The JI
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     
 

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Raap, U.
Right arrow Articles by Renz, H.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Raap, U.
Right arrow Articles by Renz, H.
The Journal of Immunology, 2003, 171: 353-359.
Copyright © 2003 by The American Association of Immunologists

{alpha}-Melanocyte-Stimulating Hormone Inhibits Allergic Airway Inflammation1

Ulrike Raap*, Thomas Brzoska{dagger}, Sirius Sohl*, Günter Päth*, Jörg Emmel*, Udo Herz*, Armin Braun*, Thomas Luger{dagger} and Harald Renz2,*

* Department of Clinical Chemistry and Molecular Diagnostics, Philipps-University Marburg, Marburg, Germany; and {dagger} Department of Dermatology, Ludwig-Boltzmann Institute, University Muenster, Muenster, Germany


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
{alpha}-Melanocyte-stimulating hormone ({alpha}-MSH) is a neuropeptide controlling melanogenesis in pigmentary cells. In addition, its potent immunomodulatory and immunosuppressive activity has been recently described in cutaneous inflammatory disorders. Whether {alpha}-MSH is also produced in the lung and might play a role in the pathogenesis of inflammatory lung conditions, including allergic bronchial asthma, is unknown. Production and functional role of {alpha}-MSH were investigated in a murine model of allergic airway inflammation. {alpha}-MSH production was detected in bronchoalveolar lavage fluids. Although aerosol challenges stimulate {alpha}-MSH production in nonsensitized mice, this rapid and marked stimulation was absent in allergic animals. Treatment of allergic mice with {alpha}-MSH resulted in suppression of airway inflammation. These effects were mediated via IL-10 production, because IL-10 knockout mice were resistant to {alpha}-MSH treatment. This study provides evidence for a novel function of {alpha}-MSH linking neuroimmune functions in allergic airway inflammation.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
{alpha}-Melanocyte-stimulating hormone (MSH)3 is an endogenous neuroimmunomodulatory peptide derived from the precursor molecule proopiomelanocortin (POMC). Originally, POMC was discovered in the pituitary gland, but subsequently, POMC-derived peptides were also detected in lymphocytes, monocytes, Langerhans cells, and epithelial cells (1, 2, 3, 4, 5).

{alpha}-MSH is well known for its role in the control of melanogenesis in pigmentary cells. However, recent studies demonstrated a potent and broad spectrum of activities as an antipyretic, antimicrobial, anti-inflammatory, and immunomodulatory peptide (6, 7, 8). The actions of {alpha}-MSH are transmitted via specific melanocortin (MC) receptors (9, 10, 11). Five different subtypes of MC receptors have been identified, designated MC-1 to MC-5. Melanocortin receptor-1 (MC-1), which is specific for {alpha}-MSH, is expressed on skin keratinocytes, dendritic cells, macrophages, endothelial cells, and epithelial cells (10, 12).

The skin is an extrapituitary site of {alpha}-MSH generation and secretion. Elevated {alpha}-MSH levels have been reported in several cutaneous inflammatory disorders, including psoriasis vulgaris and eczema (13, 14). The potent anti-inflammatory property of {alpha}-MSH was shown in a murine model of delayed-type hypersensitivity and hapten-specific tolerance (15, 16, 17). In the latter model, {alpha}-MSH-induced hapten-specific tolerance in both a preventive as well as therapeutic treatment regimen.

In addition to skin, the airways represent another primary site of the allergic response. To date, it remains unknown whether {alpha}-MSH is also produced by cells residing in or invading lung and airways during allergic inflammation. Furthermore, MC receptor expression and subsequent effects of {alpha}-MSH have not been studied in this compartment. These questions were addressed in the present study, which was conducted in a well-characterized model of allergic airway inflammation with Th2 cell influx and eosinophilia, as well as the presence of immediate-type hypersensitivity responses related to IgE/IgG1 Ab production (18, 19).


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Animals

Female BALB/c and C57BL/6 mice (6–8 wk old) were obtained from Harlan Winkelmann (Borchen, Germany); homozygous C57BL/6-IL-10tm1Cgn (6–8 wk old) were purchased from The Jackson Laboratory (Bar Harbor, ME). Animals were housed in a pathogen-free facility with single ventilated cages and received OVA-free diet and water ad libitum.

Allergic sensitization, allergen challenge, and administration of {alpha}-MSH

Mice were sensitized by three i.p. injections of 10 µg OVA grade VI (Sigma-Aldrich, Deisenhofen, Germany) absorbed to 1.5 mg AL(OH)3 (Imject Alum; Pierce, Rockford, IL) diluted in 200 µl PBS on days 1, 14, and 21. Nonsensitized animals (control) received adjuvant 1.5 mg AL(OH)3 diluted in PBS alone. {alpha}-MSH and {gamma}-MSH (Bachem, Heidelberg, Germany) were stored in aliquots until use at -80°C. Immediately before injection, the peptides were dissolved in PBS containing 0.1% BSA and kept on ice until application. A total of 1 mg/kg body weight of {alpha}-MSH or {gamma}-MSH was injected into the tail vein 30 min before sensitization or allergen aerosolization ({alpha}-MSH/OVA and {gamma}-MSH/OVA). {gamma}-MSH is a POMC gene-derived peptide, with an amino acid sequence different from {alpha}-MSH and no published immunomodulatory activity. All animals received two local allergen challenges performed by an exposure to 1% OVA diluted in PBS for 20 min on days 26 and 27. Experimental groups corresponding to this protocol are depicted in Fig. 1.



View larger version (23K):
[in this window]
[in a new window]
  		FIGURE 1. Study protocol. Experimental groups (code) and substances used during sensitization (PBS, OVA), treatment ({gamma}-MSH, {alpha}-MSH), and challenges (OVA). Control animals were not sensitzed to OVA in contrast to all other groups. {alpha}-MSH/OVA and {gamma}-MSH/OVA mice received peptides 30 min before sensitization to and challenge with OVA. All animals were OVA aerosol challenged on days 26 and 27.

