International Immunology, Vol. 24, No. 2, pp. 107–116
Advance Access publication 9 January 2012
ª The Japanese Society for Immunology. 2012. All rights reserved.
For permissions, please e-mail: firstname.lastname@example.org
Histamine modulates multiple functional activities of
monocyte-derived dendritic cell subsets via
histamine receptor 2
Tu ¨nde Simon1,2, Pe ´ter Gogola ´k1, Katalin Kis-To ´th1, Ivett Jelinek2, Vale ´ria La ´szlo ´2and
E´va Rajnavo ¨lgyi1
1Department of Immunology, Medical and Health Science Center, University of Debrecen, Debrecen H-4032, Hungary
2Department of Genetics, Cell- and Immunobiology, Semmelweis University, Budapest H-1089, Hungary
Correspondence to: E´. Rajnavo ¨lgyi, Institute of Immunology, Medical and Health Science Centre, University of Debrecen, Debrecen H-4032,
Nagyerdei Boulevard 98, Hungary; E-mail: email@example.com
Received 5 August 2011, accepted 22 November 2011
Expression of CD1a proteins in human monocyte-derived dendritic cells (DCs) specifies functionally
distinct subsets with different inflammatory properties. Histamine is recognized as an inflammatory
mediator released by various cell types including DCs. The diverse biological effects of histamine are
mediated by G-protein-coupled histamine receptors (HRs), which are able to modulate the functional
activities of DC subsets. The goal of the present study was to compare the expression and activity of
HRs in the CD1a2and CD1a1monocyte-derived DC subsets and to test the effects of histamine on the
differentiation, activation and functional activities of these subsets. We show that H2R is present at
high levels in both DC subsets, whereas H1R and H4R are expressed in a subset-specific manner.
Histamine shifts DC differentiation to the development of CD1a2DCs and modulates DC activation
through its inhibitory effect on CD1a1DC differentiation. Histamine-induced reduction of CD1a1DCs
is associated with increased secretion of IL-6 and IL-10, up-regulation of a typical combination of
chemokines, expression C5aR1 by the CD1a2DC subset and enhanced migration of both activated
DC subsets supported by the production of MMP-9 and MMP-12 enzymes. All these effects were
shown to be mediated in a H2R-specific manner as revealed by the specific antagonist of the receptor.
As H2R is expressed at high levels in both DC subsets, we propose that it may dominate the
regulation of multiple DC functions. In contrast, H1R and H4R with opposing subset-related
expression may have a regulatory or fine-tuning role in histamine-induced functional activities.
Keywords: CD1a molecule, cytokine, DC activation, migration
Dendritic cells (DCs) involve bone marrow-derived cells with
distinct life cycles, migratory properties and antigen present-
ing functions (1, 2). Being highly potent professional antigen-
presenting cells, DCs possess the unique potential to acti-
vate naive T cells, instruct Th-cell differentiation and thus
play a critical role in the induction and outcome of adaptive
immune responses (3, 4). CD34+hematopoietic progenitors
are widely studied precursors of DCs that give rise to
several classical DC subsets when cultured in the presence
of appropriate cytokine combinations (5, 6). DCs can also
differentiate from CD14+blood monocytes in peripheral tis-
sues and lymphoid organs or under in vitro conditions
(7, 8). Recent studies identified several subsets of conven-
tional circulating DCs with unique phenotypic and functional
properties both in mouse and human (9–12) demonstrating
the functional specialization of these cells.
Originally, in vitro generated human monocyte-derived
DCs have been considered as a homogeneous and func-
tionally competent cell population appropriate for clinical util-
ity (13). However, the expression pattern of human CD1
membrane proteins has been shown to specify distinct hu-
man DC subtypes independent on their activation state
(14, 15). We have previously shown that human monocyte-
derived DCs generated in vitro involve both CD1a?and
CD1a+cells, their ratio varies among individuals and is con-
trolled by the lipid and lipoprotein environment of DCs via
the ligand-activated nuclear hormone receptor peroxisome
proliferator-activated receptor gamma (16). It has also been
by guest on November 24, 2015
demonstrated that CD1a?and CD1a+DCs exhibit distinct
functional characteristics being the CD1a+subset more
inflammatory as compared with its CD1a?counterpart (8).
