed] [Google Scholar]
  • 21.Husband AJ, Gowans JL. The origin and antigen-dependent distribution of IgA-containing cells in the intestine. J Exp Med. 1978;148:1146–60. doi: 10.1084/jem.148.5.1146. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 22.Ichinose M, Nakajima N, Takahashi T, Yamauchi H, Inoue H, Takishima T. Protection against bradykinin-induced bronchoconstriction in asthmatic patients by neurokinin receptor antagonist. Lancet. 1992;340:1248–51. doi: 10.1016/0140-6736(92)92948-f. [DOI] [PubMed] [Google Scholar]
  • 23.Johnston JA, Taub DD, Lloyd AR, Conlon K, Oppenheim JJ, Kevlin DJ. Human T Lymphocyte Chemotaxis and Adhesion Induced by Vasoactive Intestinal Peptide. The Journal of Immunology. 1994;153:1762–8. [PubMed] [Google Scholar]
  • 24.Joos GF. The role of neuroeffector mechanisms in the pathogenesis of asthma. Curr Allergy Asthma Rep. 2001;1:134–43. doi: 10.1007/s11882-001-0081-8. [DOI] [PubMed] [Google Scholar]
  • 25.Joos GF, De Swert KO, Pauwels RA. Airway inflammation and tachykinins: prospects for the development of tachykinin receptor antagonists. Eur J Pharmacol. 2001;429:239–50. doi: 10.1016/s0014-2999(01)01323-1. [DOI] [PubMed] [Google Scholar]
  • 26.Karmouty-Quintana H, Cannet C, Sugar R, Fozard JR, Page CP, Beckmann N. Capsaicin-induced mucus secretion in rat airways assessed in vivo and non-invasively by magnetic resonance imaging. Br J Pharmacol. 2007;150:1022–30. doi: 10.1038/sj.bjp.0707168. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 27.Kimata H, Fujimoto M. Induction of IgA1 and IgA2 production in immature human fetal B cells and pre-B cells by vasoactive intestinal peptide. Blood. 1995;85:2098–104. [PubMed] [Google Scholar]
  • 28.Lutz EM, Sheward WJ, West KM, Morrow JA, Fink G, Harmar AJ. The VIP2 receptor: molecular characterisation of a cDNA encoding a novel receptor for vasoactive intestinal peptide. FEBS Lett. 1993;334:3–8. doi: 10.1016/0014-5793(93)81668-p. [DOI] [PubMed] [Google Scholar]
  • 29.Mukaiyama O, Morimoto K, Nosaka E, Takahashi S, Yamashita M. Involvement of enhanced neurokinin NK3 receptor expression in the severe asthma guinea pig model. Eur J Pharmacol. 2004;498:287–94. doi: 10.1016/j.ejphar.2004.07.071. [DOI] [PubMed] [Google Scholar]
  • 30.Peebles RS, Jr, Liu MC, Adkinson NF, Jr, Lichtenstein LM, Hamilton RG. Ragweed-specific antibodies in bronchoalveolar lavage fluids and serum before and after segmental lung challenge: IgE and IgA associated with eosinophil degranulation. J Allergy Clin Immunol. 1998;101:265–73. doi: 10.1016/S0091-6749(98)70392-6. [DOI] [PubMed] [Google Scholar]
  • 31.Pozo D, Delgado M. The many faces of VIP in neuroimmunology: a cytokine rather a neuropeptide? FASEB J. 2004;18:1325–34. doi: 10.1096/fj.03-1440hyp. [DOI] [PubMed] [Google Scholar]
  • 32.Renz H. Neurotrophins in bronchial asthma. Respir Res. 2001;2:265–8. doi: 10.1186/rr66. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 33.Said SI, Mutt V. Polypeptide with broad biological activity: isolation from small intestine. Science. 1970;169:1217–8. doi: 10.1126/science.169.3951.1217. [DOI] [PubMed] [Google Scholar]
  • 34.Samarasinghe AE, Hoselton SA, Schuh JM. The absence of the VPAC(2) receptor does not protect mice from Aspergillus induced allergic asthma. Peptides. 2010;31:1068–75. doi: 10.1016/j.peptides.2010.03.001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 35.Tiberio IF, Leick-Maldonado EA, Miyahara L, Kasahara DI, Spilborghs GM, Martins MA, et al. Effects of neurokinins on airway and alveolar eosinophil recruitment. Exp Lung Res. 2003;29:165–77. doi: 10.1080/01902140303772. [DOI] [PubMed] [Google Scholar]
  • 36.Usdin TB, Bonner TI, Mezey E. Two receptors for vasoactive intestinal polypeptide with similar specificity and complementary distributions. Endocrinology. 1994;135:2662–80. doi: 10.1210/endo.135.6.7988457. [DOI] [PubMed] [Google Scholar]
  • 37.Veres TZ, Shevchenko M, Krasteva G, Spies E, Prenzler F, Rochlitzer S, et al. Dendritic cell-nerve clusters are sites of T cell proliferation in allergic airway inflammation. Am J Pathol. 2009;174:808–17. doi: 10.2353/ajpath.2009.080800. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 38.Voice JK, Dorsam G, Lee H, Kong Y, Goetzl EJ. Allergic diathesis in transgenic mice with constitutive T cell expression of inducible vasoactive intestinal peptide receptor. FASEB J. 2001;15:2489–96. doi: 10.1096/fj.01-0671com. [DOI] [PubMed] [Google Scholar]
  • 39.Wykes M, Pombo A, Jenkins C, MacPherson G. Dendritic Cells Interact Directly with Naive B Lymphocytes to Transfer Antigen and Initiate Class Switching in a Primary T-Dependent Response. Journal of Immunology. 1998;161:1313–9. [PubMed] [Google Scholar]
  • 40.Zuberi RI, Apgar JR, Chen SS, Liu FT. Role for IgE in airway secretions: IgE immune complexes are more potent inducers than antigen alone of airway inflammation in a murine model. J Immunol. 2000;164:2667–73. doi: 10.4049/jimmunol.164.5.2667. [DOI] [PubMed] [Google Scholar]

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