 
Ig assays

Blood was collected by incision of the tail vein. Total IgE and allergen-specific IgE, IgG1, and IgG2a Ab concentrations were measured in serum samples by ELISA, as previously described (18, 20).

Bronchoalveolar lavage and lung histology

Bronchoalveolar lavage (BAL) fluid was taken 48 h after the last allergen challenge using a method previously described (21). BAL samples with a volume of 1.4 ± 0.2 ml were assessed for further analysis. BAL cells were counted, an aliquot was taken for cytospin centrifugation (700 x g), and cytospins were stained with Diff-Quik (Dade Behring, Düdingen, Switzerland) for differential cell counting. From remaining BAL fluids, cells were removed and supernatants were aliquoted for further ELISA analysis. For cryostat sections, the lung was removed, embedded with Tissue-Tek (Sakura, Zoeterwute, The Netherlands), diluted in PBS 1/4, and cryoconservated. Sections (8 µm) were cut, dried for 30 min, and stained with 1% H&E (Merck, Darmstadt, Germany).

Cytokine assays

The cytokines IL-4, IL-5, IL-10, IL-13, and IFN-{gamma} were measured in BAL fluids by ELISA, as described elsewhere (18). Kits for cytokine immunoassays were purchased from B&D (Heidelberg, Germany) for IL-4, IL-5, and IFN-{gamma}, and from R&D Systems (Minneapolis, MN) for IL-10 and IL-13. Detection limits were 3.4 pg/ml for IL-4, 7.8 pg/ml for IL-5 and IL-10, and 31.25 pg/ml for IL-13 and IFN-{gamma}, respectively.

{alpha}-MSH RIA

{alpha}-MSH concentrations in BAL fluids were determined using a competitive RIA. {alpha}-MSH in samples competed with 125I-labeled {alpha}-MSH (125I-{alpha}-MSH) in binding to an antiserum, which was raised against an {alpha}-MSH-albumin conjugate. To increase the sensitivity of the assay, 125I-{alpha}-MSH was added delayed. Ab-bound 125I-{alpha}-MSH was separated from the free fraction using the double Ab polyethylene glycol precipitation technique. The radioactivity of the precipitate was measured. The antiserum used in this assay was directed to the C-terminal part of the {alpha}-MSH molecule and showed no cross-reactivity with adrenocorticotropic hormone. Briefly, 100 µl of samples was pipetted in 3-ml glass tubes, 200 µl anti-{alpha}-MSH serum was added, and the mixture was incubated for 24 h at 4°C. On the following day, 200 µl 125I-{alpha}-MSH was added and the mix was incubated for further 24 h at 4°C. On day 3, double Ab polyethylene glycol (500 µl) was added and the mixture was incubated for another 60 min. Finally, vials were centrifuged, supernatants were decanted, and the radioactivity was measured in the precipitates using a gamma counter (counting time, 3 min).

Flow cytometry and FACS analysis

BAL samples with a volume of 1.4 ± 0.2 ml were taken 48 h after the last allergen challenge. BAL fluids of five mice per group were pooled and pelleted (700 x g). Cells were prepared for further analysis (22). Cell samples were analyzed using a FACScan flow cytometer (BD Biosciences, San Jose, CA). The data were processed with CellQuest software (BD Biosciences). Before staining procedure, cells were incubated with 2% heat-inactivated mouse serum so as to avoid unspecific bindings. All incubations were performed in HBSS, supplemented with 2% BSA. MAC-3 surface expression was measured with a mAb clone M3/M84 (BD PharMingen, Heidelberg, Germany), PE-conjugated, isotype control: rat IgG1 {kappa} PE. MC-1 surface expression was assessed using a polyclonal rabbit anti-mouse Ab custom made by Inventus Biotec (Muenster, Germany) directed against the aa sequences 2–18 of the extracellular domain. As a secondary Ab, a FITC-conjugated polyclonal goat anti-rabbit Ab was used (Dianova, Hamburg, Germany). Staining procedure was performed on ice for 15 min, and cells were washed twice with cold PBS. Total leukocytes were gated, and counting consisted of 10,000 events. Quadrants were defined by isotype/secondary Ab controls.

Determination of airway responsiveness

Measurements of airway responsiveness were performed 24 h after the last challenge (day 28) by head-out body plethysmography, as previously described (23). Air flow signals were recorded in response to methacholine (0, 12.5, 25, 50, 100, 200 mg/ml) and aerosolized with a nebulizer. The concentration of methacholine that caused a 50% reduction in expiratory air flow (MCh50, milligrams per milliliter) was determined.

Data analysis

Results are presented as mean ± SEM or SD. The statistical significance between study groups was analyzed by Student’s t test. A p value of <0.05 was accepted as statistically significant.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
Production of {alpha}-MSH in the lung during allergic airway inflammation

The sensitization and airway allergen challenge protocol resulted in a rise of total and allergen-specific IgE, IgG1, and IgG2a Ab production (Fig. 2). A marked influx of inflammatory cells into the airways with predominance of eosinophils (Fig. 3) together with increased levels of IL-4, IL-5, and IL-13 cytokine production (Table I) was observed following airway allergen challenge. The inflammatory response was paralleled by decreased levels of IL-10 in BAL fluids (Fig. 4). The levels of {alpha}-MSH production were assessed in BAL at consecutive time points after airway allergen challenge. {alpha}-MSH was detected in all animals. Airway allergen challenge of nonsensitized mice triggered a rapid and marked rise in {alpha}-MSH levels. By day 4, levels returned to baseline. This increase did not occur in allergic animals (Fig. 5).