One of the major functions of DCs is the continuous sam-
pling of their environment by receptors expressed in the cell
or vesicular membranes or in the cytosol and transfers this
information to T lymphocytes (17). Some of these receptors
mediate internalization of soluble material or particles; others
are involved in signal transduction, cell migration or cell-to-
cell communication (18). As a result, the differentiation and
activation of DCs occur in an environment-dependent man-
ner and result in considerable functional flexibility. Depend-
ing on the actual tissue environment, phenotypically defined
monocyte subsets develop to monocyte-derived DCs, which
also represent a functionally heterogeneous cell population
(11). Under in vivo conditions, DC differentiation and activa-
tion depend on the modulatory effects of metabolites, cyto-
kines,chemokines and other
a multifunctional small-sized biogenic amine that regulates
cellular responses and plays diverse roles in physiological
and pathological processes (19, 20). Histamine was also
shown to influence DC differentiation, as both the intracellu-
lar histamine content and the expression of the histidine
decarboxylase enzyme were found to be increased in the
course of in vitro cytokine-induced monocyte-derived DC dif-
ferentiation in parallel with the induction of the co-stimulatory
molecules CD40, CD80 and CD86 (21).
A rapidly growing body of evidence highlighted that hista-
mine, a small biogenic amine, is involved in the regulation
of DC functions and consequently T cell-mediated inflamma-
tion (22). Histamine is recognized as an inflammatory media-
tor released by mast cells, basophils and other cell types,
including DCs themselves (21). The diverse biological
effects of histamine are mediated via four distinct G-protein-
coupled receptors, namely H1, H2, H3 and the lastly discov-
ered H4 receptors (23). It is well documented that H1, H2
and H4 receptor mRNAs and proteins are expressed in hu-
man DCs (24–27) but the presence of H3R is still controver-
sial (28, 29). Interestingly, neither H1R nor H2R could be
detected on the surface of Langerhans cells (LCs) isolated
from human epidermis or differentiated in vitro (30). In con-
trast to these results, the expression of the recently discov-
ered H4R was detected at both mRNA and protein levels on
primary LCs of both murine and human skin samples and
also on in vitro generated monocyte-derived LCs (31, 32).
The expression pattern and functionality of histamine recep-
tors (HRs) in various DC subpopulations, however, are
The majority of the experimental data support that hista-
mine induces an altered cytokine expression profile in DCs
and favor Th2 polarization. Histamine was also shown to in-
hibit the secretion of the pro-inflammatory cytokines IL-1b
and IL-6 as well as the Th1 polarizing cytokine IL-12p70 (24).
These effects could be antagonized by specific HR antago-
nists (24, 25, 33), whereas the production of Th2 cytokines
and LPS-induced IL-8 and IL-10 production was increased in
histamine-treated human monocyte-derived DCs (28, 33). The
reduced production of the Th1 polarizing cytokine IL-12p70
was mediated not only by H2R but also by H4R (29). Several
independent research groups also established that histamine
(26, 29) or histamine agonists (28, 29) act as strong chemo-
taxins for resting but not for activated DCs. The involvement
of H1R (28), H2R (34, 35), H3R (26, 28) and H4R (36, 37) has
been shown to exert modulatory effects on DC migration.
The major goal of the present study was to compare the
expression of HRs in the previously identified and character-
ized CD1a?and CD1a+human monocyte-derived DC sub-
populations and to test the modulatory effects of histamine
on the differentiation, activation, cytokine and chemokine se-
cretion and migration of these DC subsets. Specific inhibi-
tors of the H1, H2 and H4 receptors were used to validate
the functionally relevant effects on DC activities and to iden-
tify the HRs responsible for the DC-associated modulatory
effects of histamine.
Monocyte separation and differentiation of DCs
Buffy coats were obtained with the written permission of the
National Blood Transfusion Centre, Budapest Hungary. Pe-
ripheral blood mononuclear cells (PBMC) were obtained by
Uppsala, Sweden). Monocytes were isolated from PBMC by
microbeads (Miltenyi Biotech, Bergisch Gladbach, Germany).