View larger version (21K):
[in this window]
[in a new window]
  		FIGURE 2. {alpha}-MSH inhibits allergen-specific Ab production. Depicted are total IgE (A), allergen-specific IgE (B), IgG1 (C), and IgG2a (D) serum levels of controls, NIL/OVA-, {gamma}-MSH/OVA-, and {alpha}-MSH/OVA-treated BALB/c mice. In each group, 7–12 mice were analyzed. Presented are mean ± SEM. Significances are shown for {alpha}-MSH/OVA-treated mice compared with NIL/OVA mice (*, p < 0.05; ***, p < 0.001).

 


View larger version (16K):
[in this window]
[in a new window]
  		FIGURE 3. {alpha}-MSH inhibits eosinophil influx into the airways. Cells from BAL fluids stained with Diff-Quik for differential cell counting of controls (n = 12), NIL/OVA (n = 12), {gamma}-MSH/OVA (n = 10), and {alpha}-MSH/OVA (n = 12). Indicated are statistically significant differences between NIL/OVA- and {alpha}-MSH/OVA-treated mice concerning total cell count (A), macrophages (B), eosinophils (C), neutrophils (D), and lymphocytes (E) (*, p < 0.05; ***, p < 0.001). Presented are mean ± SEM.

 

View this table:
[in this window]
[in a new window]
  		Table I. {alpha}-MSH decreases IL-4 and IL-13 production in BAL fluida

 


View larger version (14K):
[in this window]
[in a new window]
  		FIGURE 4. {alpha}-MSH significantly increases IL-10 production. Detection of IL-10 in BAL fluids 48 h after last allergen challenge of controls (n = 12), during acute airway inflammation (NIL/OVA, n = 9), and in {gamma}-MSH ({gamma}-MSH/OVA, n = 8)- and {alpha}-MSH-treated mice ({alpha}-MSH/OVA, n = 8) (*, p < 0.05; ***, p < 0.001; shown are mean ± SEM).

 


View larger version (14K):
[in this window]
[in a new window]
  		FIGURE 5. {alpha}-MSH is detectable in BAL fluids. At indicated time points after the last nasal allergen challenge, murine {alpha}-MSH was measured in BAL fluids of BALB/c mice with acute airway inflammation (NIL/OVA; open bars, n = 5), compared with controls (nasal challenge alone; filled bars, n = 5; **, p < 0.01; *, p < 0.05). Baseline {alpha}-MSH levels were assessed from animals that received neither allergen sensitization nor nasal challenges (n = 7, gray bars) as shown mean ± SEM.

 
{alpha}-MSH inhibits allergic immunoresponses

To assess possible effects of {alpha}-MSH on the allergic response, mice were treated with synthetic {alpha}-MSH peptide according to the treatment protocol (Fig. 1). Significantly decreased levels of allergen-specific IgE (p < 0.05), IgG1 (p < 0.05), and IgG2a (p < 0.001), but not total IgE, were detected in {alpha}-MSH/OVA mice as compared with untreated (NIL)/OVA (Fig. 2). Treatment with {gamma}-MSH was without effect on IgE production, but resulted in a small, nonsignificant decrease in IgG1 and IgG2a Ab levels.

Cell numbers and distribution of leukocyte subpopulations were determined in BAL fluids obtained 48 h after the last allergen challenge. Total cell numbers did not differ among NIL/OVA, {alpha}-MSH/OVA, and {gamma}-MSH/OVA mice (Fig. 3). In NIL/OVA mice, the inflammatory infiltrate consisted mainly of eosinophils->macrophages->neutrophils and lymphocytes. {alpha}-MSH treatment prevented the influx of eosinophils to a large extent (p < 0.001). In parallel, a higher number of neutrophils was detected in these mice. The latter effect was also observed in {gamma}-MSH-treated mice, although it did not reach statistical significance. On cryoconservated lung sections from NIL/OVA animals, a peribronchial and perivascular inflammatory cell infiltrate was detectable, and airway epithelium showed signs of cell metaplasia and mucus hypersecretion that were absent in control mice. In contrast, {alpha}-MSH prevented the development of perivascular inflammation (Fig. 6).



View larger version (74K):
[in this window]
[in a new window]
  		FIGURE 6. {alpha}-MSH inhibits peribronchial inflammatory cell infiltrate. Histological images of the OVA triggered influx of inflammatory cells into the lung and airways. Staining with H&E. Lung sections from control (A), NIL/OVA (B, arrow points to mucus production), and {alpha}-MSH/OVA (C, arrow points to morphologically normal airway apithelium) mice. Presented is one of three similar experiments. Magnification x50. D, High power field analysis of mucosal eosinophils (10 fields of vision; blinded investigators; mean ± SD; p < 0.05).

 
The pattern of BAL cytokine content was markedly altered in {alpha}-MSH-treated mice. {alpha}-MSH-, but not {gamma}-MSH-treated animals had significantly lower levels of IL-4 and IL-13 as compared with NIL/OVA mice. In contrast, the concentrations of IL-5 and IFN-{gamma} remained unchanged (Table I). Alveolar macrophages represent a major source of {alpha}-MSH-responsive cells. {alpha}-MSH treatment resulted in marked up-regulation of MAC-3, a marker of activation. At least a subset of cells coexpressed the {alpha}-MSH receptor MC-1. MC-1-positive macrophages were also detected in control and NIL/OVA mice (Fig. 7).



View larger version (25K):
[in this window]
[in a new window]
  		FIGURE 7. Macrophage activation and MC-1 surface expression. Assessment of activated (MAC-3 receptor-positive) and MC-1 receptor-positive macrophages in BAL fluid. Presented are dot plots of total leukocyte populations of five pooled BAL pellets of control, NIL/OVA, and {alpha}-MSH/OVA mice. Numbers in the upper right corners reflect the percentage of activated macrophages (MAC-3 positive) that were MC-1 receptor positive. Numbers of the lower right cases represent activated macrophages (MAC-3 positive) that were MC-1 receptor negative. Presented is one representative experiment of two.

 
Effect of {alpha}-MSH on airway responsiveness

Airway responsiveness to methacholine was assessed 24 h after the last airway allergen challenge. The dose of methacholine causing a 50% reduction in expiratory air flow was 71 ± 14 mg/ml in the control group. In contrast, a dose of only 50 ± 9.8 mg/ml (NIL/OVA) and 49 ± 13.4 mg/ml ({gamma}-MSH/OVA) was required to reach the same effect in allergic and {gamma}-MSH-treated mice, indicating airway hyperresponsiveness (p < 0.001). {alpha}-MSH-treated allergic mice showed a tendency toward normalization with 56.6 ± 14 mg/ml (not significant compared with controls, NIL/OVA, and {gamma}-MSH/OVA mice).

Effect of {alpha}-MSH is IL-10 dependent

IL-10 levels were significantly reduced in allergic mice. This effect was also observed in {gamma}-MSH-treated mice. In {alpha}-MSH-treated allergic mice, however, this effect was reverted, as these mice showed higher IL-10 levels than NIL/OVA mice (p < 0.001) and controls (p < 0.05) (Fig. 4). To investigate whether the inhibitory effect of {alpha}-MSH on allergic airway inflammation is mediated via IL-10 production, IL-10 knockout animals were used. As IL-10 knockout mice are bred on a C57BL/6 background, the effects of {alpha}-MSH were also studied in this strain. Similar to sensitized BALB/c mice, {alpha}-MSH suppressed the influx of eosinophils (Fig. 8) in the airways, paralleled by a further increase of neutrophils in C57BL/6 animals. Allergen-specific Ab production, except for IgG1, was suppressed (Fig. 9). In IL-10-/- mice, treatment with {alpha}-MSH was without effect on Ab production and airway inflammation, indicating the dependency of {alpha}-MSH effects on IL-10.



View larger version (15K):
[in this window]
[in a new window]
  		FIGURE 8. {alpha}-MSH prevents eosinophil influx in C57BL/6 wild type (wt), but not in IL-10-/-. Indicated are total cell count (A, B), eosinophils (C, D), and neutrophils (E, F) in the BAL fluids of C57BL/6 wt (A, C, E) and IL-10-/- (B, D, F) mice. Between 8 and 10 mice were studied in each group. Differences are depicted for {alpha}-MSH/OVA compared with NIL/OVA (*, p < 0.05; ***, p < 0.001). Presented are mean ± SEM.

 


View larger version (17K):
[in this window]
[in a new window]
  		FIGURE 9. {alpha}-MSH decreases allergen-specific Ig in C57BL/6 wild type (wt), but not in IL-10-/- animals. Ig production was determined by ELISA in serum samples from nonsensitized (control), OVA-sensitized (NIL/OVA), and {alpha}-MSH/OVA-treated C57BL/6 wt and IL-10-/- (NIL/OVA, {alpha}-MSH/OVA) mice. In each group, between 6 and 10 mice were analyzed. Depicted are total IgE (A, B), allergen-specific IgE (C, D), allergen-specific IgG1 (E, F), and allergen-specific IgG2a (G, H). Indicated are statistically significant differences between {alpha}-MSH/OVA and NIL/OVA mice (*, p < 0.05). Presented are mean ± SEM.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
This is the first study demonstrating {alpha}-MSH production in the lung. In normal, nonsensitized mice, a nasal Ag challenge results in enhanced {alpha}-MSH production. This effect was absent in allergic mice, pointing to a negative regulatory loop of {alpha}-MSH production in the airways. This finding together with previous results obtained in murine models of delayed-type hypersensitivity led us to the hypothesis that {alpha}-MSH may also have anti-inflammatory functions in the lung. Therefore, sensitized mice were treated with {alpha}-MSH. The results demonstrate a strong anti-inflammatory effect of {alpha}-MSH in airways and lung that is IL-10 dependent.

Production of this neuropeptide has been reported following injury, UV light exposure, trauma, and infection (24). Increased production of {alpha}-MSH was measured in patients with psoriasis vulgaris and other cutaneous inflammatory conditions (13, 14). Monocytes and macrophages are major sources of this neuropeptide. Therefore, these cells may contribute to the {alpha}-MSH level observed in BAL fluids. {alpha}-MSH has immunomodulatory effects on several cell types. It inhibits the production of a number of proinflammatory cytokines, including IL-1{beta}, IL-1{alpha}, IL-2, IL-6, IFN-{gamma}, and TNF-{alpha} (25, 26). Because many of these cytokines are produced by macrophages, {alpha}-MSH seems to designate them as key target cells (1). In fact, the receptor with high affinity for {alpha}-MSH, MC-1, has been found on macrophages and other cell populations (10). However, to our knowledge, this is the first report demonstrating expression of this receptor on macrophages of the airways. MC-1 is expressed on at least a subset of resting cells and shows up-regulation on activated macrophages. The functional activities of human mast cells are affected (27, 28). For example, the release of histamine is altered under the influence of {alpha}-MSH (28, 29). B cells are further target cells of {alpha}-MSH action (10, 30). Modulation of IgE synthesis and down-regulation of the expression of costimulatory molecules, including CD86 and CD40, have been described (1). In addition, inhibition of the expression of VCAM and E-selectin on endothelial cells was also shown (31). Moreover, in terms of intracellular signal transduction, {alpha}-MSH inhibits the activation of NF-{kappa}B (32).