CD14highmonocytes (>97%) were cultured in 6-well plates at
a density of 2 3 106cells ml?1in serum-free AIMV medium
(Life technologies, Carlsbad, CA, USA) supplemented with
antibiotics, 100 ng ml?1IL-4 and 80 ng ml?1GM-CSF. At day
2, the same amount of cytokines was added and the cells
were cultured for another 3 days. Resting DCs were har-
vested on day 5 or were activated by LPS (100 ng ml?1,
Sigma–Aldrich, Schnelldorf, Germany) and IFNc (10 ng ml?1,
PeproTech Inc., Rocky Hill, NJ, USA) and harvested on day
6. When indicated, histamine (10?5M, Sigma–Aldrich) or his-
tamine combined with specific HR inhibitors (10?5M) (38,
39) were added at days 0 and 2 together with the differentiat-
ing cytokines and when activated, with LPS+IFNc (100 ng ml?1
and 10 ng ml?1, respectively). The following HR antagonists
were used: pyrilamine (H1R antagonist, Sigma–Aldrich),
famotidine (H2R antagonist Sigma–Aldrich), JNJ7777120 and
JNJ10191584 (H4R antagonists, Sigma–Aldrich and Tocris
Bioscience, Ellisville, MO, USA).
Phenotyping of resting and activated DCs was performed by
flow cytometry using anti-CD83-FITC and anti-CD1a-PE anti-
bodies (Beckman Coulter, Hialeah, FL, USA) and isotype-
matched control antibody (BD Pharmingen, San Diego, CA,
USA). For measuring H2R expression by DC subsets, the
washed cells were labeled with 0.1 lg rabbit anti-human
H2R (Alpha Diagnostic International Inc., San Antonio, TX,
USA) in the presence of heat-inactivated normal mouse se-
rum as a blocking reagent. After 30 min incubation on ice,
the cells were washed and labeled with 1 lg (0.5 ll)
Alexa488-conjugated goat anti-rabbit (Fab’)2fragments (Invi-
trogen, Eugene, OR, USA) together with 5 lPE-labeled
mouse anti-human CD1a antibody (BD Biosciences, Franklin
Lakes, NJ, USA). Fluorescence intensities were measured
108 H2R modulates monocyte-derived DC differentiation
by guest on November 24, 2015
29 Gutzmer, R., Diestel, C., Mommert, S. et al. 2005. Histamine
H4 receptor stimulation suppresses IL-12p.70 production and
mediates chemotaxis in human monocyte-derived dendritic cells.
J. Immunol. 174:5224.
30 Ohtani, T., Aiba, S., Mizuashi, M., Mollah, Z. U., Nakagawa, S. and
Tagami, H. 2003. H1 and H2 histamine receptors are absent on
Langerhans cells and present on dermal dendritic cells. J. Invest.
31 Dijkstra, D., Stark, H., Chazot, P. L. et al. 2008. Human
inflammatory dendritic epidermal cells express a functional
histamine H4 receptor. J. Invest. Dermatol. 128:1696.
32 Gschwandtner, M., Rossbach, K., Dijkstra, D. et al. 2010. Murine
and human Langerhans cells express a functional histamine H4
receptor: modulation of cell migration and function. Allergy
33 Mazzoni, A., Young, H. A., Spitzer, J. H., Visintin, A. and Segal, D.
M. 2001. Histamine regulates cytokine production in maturing
dendritic cells, resulting in altered T cell polarization. J. Clin.
34 Jawdat, D. M., Albert, E. J., Rowden, G., Haidl, I. D. and Marshall,
J. S. 2004. IgE-mediated mast cell activation induces Langerhans
cell migration in vivo. J. Immunol. 173:5275.
35 Dawicki, W., Jawdat, D. W., Xu, N. and Marshall, J. S. 2010. Mast
cells, histamine, and IL-6 regulate the selective influx of dendritic
cell subsets into an inflamed lymph node. J. Immunol. 184:2116.
36 Baumer, W., Wendorff, S., Gutzmer, R. et al. 2008. Histamine
H4 receptors modulate dendritic cell migration through skin–
immunomodulatory role of histamine. Allergy 63:1387.
37 Cowden, J. M., Zhang, M., Dunford, P. J. and Thurmond, R. L.
2010. The histamine H4 receptor mediates inflammation and
pruritus in Th2-dependent dermal inflammation. J. Invest. Derma-
38 Dijkstra, D., Leurs, R., Chazot, P. et al. 2007. Histamine down-
regulates monocyte CCL2 production through the histamine H4
receptor. J. Allergy Clin. Immunol. 120:300.
39 Dunford, P. J., O’Donnell, N., Riley, J. P., Williams, K. N., Karlsson,
L. and Thurmond, R. L. 2006. The histamine H4 receptor mediates
allergic airway inflammation by regulating the activation of CD4+ T
cells. J. Immunol. 176:7062.
40 de Saint-Vis, B., Fugier-Vivier, I., Massacrier, C. et al. 1998. The
cytokine profile expressed by human dendritic cells is dependent
on cell subtype and mode of activation. J. Immunol. 160:1666.
41 Tetlow, L. C. and Woolley, D. E. 2002. Histamine stimulates matrix
metalloproteinase-3 and -13 production by human articular
chondrocytes in vitro. Ann. Rheum. Dis. 61:737.
42 Del Prete, A., Locati, M., Otero, K. et al. 2006. Migration of
dendritic cells across blood and lymphatic endothelial barriers.
Thromb. Haemost. 95:22.
43 Randolph, G. J., Sanchez-Schmitz, G. and Angeli, V. 2005.
Factors and signals that govern the migration of dendritic cells via
lymphatics: recent advances. Springer Semin. Immunopathol.
44 Schiefner, A. and Wilson, I. A. 2009. Presentation of lipid antigens
by CD1 glycoproteins. Curr. Pharm. Des. 15:3311.
45 Katoh, N., Soga, F., Nara, T., Masuda, K. and Kishimoto, S. 2005.
Histamine induces the generation of monocyte-derived dendritic
cells that express CD14 but not CD1a. J. Invest. Dermatol.
46 Caron, G., Delneste, Y., Roelandts, E. et al. 2001. Histamine
polarizes human dendritic cells into Th2 cell-promoting effector
dendritic cells. J. Immunol. 167:3682.
47 Chang, C. C., Wright, A. and Punnonen, J. 2000. Monocyte-
derived CD1a+ and CD1a- dendritic cell subsets differ in their
cytokine production profiles, susceptibilities to transfection, and
capacities to direct Th cell differentiation. J. Immunol. 165:3584.
48 Simon, T., Laszlo, V., Lang, O., Buzas, E. and Falus, A. 2011.
Histamine regulates relevant murine dendritic cell functions via H4
receptor. Front. Biosci. (Elite Ed.) 3:1414.
49 Osman, M., Tortorella, M., Londei, M. and Quaratino, S. 2002.
Expression of matrix metalloproteinases and tissue inhibitors of
metalloproteinases define the migratory characteristics of human
monocyte-derived dendritic cells. Immunology 105:73.
50 Bracke, K., Cataldo, D., Maes, T. et al. 2005. Matrix metalloproteinase-
12 and cathepsin D expression in pulmonary macrophages and
dendritic cells of cigarette smoke-exposed mice. Int. Arch. Allergy
51 Schakel, K., Mayer, E., Federle, C., Schmitz, M., Riethmuller, G.
and Rieber, E. P. 1998. A novel dendritic cell population in human
blood: one-step immunomagnetic isolation by a specific mAb (M-
DC8) and in vitro priming of cytotoxic T lymphocytes. Eur. J.
52 Li, K., Fazekasova, H., Wang, N. et al. 2011. Expression of
complement components, receptors and regulators by human
dendritic cells. Mol. Immunol. 48:1121.
53 Weaver, D. J. Jr, Reis, E. S., Pandey, M. K. et al. 2010. C5a
receptor-deficient dendritic cells promote induction of Treg and
Th17 cells. Eur. J. Immunol. 40:710.
54 Sallusto, F., Schaerli, P., Loetscher, P. et al. 1998. Rapid and
coordinated switch in chemokine receptor expression during
dendritic cell maturation. Eur. J. Immunol. 28:2760.
55 Kabashima, K., Shiraishi, N., Sugita, K. et al. 2007. CXCL12-
CXCR4 engagement is required for migration of cutaneous
dendritic cells. Am. J. Pathol. 171:1249.
56 Chong, S. Z., Wong, K. L., Lin, G. et al. 2011. Human CD8 Tcells
drive Th1 responses through the differentiation of TNF/iNOS-
producing dendritic cells. Eur. J. Immunol. 41:1639.
57 Rogers, N. M., Matthews, T. J., Kausman, J. Y., Kitching, A. R. and
Coates, P. T. 2009. Review article: kidney dendritic cells: their role
in homeostasis, inflammation and transplantation. Nephrology
116H2R modulates monocyte-derived DC differentiation
by guest on November 24, 2015