In the model of allergic bronchial asthma, {alpha}-MSH suppressed allergen-specific IgE, IgG1, and IgG2a Ab production. Based on the known effects of {alpha}-MSH on B cells, it is likely that B cells are among the targets of {alpha}-MSH action. The local anti-inflammatory properties of {alpha}-MSH are characterized by a marked suppression in airway eosinophilia. This was accompanied by an increase in the influx of neutrophils. However, the suppression of airway eosinophilia was not related to decreases in IL-5 production, although IL-5 is known as a potent cytokine for eosinophil development, recruitment, activation, and survival (33, 34). The reduction of eosinophils may be related to the down-regulation of certain endothelial adhesion molecules including VCAM. The expression of VCAM, in turn, is under the close control of cytokines, including IL-4, IL-13, and TNF-{alpha} (35). The production of all these cytokines is apparently modulated by {alpha}-MSH. Why treatment with {alpha}-MSH resulted in an increased influx of neutrophils into the airways remains currently unclear. However, because this effect was also observed with {gamma}-MSH, a peptide also derived from POMC that has to date no effects on the immune system, the observed effects might be considered as unspecific. From our data, we have no indication that the influx in neutrophils was associated by a Th1 response. There are no increases in IFN-{gamma}, and IgG2a Abs were even significantly reduced by {alpha}-MSH. Furthermore, the influx in neutrophils was rather small as compared with a recent study investigating Th1 responses in the lung (36).

Similar to the effects observed in the model of delayed-type hypersensitivity, {alpha}-MSH exerts its anti-inflammatory potency also in the lung via the induction of IL-10. IL-10 is a pleiotropic cytokine with potent immunostimulatory effects on a variety of cell types. One major effect that has recently been extensively investigated is the suppression of effector functions on macrophages, monocytes, NK cells, and Th cell subsets (37). IL-10 is mainly produced by T cell subsets of the Th2 phenotype, but also by other cells, including B cells, macrophages, monocytes, keratinocytes, and mast cells (37). Increased production of IL-10 has been observed during antiallergic specific immunotherapy and corresponds to the success of this mode of therapy (38). Further analysis indicates that this effect is related to the induction of regulatory T cells by IL-10 (39). Most recently, the in vitro induction of CD25+ CD4+ regulatory T cells by {alpha}-MSH has been described. In that system of experimental autoimmune uveoretinitis, {alpha}-MSH appeared to convert a population of effector T cells polarized to mediate hypersensitivity into a population of T cells with no mediated immunoregulation (40). These regulatory T cells are characterized by the production of IL-10 among other cytokines (41). It is also well known that IL-10 prevents IL-4-induced IgE synthesis by inhibiting the accessory cell function of monocytes (42).

The association between airway inflammation and airway hyperresponsiveness is still a matter of debate. There are a number of papers indicating a link between eosinophilia and/or IL-5 production and increased airway responsiveness (43, 44, 45, 46, 47). In contrast, there is also a substantial body of literature pointing toward a dissociation between airway inflammation and airway hyperresponsiveness (18, 48, 49, 50). Even more recently, data from clinical trials using anti-IL-5 mAbs revealed unchanged lung function in the absence of eosinophils (51). We have measured airway responsiveness to methacholine. In addition, there are several other pathways controlling airway smooth muscle contractility. They include serotonin, neurokinin, and substance P-dependent mechanisms. Whether {alpha}-MSH has any effect on these other pathways still remains unknown. In addition to a qualitative relationship between eosinophilia, airway inflammation, and airway hyperresponsiveness, there might also be a quantitative relationship between these events. For example, the number of (remaining) eosinophils might be still sufficient to maintain a level of airway hyperresponsiveness.

Elevated levels of {alpha}-MSH were detected in BAL fluids of OVA-challenged, but nonsensitized mice. It is well known that repeated Ag exposure via this route induces tolerance. Furthermore, it has been shown more recently that tolerance induced by similar protocols is mediated to a large degree by IL-10 (52, 53). It might be, therefore, possible that the importance of endogenous {alpha}-MSH lies in its ability to induce airway tolerance. This would make triggering the {alpha}-MSH/{alpha}-MSH receptor pathway an interesting therapeutic target.


   Acknowledgments
 
For their excellent technical assistance, we thank Brigitte Auffarth, Helga Fischer, Steffen Heuser, Britta Kosche, Verena Kräling, Delia Miklay, Jürgen Kurz, and Caroline Schwalm.


   Footnotes
 
1 This study was supported by Deutsche Forschungsgemeinschaft Grant RE 737/7, Volkswagen Stiftung, and Kempke Stiftung 23/01. Back

2 Address correspondence and reprint requests to Dr. Harald Renz, Department of Clinical Chemistry and Molecular Diagnostics, Central Laboratory, Hospital of the Philipps University, Baldingerstr., D-35033 Marburg, Germany. E-mail address: renzh{at}med.uni-marburg.de Back

3 Abbreviations used in this paper: MSH, melanocyte-stimulating hormone; BAL, bronchoalveolar lavage; 125I-{alpha}-MSH, 125I-labeled {alpha}-MSH; MC, melanocortin; NIL, untreated; POMC, proopiomelanocortin. Back

Received for publication December 19, 2002. Accepted for publication April 17, 2003.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Luger, T. A., T. Brzoska, T. E. Scholzen, D. H. Kalden, C. Sunderkotter, C. Armstrong, J. Ansel. 2000. The role of {alpha}-MSH as a modulator of cutaneous inflammation. Ann. NY Acad. Sci. 917:232.[Abstract/Free Full Text]
  2. Bhardwaj, R. S., T. A. Luger. 1994. Proopiomelanocortin production by epidermal cells: evidence for an immune neuroendocrine network in the epidermis. Arch. Dermatol. Res. 287:85.[Medline]
  3. Scholzen, T. E., D. H. Kalden, T. Brzoska, T. Fisbeck, M. Fastrich, M. Schiller, M. Bohm, T. Schwarz, C. A. Armstrong, J. C. Ansel, T. A. Luger. 2000. Expression of proopiomelanocortin peptides in human dermal microvascular endothelial cells: evidence for a regulation by ultraviolet light and interleukin-1. J. Invest. Dermatol. 115:1021.[Medline]
  4. Botchkarev, V. A., N. V. Botchkareva, A. Slominski, B. Roloff, T. Luger, R. Paus. 1999. Developmentally regulated expression of {alpha}-MSH and MC-1 receptor in C57BL/6 mouse skin suggests functions beyond pigmentation. Ann. NY Acad. Sci. 885:433.[Free Full Text]
  5. Wintzen, M., B. A. Gilchrest. 1996. Proopiomelanocortin, its derived peptides, and the skin. J. Invest. Dermatol. 106:3.[Medline]
  6. Lipton, J. M., H. Zhao, T. Ichiyama, G. S. Barsh, A. Catania. 1999. Mechanisms of antiinflammatory action of {alpha}-MSH peptides: in vivo and in vitro evidence. Ann. NY Acad. Sci. 885:173.[Abstract/Free Full Text]
  7. Luger, T. A., R. S. Bhardwaj, S. Grabbe, T. Schwarz. 1996. Regulation of the immune response by epidermal cytokines and neurohormones. J. Dermatol. Sci. 13:5.[Medline]
  8. Cutuli, M., S. Cristiani, J. M. Lipton, A. Catania. 2000. Antimicrobial effects of {alpha}-MSH peptides. J. Leukocyte Biol. 67:233.[Abstract]
  9. Hartmeyer, M., T. Scholzen, E. Becher, R. S. Bhardwaj, T. Schwarz, T. A. Luger. 1997. Human dermal microvascular endothelial cells express the melanocortin receptor type 1 and produce increased levels of IL-8 upon stimulation with {alpha}-melanocyte-stimulating hormone. J. Immunol. 159:1930.[Abstract]
  10. Bhardwaj, R., E. Becher, K. Mahnke, M. Hartmeyer, T. Schwarz, T. Scholzen, T. A. Luger. 1997. Evidence for the differential expression of the functional {alpha}-melanocyte-stimulating hormone receptor MC-1 on human monocytes. J. Immunol. 158:3378.[Abstract]
  11. Chhajlani, V., R. Muceniece, J. E. Wikberg. 1996. Molecular cloning of a novel human melanocortin receptor. Biochem. Biophys. Res. Commun. 218:638.[Medline]
  12. Curry, J. L., W. Pinto, B. J. Nickoloff, A. T. Slominski. 2001. Human keratinocytes express functional {alpha}-MSH (MC1-R) receptors. In Vitro Cell. Dev. Biol. Anim. 37:234.[Medline]
  13. Slominski, A., J. Wortsman, J. E. Mazurkiewicz, L. Matsuoka, J. Dietrich, K. Lawrence, A. Gorbani, R. Paus. 1993. Detection of proopiomelanocortin-derived antigens in normal and pathologic human skin. J. Lab. Clin. Med. 122:658.[Medline]
  14. Luger, T. A., T. Scholzen, S. Grabbe. 1997. The role of {alpha}-melanocyte-stimulating hormone in cutaneous biology. J. Invest. Dermatol. Symp. Proc. 2:87.
  15. Rheins, L. A., A. L. Cotleur, R. S. Kleier, W. B. Hoppenjans, D. N. Saunder, J. J. Nordlund. 1989. {alpha}-Melanocyte stimulating hormone modulates contact hypersensitivity responsiveness in C57/BL6 mice. J. Invest. Dermatol. 93:511.[Medline]
  16. Lipton, J. M., A. Macaluso, M. E. Hiltz, A. Catania. 1991. Central administration of the peptide {alpha}-MSH inhibits inflammation in the skin. Peptides 12:795.[Medline]
  17. Grabbe, S., R. S. Bhardwaj, K. Mahnke, M. M. Simon, T. Schwarz, T. A. Luger. 1996. {alpha}-Melanocyte-stimulating hormone induces hapten-specific tolerance in mice. J. Immunol. 156:473.[Abstract]
  18. Herz, U., A. Braun, R. Ruckert, H. Renz. 1998. Various immunological phenotypes are associated with increased airway responsiveness. Clin. Exp. Allergy 28:625.[Medline]
  19. Finotto, S., G. T. De Sanctis, H. A. Lehr, U. Herz, M. Buerke, M. Schipp, B. Bartsch, R. Atreya, E. Schmitt, P. R. Galle, et al 2001. Treatment of allergic airway inflammation and hyperresponsiveness by antisense-induced local blockade of GATA-3 expression. J. Exp. Med. 193:1247.[Abstract/Free Full Text]
  20. Renz, H., H. R. Smith, J. E. Henson, B. S. Ray, C. G. Irvin, E. W. Gelfand. 1992. Aerosolized antigen exposure without adjuvant causes increased IgE production and increased airway responsiveness in the mouse. J. Allergy Clin. Immunol. 89:1127.[Medline]
  21. Braun, A., M. Lommatzsch, A. Mannsfeldt, U. Neuhaus-Steinmetz, A. Fischer, N. Schnoy, G. R. Lewin, H. Renz. 1999. Cellular sources of enhanced brain-derived neurotrophic factor production in a mouse model of allergic inflammation. Am. J. Respir. Cell Mol. Biol. 21:537.[Abstract/Free Full Text]
  22. Herz, U., B. Ahrens, A. Scheffold, R. Joachim, A. Radbruch, H. Renz. 2000. Impact of in utero Th2 immunity on T cell deviation and subsequent immediate-type hypersensitivity in the neonate. Eur. J. Immunol. 30:714.[Medline]
  23. Neuhaus-Steinmetz, U., T. Glaab, A. Daser, A. Braun, M. Lommatzsch, U. Herz, J. Kips, Y. Alarie, H. Renz. 2000. Sequential development of airway hyperresponsiveness and acute airway obstruction in a mouse model of allergic inflammation. Int. Arch. Allergy Immunol. 121:57.[Medline]
  24. Luger, T. A., T. Scholzen, T. Brzoska, E. Becher, A. Slominski, R. Paus. 1998. Cutaneous immunomodulation and coordination of skin stress responses by {alpha}-melanocyte-stimulating hormone. Ann. NY Acad. Sci. 840:381.[Abstract/Free Full Text]
  25. Cannon, J. G., J. B. Tatro, S. Reichlin, C. A. Dinarello. 1986. {alpha} Melanocyte stimulating hormone inhibits immunostimulatory and inflammatory actions of interleukin 1. J. Immunol. 137:2232.[Abstract]
  26. Hiltz, M. E., A. Catania, J. M. Lipton. 1992. {alpha}-MSH peptides inhibit acute inflammation induced in mice by rIL-1{beta}, rIL-6, rTNF-{alpha} and endogenous pyrogen but not that caused by LTB4, PAF and rIL-8. Cytokine 4:320.[Medline]
  27. Artuc, M., A. Grutzkau, T. Luger, B. M. Henz. 1999. Expression of MC1- and MC5-receptors on the human mast cell line HMC-1. Ann. NY Acad. Sci. 885:364.[Free Full Text]
  28. Adachi, S., T. Nakano, H. Vliagoftis, D. D. Metcalfe. 1999. Receptor-mediated modulation of murine mast cell function by {alpha}-melanocyte stimulating hormone. J. Immunol. 163:3363.[Abstract/Free Full Text]
  29. Grutzkau, A., B. M. Henz, L. Kirchhof, T. Luger, M. Artuc. 2000. {alpha}-Melanocyte stimulating hormone acts as a selective inducer of secretory functions in human mast cells. Biochem. Biophys. Res. Commun. 278:14.[Medline]
  30. Aebischer, I., M. R. Stampfli, A. Zurcher, S. Miescher, A. Urwyler, B. Frey, T. Luger, R. R. White, B. M. Stadler. 1994. Neuropeptides are potent modulators of human in vitro immunoglobulin E synthesis. Eur. J. Immunol. 24:1908.[Medline]
  31. Kalden, D. H., T. Scholzen, T. Brzoska, T. A. Luger. 1999. Mechanisms of the antiinflammatory effects of {alpha}-MSH: role of transcription factor NF-{kappa}B and adhesion molecule expression. Ann. NY Acad. Sci. 885:254.[Abstract/Free Full Text]
  32. Ichiyama, T., H. Zhao, A. Catania, S. Furukawa, J. M. Lipton. 1999. {alpha}-Melanocyte-stimulating hormone inhibits NF-{kappa}B activation and I{kappa}B{alpha} degradation in human glioma cells and in experimental brain inflammation. Exp. Neurol. 157:359.[Medline]
  33. Wedi, B., U. Raap, H. Lewrick, A. Kapp. 1997. Delayed eosinophil programmed cell death in vitro: a common feature of inhalant allergy and extrinsic and intrinsic atopic dermatitis. J. Allergy Clin. Immunol. 100:536.[Medline]
  34. Wardlaw, A. J.. 2001. Eosinophil trafficking in asthma. Clin. Med. 1:214.[Medline]
  35. Croft, D., P. McIntyre, A. Wibulswas, I. Kramer. 1999. Sustained elevated levels of VCAM-1 in cultured fibroblast-like synoviocytes can be achieved by TNF-{alpha} in combination with either IL-4 or IL-13 through increased mRNA stability. Am. J. Pathol. 154:1149.[Abstract/Free Full Text]
  36. Cohn, L., R. J. Homer, A. Marinov, J. Rankin, K. Bottomly. 1997. Induction of airway mucus production by T helper 2 (Th2) cells: a critical role for interleukin 4 in cell recruitment but not mucus production. J. Exp. Med. 186:1737.[Abstract/Free Full Text]
  37. Moore, K. W., R. de Waal Malefyt, R. L. Coffman, A. O’Garra. 2001. Interleukin-10 and the interleukin-10 receptor. Annu. Rev. Immunol. 19:683.[Medline]
  38. Blaser, K., C. A. Akdis, A. Faith. 1999. Differential regulation of allergen-specific antibodies in allergy and specific immunotherapy. Arb. Paul. Ehrlich. Inst. Bundesamt Sera Impfstoffe Frankf. AM 93:243.
  39. Akdis, C. A., K. Blaser. 2001. Mechanisms of interleukin-10-mediated immune suppression. Immunology 103:131.[Medline]
  40. Taylor, A., K. Namba. 2001. In vitro induction of CD25 CD4 regulatory T cells by the neuropeptide {alpha}-melanocyte stimulating hormone ({alpha}-MSH). Immunol. Cell Biol. 79:358.[Medline]
  41. Pantoux, C., A. Banz, M. Papiernik. 2002. Natural CD4 CD25+ regulatory T cells control the burst of superantigen induced cytokine production: the role of IL-10. Int. Immunol. 14:233.[Abstract/Free Full Text]
  42. Punnonen, J., R. de Waal Malefyt, P. van Vlasselaer, J. F. Gauchat, J. E. de Vries. 1993. IL-10 and viral IL-10 prevent IL-4-induced IgE synthesis by inhibiting the accessory cell function of monocytes. J. Immunol. 151:1280.[Abstract]
  43. Hogan, S. P., A. Koskinen, P. S. Foster. 1997. Interleukin-5 and eosinophils induce airway damage and bronchial hyperreactivity during allergic airway inflammation in BALB/c mice. Immunol. Cell Biol. 75:284.[Medline]
  44. Hamelmann, E., A. Oshiba, J. Loader, G. L. Larsen, G. Gleich, J. Lee, E. W. Gelfand. 1997. Antiinterleukin-5 antibody prevents airway hyperresponsiveness in a murine model of airway sensitization. Am. J. Respir. Crit. Care Med. 155:819.[Abstract]
  45. Foster, P. S., S. P. Hogan, K. I. Matthaei, I. G. Young. 1997. Interleukin-4and interleukin-5 as targets for the inhibition of eosinophilic inflammation and allergic airways hyperreactivity. Mem. Inst. Oswaldo Cruz 92:(Suppl. 2):55.
  46. Foster, P. S., S. P. Hogan, A. J. Ramsay, K. I. Matthaei, I. G. Young. 1996. Interleukin 5 deficiency abolishes eosinophilia, airways hyperreactivity, and lung damage in a mouse asthma model. J. Exp. Med. 183:195.[Abstract/Free Full Text]
  47. Lee, J. J., M. P. McGarry, S. C. Farmer, K. L. Denzler, K. A. Larson, P. E. Carrigan, I. E. Brenneise, M. A. Horton, A. Haczku, E. W. Gelfand, et al 1997. Interleukin-5 expression in the lung epithelium of transgenic mice leads to pulmonary changes pathognomonic of asthma. J. Exp. Med. 185:2143.[Abstract/Free Full Text]
  48. Wilder, J. A., D. D. Collie, B. S. Wilson, D. E. Bice, C. R. Lyons, M. F. Lipscomb. 1999. Dissociation of airway hyperresponsiveness from immunoglobulin E and airway eosinophilia in a murine model of allergic asthma. Am. J. Respir. Cell Mol. Biol. 20:1326.[Abstract/Free Full Text]
  49. Kips, J. C., G. G. Brusselle, G. F. Joos, R. A. Peleman, R. R. Devos, J. H. Tavernier, R. A. Pauwels. 1995. Importance of interleukin-4 and interleukin-12 in allergen-induced airway changes in mice. Int. Arch. Allergy Immunol. 107:115.[Medline]
  50. Corry, D. B., H. G. Folkesson, M. L. Warnock, D. J. Erle, M. A. Matthay, J. P. Wiener Kronish, R. M. Locksley. 1996. Interleukin 4, but not interleukin 5 or eosinophils, is required in a murine model of acute airway hyperreactivity. J. Exp. Med. 183:109.[Abstract/Free Full Text]
  51. Leckie, M. J., A. ten Brinke, J. Khan, Z. Diamant, B. J. O’Connor, C. M. Walls, A. K. Mathur, H. C. Cowley, K. F. Chung, R. Djukanovic, et al 2000. Effects of an interleukin-5 blocking monoclonal antibody on eosinophils, airway hyper-responsiveness, and the late asthmatic response. Lancet 356:2144.[Medline]
  52. Akbari, O., R. H. DeKruyff, D. T. Umetsu. 2001. Pulmonary dendritic cells producing IL-10 mediate tolerance induced by respiratory exposure to antigen. Nat. Immun. 2:725.[Medline]
  53. Tsitoura, D. C., R. H. DeKruyff, J. R. Lamb, D. T. Umetsu. 1999. Intranasal exposure to protein antigen induces immunological tolerance mediated by functionally disabled CD4+ T cells. J. Immunol. 163:2592.[Abstract/Free Full Text]



This article has been cited by other articles:


Home page
 Endocr. Rev.Home page
T. Brzoska, T. A. Luger, C. Maaser, C. Abels, and M. Bohm
{alpha}-Melanocyte-Stimulating Hormone and Related Tripeptides: Biochemistry, Antiinflammatory and Protective Effects in Vitro and in Vivo, and Future Perspectives for the Treatment of Immune-Mediated Inflammatory Diseases
Endocr. Rev., August 1, 2008; 29(5): 581 - 602.
[Abstract] [Full Text] [PDF]

Home page
 Ann Rheum DisHome page
T. A Luger and T. Brzoska
{alpha}-MSH related peptides: a new class of anti-inflammatory and immunomodulating drugs
Ann Rheum Dis, November 1, 2007; 66(suppl_3): iii52 - iii55.
[Abstract] [Full Text] [PDF]

Home page
 Mol. Pharmacol.Home page
S. J. Getting, C. W. Lam, G. Leoni, F. N. E. Gavins, P. Grieco, and M. Perretti
[D-Trp8]-{gamma}-Melanocyte-Stimulating Hormone Exhibits Anti-Inflammatory Efficacy in Mice Bearing a Nonfunctional MC1R (Recessive Yellow e/e Mouse)
Mol. Pharmacol., December 1, 2006; 70(6): 1850 - 1855.
[Abstract] [Full Text] [PDF]

Home page
 FASEB J.Home page
S. J. Getting, C. W. Lam, A. S. Chen, P. Grieco, and M. Perretti
Melanocortin 3 receptors control crystal-induced inflammation
FASEB J, November 1, 2006; 20(13): 2234 - 2241.
[Abstract] [Full Text] [PDF]

Home page
 J. Leukoc. Biol.Home page
L. Kang, K. W. McIntyre, K. M. Gillooly, Y. Yang, J. Haycock, S. Roberts, A. Khanna, T. F. Herpin, G. Yu, X. Wu, et al.
A selective small molecule agonist of the melanocortin-1 receptor inhibits lipopolysaccharide-induced cytokine accumulation and leukocyte infiltration in mice
J. Leukoc. Biol., October 1, 2006; 80(4): 897 - 904.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
C. W. Lam, S. J. Getting, and M. Perretti
In Vitro and In Vivo Induction of Heme Oxygenase 1 in Mouse Macrophages following Melanocortin Receptor Activation
J. Immunol., February 15, 2005; 174(4): 2297 - 2304.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Raap, U.
Right arrow Articles by Renz, H.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Raap, U.
Right arrow Articles by Renz, H.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS