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Immunity
Review
Alternative Activation of Macrophages:
Mechanism and Functions
Siamon Gordon1,* and Fernando O. Martinez1,*
1Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK
*Correspondence: [email protected] (S.G.), [email protected] (F.O.M.)
DOI 10.1016/j.immuni.2010.05.007
The concept of an alternative pathway of macrophage activation has stimulated interest in its definition,
mechanism, and functional significance in homeostasis and disease. We assess recent research in this field,
argue for a restricted definition, and explore pathways by which the T helper 2 (Th2) cell cytokines interleukin4 (IL-4) and IL-13 mediate their effects on macrophage cell biology, their biosynthesis, and responses to
a normal and pathological microenvironment. The stage is now set to gain deeper insights into the role of
alternatively activated macrophages in immunobiology.
Introduction
The concept of an alternative pathway of macrophage activation
induced by the T helper 2 (Th2) cytokines interleukin-4 (IL-4) and
IL-13, distinct from interferon-g (IFN-g)-mediated classical activation, has gained considerable ground over the past decade.
Analysis of marker molecules in the mouse has revealed a stereotypic signature of expressed genes induced by IL-4 and IL-13,
contrasting with characteristic changes induced by IFN-g, which
mediates Th1 cell-type activation of macrophages. Phenotypes
associated with alternative activation of macrophages (AAMs)
and classical activation of macrophages (CAMs) can be distinguished from the direct effects of microbial stimuli, such as lipopolysaccharide, which induce innate macrophage activation,
often through Toll-like receptors (TLRs). Innate stimuli are able
to synergize with CAMs, to achieve full expression of macrophage effector pathways.
Investigators have implicated AAMs in a range of physiologic
and pathological processes, including homeostasis, inflammation, repair, metabolic functions, and malignancy. This is in addition to their established role in immunity, hypersensitivity, allergy,
parasitic infections, and their sequelae, such as fibrosis. Recent
genome-wide analysis, although broadly supporting earlier studies with limited numbers of markers, has underscored the
heterogeneity and plasticity of macrophage gene expression,
adding to the confusing nomenclature of monocyte and tissue
macrophage subpopulations (Geissmann et al., 2010). In this
review, we assess current knowledge, point to the need to
restrict the concept of AAM, and consider newer findings
regarding mechanisms and therapeutic potential.
Our own interest in IL-4 modulation of the macrophage phenotype arose in the late 1980s from experiments with human blood
monocyte-derived macrophages and mouse peritoneal macrophages. At the time, the role of this cytokine was believed to
be mainly anti-inflammatory, resulting from its ability to suppress
TNF and IL-6 production in macrophages. Whereas macrophage
mannose receptor (MRC1) expression and its endocytic function
were selectively downregulated by IFN-g (Mokoena and Gordon,
1985), it was upregulated by IL-4 (Stein et al., 1992) and IL-13
(Doyle et al., 1994) and depended in vitro and in vivo on
IL4Ra1, the common IL-4 and IL-13 receptor alpha chain (Linehan et al., 2003). Others showed a critical role for the transcrip-
tion factor STAT6 in their signaling (Heller et al., 2008). These
and other experiments showed that these Th2 cytokines were
not simply inhibitory, but subtle modulators of macrophage functions. We proposed that IL-4 induced an ‘‘alternative’’ form of
activation in macrophages (Stein et al., 1992).
Subsequently, other workers introduced the term M1 and M2
to parallel macrophage activation with T helper cell polarization,
subdivided M2 classes into subgroups, and in some cases,
termed all nonclassical activation phenotypes as ‘‘alternative’’
(discussed in Gratchev et al., 2001; Mantovani et al., 2008;
Mosser and Edwards, 2008). We have argued that too loose
a definition obscures the study of mechanism and discovery of
potential therapeutic agents. Even a restricted definition of IL-4
and IL-13 dependence needs to take into account the complex
effects of cell differentiation, interactions with other receptors,
and signaling pathways, and the role of tissue microenvironment
in modulating the macrophage phenotype. Monocyte and macrophage heterogeneity and nomenclature are discussed below
and elsewhere (Gabrilovich and Nagaraj, 2009; Geissmann
et al., 2010).
Analytic Approaches
There are two distinct, complementary approaches to the
study of AAMs. The first, ‘‘forward analysis,’’ flows from receptor-dependent recognition, through signal transduction, gene
expression, and protein production. The second, ‘‘reverse analysis,’’ deduces from phenotypic markers the mechanism of initiation. Studies with isolated macrophages and in situ analysis
in wild-type and genetically modified mice made it possible to
combine these approaches. Forward analysis utilizes cytokine,
receptor, or signaling-deficient animals to define an AAM phenotype, including Arginase 1 (ARG1), Ym1 and Ym2, Fizz1, MRC1,
and selected chemokines (Loke et al., 2002). Signature markers,
in combination, have then been used to infer an AAM pathway
and mechanism (reverse analysis).
There are obvious caveats to the use of reverse analysis alone,
as required in human studies in situ: substantial species differences exist in available markers, and established markers such
as Arg1 can be induced by other pathways (El Kasmi et al.,
2008). Even clusters of multiple signature markers can be independent of the IL-4 and IL-13 receptor pathway (Daley et al.,
Immunity 32, May 28, 2010 ª2010 Elsevier Inc. 593
Immunity
Review
2010; Stout, 2010). We lack markers for human AAM, but
these should become available with current microarray and
proteomic studies. Combined forward and reverse analysis
in the mouse will validate the link between recognition and
response. With humans, our dilemma mimics the problem of
diagnosis in clinical medicine, where a cluster of symptoms and
signs cannot be assigned to a particular cause without supportive evidence.
Toward an Integrated Approach
Before turning to the pathways and functions of AAM, we
summarize general features of monocyte and macrophage
heterogeneity (Gordon, 2008). Macrophages arise from hemopoietic progenitors (Geissmann et al., 2010), which differentiate,
directly or via circulating monocytes, into subpopulations of
tissue macrophages and closely related myeloid dendritic cells
(Steinman and Idoyaga, 2010), the latter specialized to present
antigens to naive T helper lymphocytes. Osteoclasts arise from
monocyte precursors to form multinucleated giant cells, implicated in resorption of bone. Monocyte and macrophages share
properties with granulocytes, especially neutrophils, and to
lesser extent with B lymphocytes. Monocyte and macrophages
migrate through several body compartments, including bone
marrow, blood, lymphoid, and all nonhematopoietic tissues.
Resident macrophages are present in organs constitutively, in
the absence of overt inflammation, and perform trophic as
well as homeostatic roles in the removal of apoptotic cells,
serving as sentinels of injury and infection. Tissue macrophages
can replicate locally, but are terminally differentiated, turning
over at different rates, depending on the stimulus and tissue
environment.
Macrophages differ morphologically and phenotypically in
organs such as liver, spleen, lung, gut, and brain, interacting
with matrix and other cell types. They are active in biosynthesis
and express a wide range of receptors, recognizing foreign as
well as normal and abnormal cells and host-derived products.
Through constitutive and induced endocytosis, phagocytosis,
and secretion of various products, including cytokines, growth
factors, and metabolites, they perform both trophic and toxic
functions, serving as a widely dispersed mononuclear phagocyte system during development and throughout adult life.
They contribute to tissue remodeling, host defense in innate
and adaptive immunity, and many disease processes.
Monocytes are the main source of newly recruited tissue
macrophages in infection, granulomata, atherosclerosis, Alzheimer’s disease, and tumors. Differentiation and activation of
macrophages depend on specific growth and differentiation
factors, their receptors, signaling pathways, and transcription
factors. Macrophages migrate to local sites of injury and infection, contributing to acute and chronic inflammation, locally
and systemically. In addition they acquire enhanced cytotoxic,
antimicrobial, and inhibitory activities, initiate repair, and resolve
inflammation.
Macrophages express opsonic and a range of nonopsonic
receptors, TLRs, RIG-I-like and Nod-like sensing receptors, on
their surface and in vacuolar and cytosolic compartments. Other
receptors include families of regulatory molecules, lectins, and
scavenger receptors. Their secretory repertoire includes antibacterial and proteolytic enzymes, chemokines, and anti-inflam594 Immunity 32, May 28, 2010 ª2010 Elsevier Inc.
matory cytokines, such as IL-10 and TGF-b. Proinflammatory
cytokines include IL-1b, TNF, and IL-6. Through potent membrane protein assemblies they generate reactive oxygen, nitrogen, and arachidonate metabolites.
The study of chromatin structure and epigenetic control of
gene expression in macrophages is in its infancy. Given the
range of receptors and potential ligands, it may be invidious to
single out IFN-g, IL-4, and IL-13 receptor pathways as central
regulators of macrophage activation. However, given their often
polarized production in the immune response, these contrasting
states of activation provide an important conceptual framework.
Other receptors participate in forming signaling platforms and
complexes, e.g., G protein-coupled receptors and integrins.
Crosstalk at all levels impinges on altered transcription. It is
therefore possible to conceive an infinite number of potential
phenotypes; however, the conserved, stereotypic macrophagespecific signature for IFN-g versus IL-4- and IL-13-dependent
activation provides useful insights into physiologic and pathologic mechanisms.
Figure 1 presents a schematic model of macrophage activation. The first stage, differentiation, depends on growth factors
such as GM-CSF or M-CSF; the second, priming by IFN-g
(CAMs) or IL-4 and IL-13 (AAMs); and the third, a localizing stimulus delivered by a TLR or analogous receptor. This additional
stimulus promotes induction of a full complement of classical
or alternative activation functions. In the case of AAMs, this
may arise in a Th2 cell-biased environment, e.g., chronic parasitic infection, followed by a secondary microbial challenge.
Counteracting negative controls contribute to a fourth stage of
resolution and repair. Once established, some properties of full
alternative activation can be reversible (plasticity) whereas
others may be irreversible (Stout et al., 2005).
Cytokines in AAM Induction
IL-4 and IL-13 are the prototypical direct inducers of AAMs, but
other cytokines such as IL-33 and IL-25 amplify AAM induction
indirectly, through Th2 cells (for review on CD4+ T cell heterogeneity, see O’Shea and Paul, 2010). IL-4 and IL-13 are produced
by various innate cells, including non-B non-T cells (Moro et al.,
2009; Neill et al., 2010), mast cells, basophils, eosinophils, NKT
cells, and even macrophages themselves. Apart from activated
Th2 cells and B cells, epithelia and tumor cells are potential sources in tissues. The availability of reporter mice has contributed to
the analysis of IL-4 production in vivo, during allergic inflammation and infection (Mohrs et al., 2005).
The role of IL-5, another Th2 cell cytokine that binds to distinct
receptors on macrophages, is unclear. IL-10, produced by CD4+
Th2 cells, macrophages, and B cells as well as tumor cells, is
a potent downregulator of macrophage gene expression, modulating IL-4 and IL-13 and IFN-g actions. IL-21 enhances AAMs by
driving IL-4R expression (Pesce et al., 2006). IL-33, a Th2 cell
product, binds to ST2L, a subunit of the IL-33 receptor, to
amplify Th2 cytokine production and AAM induction (Kurowska-Stolarska et al., 2009).
IL-25, a member of the IL-17 family also produced by Th2
cells, induces production of multipotent hemopoietic progenitors in bone marrow and at sites of extramedullary hemopoiesis,
giving rise to a range of myeloid cells, including monocytes and
macrophages (Saenz et al., 2010). IL-25 activates antigenspecific responses in mucosal tissues, such as the colon. In
Immunity
Review
RESOLUTION
IL-10, TGF-β, lipoxins
ACTIVATION
TLR stimuli
PRIMING
IFN-γ<-> IL-4
DIFFERENTIATION
M-CSF <-> GM-CSF
Figure 1. Paradigm of Macrophage Activation
Macrophages undergo activation to fulfill specific functional roles during
inflammation and its resolution. The phenotype is imparted by the complex
mixture of cytokines, metabolites, plasma proteins, and microbial ligands
present in the inflammatory milieu. The development into mature and fully activated macrophages can be artificially divided into successive stages. In this
scheme we have placed these stimuli in a hierarchical order.
A first phase is envisaged in which the recruited monocytes mature into
macrophages. In vitro experiments suggest that the balance of MCSF,
GMCSF, retinoic acids, and lipoproteins determines substantial differences
in the phenotype of the mature macrophage. During continuous recruitment,
monocytes are exposed to varying concentrations of mediators, inducing
a second phase of priming by cytokines. The priming term derives from the
fact that these stimuli per se are not very strong, but influence the inflammatory
potential of macrophages and their response to other stimuli. During the third
phase of activation, macrophages reach a mature functional phenotype in
response to microbial and opsonic stimuli such as antibody complexes.
If the macrophage survives its inflammatory task, it should undergo the final
phase commonly referred as to deactivation. We have chosen the term resolution, because while the macrophage proinflammatory potential is deactivated, it also undergoes functional changes that allow it to clear debris and
express general repair functions. IL-10, TGF-b, and a multitude of anti-inflammatory mediators such as nucleotides, lipoxins, and glucocorticoids are
included in this category.
principle, all cytokines that regulate Th1 and Th2 cell responses
can affect AAMs indirectly.
The IL-4 and IL-13 Receptors, Signaling, and Function
Although IL-4 and IL-13 have many overlapping activities on
macrophages, they have distinct functions. Receptors for IL-4
are mainly expressed on hematopoietic cells, whereas IL-13
receptors are also present on nonhematopoietic cells (Figure 2).
IL-4 and IL-13 receptors share a common alpha chain, but there
are specific IL-13Ra1 and IL-13Ra2 subunits (Kawashima et al.,
2006). The IL-4Ra1 can signal through the common gamma
chain (Type I IL-4R) or partner IL-13Ra1 (Type II IL-4R). The
IL-13Ra2 chain, thought to form a decoy receptor with extraordinary affinity for IL-13 (Lupardus et al., 2010), but with little
signaling capacity, has been implicated in STAT6-independent
fibrogenesis and tumor-host interactions (Fichtner-Feigl et al.,
2006, 2008b). There are therefore three distinct subtypes of
AAMs, depending on specific ligand-receptor interactions, and
their interplay is largely unexplored. The signaling pathways activated by the Type I and II receptors involve STAT6, although
other STATs can participate (Bhattacharjee et al., 2006).
STAT6 plays a central role in gene regulation by IL-4 and IL-13,
including the differentiation of Th2 cells, production of IgE, chemokines, and mucus at sites of allergic inflammation.
The differential effects of IL-4 and IL-13 on different macrophage populations from mice and humans have been examined
by correlating the magnitude of response with the expression of
each receptor (Junttila et al., 2008). Early, low amounts of IL-4
promote a Th2 cell type response; as the stimulus persists,
IL-13 becomes sufficient to induce responses in both hemopoietic and nonhemopoietic cells, making it the dominant Th2
effector cell cytokine.
Type I and Type II receptor complexes have also been
compared in mouse bone marrow-cultured macrophages and
human monocytoid cells (Heller et al., 2008). IL-4 action via the
Type I receptor was more potent with subtle differences in
signaling and target gene activation, which bear on the contribution of each cytokine to pathophysiology, e.g., in the airway.
Type I IL-4R selectively activated IRS-2, which interacts with
the adaptor Grb-2 and with PI-3 kinase, thereby initiating
multiple signaling pathways. IL-4 more readily induced a subset
of commonly used mouse AAM signature genes (Arg1, Fizz1,
Ym1), whereas Mrc1 mRNA was induced comparably by IL-4
and IL-13. The expression of each receptor polypeptide influenced its affinity for each cytokine.
IL-13Ra1-deficient cells were used to define distinct requirements for IL-13 and IL-4 in AAM induction by parasitic infection
(Ramalingam et al., 2008). Whereas IL-4 could stimulate both
Type I and Type II receptor signaling, the Type II receptor was
essential for protection against schistosomiasis and resistance
to Nippostrongylus braziliensis and its expulsion from the gut.
Allergen-induced airway hyperreactivity, mucus secretion, and
fibrosis were IL-13Ra1 dependent, whereas this receptor was
redundant for AAM and fibroblast activation. Expression of a
subset of genes, however, was reduced in the IL-13Ra1-deficient mice, including IL-13Ra2, Muc5a, Ccl11, and Ccl3. Surprisingly, the Th2 cell response was exacerbated, possibly because
of lack of feedback inhibition. These results show that IL-13 is the
more pathogenic cytokine, while consistent with the notion that
IL-4 contributes most to expansion of a Th2 cell response. The
role of epithelial and fibroblast receptors in vivo requires further
study.
IL-13 induced phosphorylation of IL-13Ra1 and IL-4Ra1, but
not of the IL-13Ra2 or IL-2Rgc (Bhattacharjee et al., 2006; Roy
et al., 2002). Phosphorylated Jak2 associated with IL-4Ra1 and
phosphorylated Tyk2 with IL-13Ra1. STAT1, STAT3, STAT5A
and B, and STAT6 were all tyrosine phosphorylated and activated in response to IL-13. Furthermore, IL-13 induction of the
gene that encodes 15-LO (15 lipoxygenase) expression required
p38 MAP kinase-mediated phosphorylation of STAT1 and
STAT3. 15-LO was regulated by a signaling complex with PKCd
kinase and both tyrosine and serine phosphorylation of STAT3.
PKCd was thus required for IL-13-induced 15-LO expression
and subsequent inhibition of the lipid mediator leukotriene B4.
IL-4 has substantial effects on SHIP1 and SHIP2, phosphatases that skew macrophage polarization (Rauh et al., 2005).
Immunity 32, May 28, 2010 ª2010 Elsevier Inc. 595
Immunity
Review
Type I
Figure 2. IL-4 and IL-13 Signaling
Machinery
Type II
IL-4
IL-13
IL-4
IL-13
IL-4Rα
Jak1
STAT6
IL-2Rγ
IL-4Rα
Jak3
IL-13Rα1
Jak1
IRS-2
Tyk2
STAT3 (?)
STAT6
PI3K
P
?
STAT6
Grb2
STAT6
IL-13Rα2
P
Hematopoietic cell
Nonhematopoietic cell
STAT6
STAT6
PP
PP
IL-4 binds IL-4Ra and then recruits either IL-2Rg
chain (to form the type I receptor) or the IL-13Ra1
chain (to make the type II receptor), depending
upon the cell type (Heller et al., 2008). However,
IL-13 can only signal through type II receptor as
it binds IL-13Ra1 and recruits IL-4Ra (LaPorte
et al., 2008). The common signaling pathways
shared by both the cytokines involve phosphorylation of STAT6, which leads to dimerization and
translocation of STAT6 to the nucleus where it
regulates the transcription of a number of genes.
The type I receptor is predominantly found in cells
of hemopoietic origin such as lymphocytes and
granulocytes, resulting from restricted expression
pattern of the IL-2Rg chain. The type II IL-4 receptor is mainly expressed by cells of nonhemopoietic
origin such as epithelial cells and fibroblasts.
Some cell types, such as macrophages, express
both receptor types. IL-13 also binds IL-13Ra2,
with an extraordinarily high affinity (Lupardus
et al., 2010), which is thought to function as a
decoy receptor (Chiaramonte et al., 2003), though
evidence exists for its potential to signal in
selected circumstances (Fichtner-Feigl et al.,
2006).
STAT6
PP
IL-4 triggers proteosomal degradation of both of these phosphatases in bone marrow-cultured mouse macrophages, enhancing
the PI3K and Akt kinase pathways (Ruschmann et al., 2010).
SHIP represses IL-4 production from basophils, thereby indirectly controlling the AAM phenotype (Kuroda et al., 2009).
The mTOR adaptor is an evolutionarily conserved member of
the PI3K family that forms the core of distinct TORC1 and
TORC2 signaling complexes, broadly responsible for the regulation of metabolism, growth, and proliferation (Delgoffe et al.,
2009). Recent studies have shown that mTOR is essential in
dictating T helper cell differentiation, TORC1 for Th1 and Th17,
and TORC2 for Th2 cell differentiation. It would be interesting
to examine macrophage polarization in mice after genetic ablation of these distinct pathways.
There are few reports of epigenetic modification or microRNA
analysis in AAMs. Signature marker genes of IL-4-treated mouse
macrophages, Arg1, Ym1, Fizz1, and MRC1, revealed reciprocal
changes in histone H3K4 and H3K27 methylation; the latter
methylation markers were removed by the H3K27 demethylase
Jmjd3 (Ishii et al., 2009). These changes depended on activation
of STAT6, which bound to consensus sites at the Jmjd3 promoter, contributing decreased di- and trimethylation of H3K27,
as well as transcriptional activation of AAM marker genes. In
vivo studies with Schistosome egg granulomas were consistent
with evidence of chromatin remodeling in AAM.
The kinase Akt regulates LPS-induced microRNA in macrophages and is implicated in LPS tolerance (Androulidaki et al.,
2009). The isoforms Akt1 and Akt2 have differential effects on
TLR4 and SOCS1 signaling in macrophages, depending on the
microRNAs let7e and miR155. The effect of IL-4 and IL-13 on
Akt isoforms has not been reported. Other studies have implicated microRNAs in regulation of macrophage activation
(Taganov et al., 2006; Tili et al., 2007).
596 Immunity 32, May 28, 2010 ª2010 Elsevier Inc.
Before global mRNA and protein repertoire analysis, knowledge of AAM markers was restricted to a few molecules.
MRC1, MHC class II, FN1, and CCL22 were upregulated in common in both human and mouse macrophages, leading to the
impression that they responded similarly to IL-4 (Gratchev
et al., 2001; Mantovani et al., 2005; Martinez et al., 2009). Subsequent reports showed that important AAM markers in mouse,
such as Fizz1, Ym1, and Ym2, lacked homologs in humans,
and others such as Arg1 were not upregulated in human macrophages by IL-4, despite gene homology.
Microarrays have made it possible to investigate effects of IL-4
in macrophages of both species; however, given the limited
number of published studies in human or mouse macrophages,
a consensus of markers is hard to establish, particularly for
humans (Syed et al., 2007; Martinez et al., 2006; Scotton et al.,
2005). Genes that appear to be modulated in common by IL-4
in macrophages, monocytes, and perhaps circulating precursors include ALOX15, FN1, CD23a, MAO-A, GAS6, FXIIIA1,
CCL26, CCL22, and IL-17RB. Many of the expected AA markers
could also be regulated by M-CSF, while a proportion of the
genes were uniquely regulated by IL-4.
Cell Biology and Membrane Dynamics in AAM
Apart from their effects on specific receptors and antigen
markers, IL-4 and IL-13 profoundly alter membrane flow in
macrophages (Kzhyshkowska and Krusell, 2009; Montaner
et al., 1999; Varin and Gordon, 2009). Human blood monocytederived macrophages displayed enhanced flow from the
plasma membrane, through early endosomes into an expanded
vacuolar compartment, accompanied by enhanced fusion with
lysosomes (Montaner et al., 1999). These effects are distinct
from those induced by other cytokines, such as IFN-g and
TNF. Apart from receptor-mediated endocytosis, e.g., via mannose, transferrin, and folate receptors (Puig-Kroger et al.,
Immunity
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2009), fluid phase endocytosis is also enhanced. The effects of
IL-4 and IL-13 on phagocytosis have been variable (Varin et al.,
2009).
In our hands, with mouse peritoneal macrophages, these cytokines selectively downregulate phagocytosis of a range of particles, by approximately 50%, independent of specific receptor
expression (Varin et al., 2009). Uptake of Neisseria meningitidis
via Class A Scavenger receptors, of zymosan particles via Dectin-1, and of latex particles is similarly decreased. Reduced
uptake is associated with reduced Akt phosphorylation, phagocytic cup closure, and PI3 Kinase activation. The mechanism
and functional significance of these observations require further
study.
In contrast with reduction of phagocytosis in AAMs, IL-4 is
required for the efficient uptake of apoptotic neutrophils by
macrophages from murine gp91 phox-deficient mice, a model
of chronic granulomatous disease (Fernandez-Boyanapalli
et al., 2009).
Another feature of IL-4 and IL-13 effects on macrophages is
the induction of homokaryon formation by macrophage fusion
(Helming and Gordon, 2009; Moreno et al., 2007). Foreign
body giant cell formation depends on IL-4, whereas osteoclast
formation, induced by RANKL cytokine, is inhibited by IL-4
(Moreno et al., 2003). Independent studies have demonstrated
murine multinucleated giant cell formation by IL-4 and IL-13
and studied its mechanism. Fusion depends on STAT6 signaling
and is homotypic, i.e., both fusing partners need to express
STAT6 (Moreno et al., 2007). Several surface molecules have
been implicated in fusion, including E-Cadherin, DC-STAMP
(dendritic cell-specific transmembrane protein), and CD36,
which is thought to interact with patches of exposed PS on
fusion partners (Helming and Gordon, 2009). Other molecules
such as DAP-12 induce a fusogenic phenotype required on
both fusion partners (Helming et al., 2008). The effects of IL-4
and IL-13 are not necessarily associated with enhanced expression of fusogenic surface molecules, but may result from clustering in the plasma membrane or altered signaling.
With human monocyte-derived macrophages, IL-4 and IL-13
accelerate fusion in vitro, but other culture variables, e.g.,
substratum, growth factors, and serum, also play a role. The
significance of giant cell formation in granulomatous inflammation is obscure, nor is it known whether IL-4 and IL-13 play
a role in macrophage fusion in nonparasitic infections, such as
the prototypical Langhans giant cells of tuberculosis.
Function of Signature Markers in AAM
In the case of enzyme markers such as Arg1, there has been
extensive investigation of its possible role in alternative versus
classical activation (De’Broski et al., 2010; El Kasmi et al.,
2008; Herbert et al., 2004; Pesce et al., 2009a). Although its
upregulation in the mouse can be a useful indicator of IL-4 and
IL-13 effects on macrophages, it can be upregulated by other
pathways. Its demonstration in human macrophages has been
problematic. Its possible role in fibrosis is described below.
Enhanced expression and endocytosis of MRC1 are conserved
features in both mouse and human macrophages; enhanced
uptake of pathogens through carbohydrate recognition may
contribute to infection. However, its immunologic role is
unknown. A possible clue is via a role in Th17 cell differentiation
(van de Veerdonk et al., 2009).
Ym1 and Ym2 belong to a family of chitinase-like molecules.
Chilectins are strongly induced by Th2 cell stimuli on macrophages as well as epithelium. Chilectins have undergone considerable diversification during evolution, in many cases losing
enzyme activity, as with Ym1 and Ym2. These molecules bind
chitin in insects, Helminth eggs, and Nematode pharynx. Chitin
induces Th2 cell responses (Reese et al., 2007; Voehringer
et al., 2006) and can be degraded by glucanases, and MRC1
contributes to chitin binding.
Fizz1, also known as Relma or Retnla, a secreted protein
produced by macrophages, epithelium, and eosinophils, is
highly induced by IL-13. Fizz1 found in lung allergy and asthma
may inhibit inflammation (Nair et al., 2009) Fizz / AAM display
exaggerated antigen-specific Th2 cell differentiation. Fizz1 protein binds to macrophages and effector CD4+ Th2 cells, inhibiting Th2 cytokine production via Bruton’s tyrosine kinase. Fizz1
plays a role in metabolism, as well as regulating T cell differentiation and eosinophil migration. Others have reported a proinflammatory role for Fizz1 and related resistin-like molecules (Munitz
et al., 2008).
Involvement of AAMs in Selected Pathological
Processes
Role of AAM in Inflammation and Immunity
AAMs are influenced by, and act upon, many processes associated with inflammation, immunity, and hypersensitivity. AAMs
counteract proinflammatory and cellular immune effector mechanisms, serving regulatory as well as inhibitory functions, to
shape the nature of the response. For example, IL-4 induces
expression of SOCS1 in human and mouse macrophages via
STAT6. SOCS1 feedback inhibits IL-4 signaling to limit expression of STAT6 responsive genes and the AAM pathway (Dickensheets et al., 2007).
Although intimately associated with IL-10, TGF-b, and other
agents of more profound deactivation, IL-4 and IL-13 promote
a distinctive inflammation and cellular immune response. For
example, NO generated by IFN-g-induced iNOS is shut down
in macrophages (it is induced in epithelial targets), with a shift
to Arg1. Interactions with mast cells, basophils, eosinophils,
and NKT cells, as well as IgE and selected subclasses of IgG,
promote allergy and hypersensitivity. Together they influence
vascular permeability, angiogenesis, cell recruitment, smooth
muscle contraction, goblet cell secretion, mucus production,
and collagen deposition by myofibroblasts (Locksley, 2010).
Several questions remain understudied. For example, the
interactions between AAMs and other cells of the innate and
acquired immune system, by contact, and secretion, require
attention. Do AAMs influence DCs, which themselves are
affected by Th2 cytokines, and can AAMs function as antigenpresenting cells (Loke et al., 2000; Rodriguez-Sosa et al.,
2002)? Is the reported upregulation of MHCII significant? Apart
from modulating inflammation, can AAM kill microbial and host
targets, particularly parasites? Arg1 has been put forward as
a candidate. Do IL-4-induced homokaryons participate in this
process? Perhaps killing is indirect through eosinophils or
neutrophils. In sum, the metabolic and secretory activities of
AAM favor trophic rather than lytic functions, clearance of
apoptotic corpses rather than induction of necrosis, and induction of tolerance rather than autoimmunity.
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Inflammasome Activation
Little is known about inflammasome activation and IL-1b release
by AAMs, compared with CAMs. Decrease of caspase 1 expression and pro-IL-1b processing has been described for human
monocytes stimulated with IL-13 (Scotton et al., 2005). A similar downregulation is observed in some human macrophage
models. An intriguing study by Pelegrin and Surprenant examined the role of extracellular ATP and purinergic receptors in
a spectrum of polarized macrophages. In intermediate, but not
fully AAMs, P2X7R activation uncouples NLRP3 inflammasome
maturation (Pelegrin and Surprenant, 2009). ATP acts, instead,
through pyrophosphate to inhibit IL-1b release.
Role of AAMs in Infection, Fibrosis,
and Immunopathology
The generation and role of AAMs has been studied extensively in
parasitic disease models (e.g., Anthony et al., 2006; Fallon et al.,
2006; Hesse et al., 2001; Holscher et al., 2006; Reece et al.,
2006), providing the best evidence for a regulatory, protective
role of AAMs in these Th2 cell-dominated infections. Earlier
studies, culminating in macrophage-specific genetic ablation
experiments, implicated the common IL-4Ra1 chain in Nippostrongylus resistance. IL-13 is critical for worm expulsion and
also plays an important role in Schistosome egg granuloma
formation and fibrosis. Murine markers of AAMs have provided
a remarkably conserved signature of gene expression, considering the diversity of organisms studied. Parasite-specific
features are associated with particular sites of migration during
complex life cycles, e.g., in skin, lung, gut, and liver, illustrating
the importance of tissue microenvironment in tuning the Th2
cell response. AAMs respond to Th2 cytokines and in turn regulate T helper cell functions. Delayed healing or excessive fibrosis
occurs in mice with dysfunctional AAMs (Wilson and Wynn,
2009).
Major questions remain unresolved regarding the role, if any,
of AAM in parasite killing and in activation or suppression of
different T cell subsets. Can AAM serve as parasite antigen-presenting cells for T cells? Chemokines produced by AAMs
promote recruitment and activation of eosinophils, mast cells,
and basophils, themselves sources of Th2 cytokines, as well
as neutrophils.
Interactions of AAMs with antibody subclasses promoted by
Th2 cytokines have received less attention, but include upregulation of FcR epsilon for IgE. The range of organisms studied in
the mouse includes H. polygyrus and flatworms, and wellstudied unicellular parasites include Leishmania and Trypanosoma brucei. In humans, filariasis has attracted investigators,
but the lack of reliable AAM markers has hampered progress.
Further details on the diversity of host responses to parasitic
infection can be found in excellent reviews (Jenkins and Allen,
2010; Wilson and Wynn, 2009).
A striking example of the way in which a bacterial pathogen
can subvert the host response to favor its intracellular survival
is provided by Francisella tularensis (Ft), the causal agent of tularaemia (Shirey et al., 2008), illustrated in Figure 3A. This highly
virulent coccobacillus has been studied in the form of a vaccine
LVS strain, attenuated for humans but still pathogenic for mice.
It enters macrophages by a TLR-2-dependent mechanism,
utilizing mannose and scavenger receptors, and initiates a transient inflammatory response, but the host CAM reaction is then
598 Immunity 32, May 28, 2010 ª2010 Elsevier Inc.
subverted by pathogen-induced reprogramming of the macrophage to acquire an AAM phenotype. Instead of the host-protective CAMs, it induces many characteristics of AAMs, by unknown
sensing receptors within the phagocytic vacuole and, on escape
into the cytoplasm, by additional cytosolic sensors. The macrophages are induced to produce IL-4 and IL-13, which amplify the
AAM phenotype through autocrine and paracrine mechanisms.
The canonical signature markers are induced, depending on
the IL-4 and IL-13 common alpha chain receptor and STAT6,
incapacitating the cell’s antimicrobial responses, and allowing
FT to replicate freely.
Similar though less dramatic exploitation of AAMs may contribute to persistent mycobacterial infection (Kahnert et al.,
2006). Another issue is the secondary infection of Th2 cellbiased infected hosts with opportunistic or pathogenic bacteria.
Challenge of IL-4 or IL-13 pretreated macrophages with Neisseria meningitidis can potentiate the release of proinflammatory
cytokines (Varin et al., 2009); AAMs can thus contribute to septic
shock.
Pathogens such as Aspergillus and Candida utilize TLR-2
and TLR-6 and the beta glucan receptor Dectin-1 to infect
macrophages (Brown, 2006). AAMs express enhanced Dectin-1 as well as mannose receptor, which is delivered to the
nascent phagosome intracellularly (Heinsbroek et al., 2009).
Dectin-1 contains a hemi-ITAM (immunoreceptor tyrosinebased activation) motif in its cytoplasmic domain, essential
for signaling, through Syk kinase and CARD9, thus inducing
proinflammatory cytokines and activating a respiratory burst.
The receptor contributes to host resistance to Candida and
Pneumocystis. However, it is not clear whether AAMs are beneficial or deleterious to the host, because other innate (complement, neutrophils) and adaptive mechanisms, including classical activation by IFN-g and Th17 cell activation, come into
play.
Respiratory syncytial virus (RSV), a major cause of morbidity in
children, employs a similar mechanism of AAM induction to that
described for Francisella, although in this instance the outcome
is beneficial to host repair and diminishes host pathology
(Figure 3B; Shirey et al., 2010). Studies with mouse macrophages in vitro and with cotton rats in vivo show that AAMs
play a role in the response to infection. The macrophages are
induced to express the marker signature of AAMs, which
depends on the IL-4 and IL-13 receptor common alpha chain
and STAT6, as well as TLR-4 and IFN-b. Infection in humans
may also involve Th2 cytokines and AAMs. Although markers
are insufficient to establish their role, other cell types, including
innate and adaptive immune and epithelial cells, also modulate
the host response.
Since the earliest reports on AAMs, it has been assumed that
these cells promote repair of host tissues after inflammation.
Induction of Arg1 by IL-4 and IL-13 has been implicated in
collagen deposition (Hesse et al., 2001) and degradation
(reviewed in Wilson and Wynn, 2009). Imbalance in production
and catabolism, associated with prolonged IL-13 effects on
macrophages, promotes excessive fibrosis. Wynn and his
colleagues and Murray, who has produced a conditional macrophage-specific deficiency of Arg1, provide evidence to support
as well as question these hypotheses (El Kasmi et al., 2008;
Pesce et al., 2009a, 2009b).
Immunity
Review
A
Ft
IL-4, IL-13
TLR2
IL-4, IL-13
(requires IL-4Rα and STAT6)
Inhibit Ft
replication
TLR2-dependent increase in:
FIZZ1, Ym1, mannose receptor,
arginase 1
Macrophage
Alternatively
activated macrophage
Exogenous
and endogenous IFN-β
IL-10
Proinflammatory cytokines
B
Promotes Ft replication
Kills host
RSV
IL-4, IL-13
Antiviral
Increase IL-4Rα
IFN-β
TLR4-dependent increase in:
FIZZ1, Ym1, mannose receptor,
arginase 1
IL-4, IL-13
(requires IL-4Rα and STAT6)
Lung macrophage
Steroids
COX-2
Repair of tissue damage
IL-10
Parecoxib
PGs
Alternatively
activated macrophage
Pathology
? Predisposition to
airway hyperreactivity
Figure 3. Alternative Activation Amplification Loops in Infection
(A) Model for the role of AAMs during Francisella tularensis (Ft) LVS infection. Ft infection of macrophages results in an initial proinflammatory response through
a TLR2-dependent pathway (Cole et al., 2007), as well as an unknown intracellular sensor that leads to IFN-b production (Cole et al., 2008) and activation of
caspase-1 through AIM2 and ASC sensors (Cole et al., 2008; Rathinam et al., 2010). IFN-b acts to inhibit bacterial replication, but production of IL-4 and
IL-13 differentiates the macrophages to an AAM phenotype through an IL-4Ra- and TLR2-dependent mechanism that results in a microenvironment that
promotes Ft replication, which is ultimately detrimental to the host.
(B) Model for the role of AAM during RSV infection. RSV infection of macrophages leads to an early induction of proinflammatory cytokines and COX-2, which has
been shown to mediate the observed lung pathology (Richardson et al., 2005). After this early proinflammatory period, macrophages produce their own IL-4 and
IL-13 and differentiate into AAM through an IL-4Ra-, TLR4-, and IFN-b-dependent mechanism. This is thought to promote repair of damage caused by the earlier
inflammatory process by inducing AAM markers including Arg1, FIZZ1, and Ym1. IL-10 acts to downregulate production of inflammatory cytokines.
Figure adapted from Shirey et al. (2010) with permission.
Although sterile foreign body reactions can depend on IL-4,
IL-4Ra1 chain, and STAT6, repair of surgical wounds was independent of this pathway (Daley et al., 2010). The IL-13-specific
receptor was not investigated although it has been shown to
be essential for fibrosis in a parasitic model of S. mansoni granuloma formation (Ramalingam et al., 2008). As noted, Arg1 can
be induced by a variety of different pathways, and cells other
than macrophages also express receptors for IL-13. The reduction of fibrosis in egg granulomata in macrophage-selective
ablation of the IL-13Ra1, in which induction of AAM is unaffected, confirms the existence of AAM-independent pathways
of fibrogenesis, and also independent of TGF-b or MMP9, which
are enhanced in this model.
Macrophage-specific ablation of Arg1 enhances S. mansoni
egg-induced fibrosis, an unexpected result. In wild-type mice,
granulomata appear in liver after 7–10 weeks and diminish
subsequently by Arg1-dependent downregulation, which is
absent in Arg1-deficient mice. Macrophages express several
transporters such as Cat2 (Thompson et al., 2008) as well as
enzymes involved in L-arginine uptake and metabolism, but
not a complete urea cycle. Ornithine decarboxylase, hydroxyproline, and polyamines have been implicated in additional
properties of AAMs. Polyamines contribute to induction of
Cadherin-1, after IL-4 treatment in vitro or Taenia crassiceps or
allergen-induced activation in vivo (Van den Bossche et al.,
2009). Cadherin-1 expression at the macrophage surface,
together with catenin, mediates homotypic (cell fusion) and
heterotypic interactions with CD103+ T cells. Alternative activation of macrophages has also been implicated in fibrosis after
gamma herpes virus infection in mice (Gangadharan et al., 2008).
It is not clear to what extent Th2 cytokines, AAMs, or fibroblasts contribute to noninfectious causes of fibrosis or whether
combined targeting of IL-13 and TGF-b might prevent or reverse
fibrosis.
A remarkable example of a pathogenic role of IL-13 in chronic
obstructive pulmonary disease (COPD) has been reported
(Holtzman et al., 2009; Kim et al., 2008). COPD is a major cause
of morbidity and mortality, in which lung failure is associated with
chronic inflammation, mucous cell metaplasia, bronchial hyperreactivity, and fibrosis. The condition is often attributed to allergens and adaptive immune mechanisms, but this study provides
convincing evidence that an initial transient acute viral infection
in the lung can be followed after a period by activation of NKT
cells, production of IL-13 by these cells, and by macrophages
Immunity 32, May 28, 2010 ª2010 Elsevier Inc. 599
Immunity
Review
themselves. The macrophages upregulate IL-13Ra1 expression
and become alternatively activated by an autocrine or paracrine
mechanism.
In an experimental mouse model of Sendai virus infection,
bronchiolitis is associated with mucus production and airway
hyperreactivity to methacholine. After the virus becomes barely
detectable, NKT cells expressing semi-invariant Valpha14-Jalpha 18 TCR produce IL-13 through a CD1d-glycolipid-dependent mechanism, initiating a vicious circle of AAM amplification.
In the mouse, the characteristic signature markers Arg1, Ym1
and Ym2, Fizz1, Mmp12, and Alox12e are now present.
An analogous mechanism has been proposed for human
COPD, with patient transplant-derived material to demonstrate
the presence of NKT cells and IL-13 message and protein in
macrophages (Holtzman et al., 2009). The critical role of IL-13
in the mouse model was demonstrated by macrophage depletion studies and blockade with soluble IL-13R2 protein. This
provocative study shifts the pathogenic mechanism to the innate
immune system. Improved AAM markers are needed for human
tissue analysis and prospective studies in human subjects.
Plasma Chitinase-1, a member of the chitinase family, allows
quantitative stratification of COPD patients (Agapov et al., 2009).
The question arises of genetic susceptibility to these sequelae of
common virus infections, as does the possibility of therapeutic
targeting of the IL-13 pathway.
LPS tolerance, a refractory state in responses of the host and
macrophages to repeated challenge with LPS, is known to
involve NF-kB. Patients with sepsis can display features resembling alternative activation of monocytes. Porta and colleagues
have shown that tolerance and alternative activation (M2 polarization) are related processes mediated by p50 homodimers,
which lack a transcription activation domain of members of the
NF-kB pathway (Porta et al., 2009). Studies were performed
with human monocyte-derived macrophages and with mouse
primary macrophages, in vitro and in vivo. P50 was shown to
be a negative regulator of classical, M1 polarization and of
IFN-b production, but played a positive, nonredundant role in
induction of several signature M2-associated genes. Markers
included selected chemokines (CCL2, CCL17, and CCL22),
Arg1, Ym1, and Fizz1. There were some discrepancies between
human and mouse phenotypes, however, and no reported
involvement of IL-4 and IL-13 expression or of their receptors.
P50 / mice lacked the characteristic AAM phenotype associated with Taenia crassiceps infection, consistent with the overall
hypothesis.
In another model, the prototypical proinflammatory function of
TLR4 can be subverted by house dust mite allergen, which
mimics MD2, its coreceptor, to induce IL-4 production (Trompette et al., 2009).
Although IL-13 has been associated with disease-promoting
activities, it can also play a protective role by virtue of its ability
to regulate macrophages in an autoimmune myocarditis model
of disease (Cihakova et al., 2008). Infection of mice with Coxsackie virus B3 induces a self-limiting myocarditis in most mouse
strains, but a few strains develop severe myocardiopathy, with
Th1 cell type inflammation, myosin antibodies, and fibrosis,
especially in IL-13 / , but not IL-4 / , BALB/c mice. T cells are
not known to express IL-13Ra, and the protective role of IL-13
has been attributed to macrophages. MRC1 expression was
600 Immunity 32, May 28, 2010 ª2010 Elsevier Inc.
reduced in macrophages from the hearts of IL-13 or doubly deficient IL-13 and IL-4, but not IL-4-deficient, mice.
PPARs and AAMs: Role in Metabolism, Inflammation,
and Atherosclerosis
Since the early observation by Glass and colleagues that PPAR
gamma is strongly induced by IL-4, PPARs have been investigated in relation to lipid metabolism, macrophage polarization,
obesity and insulin resistance, and atherogenesis (reviewed by
Glass [Glass and Saijo, 2010] and Chawla [Chawla, 2010]).
Macrophages express PPARg and PPARd; macrophage-specific deficient mice have been utilized in bone marrow chimera
experiments of experimental atherosclerosis. Although there
has been progress in analyzing the control of inflammation, the
therapeutic application of PPAR ligand agonists via these receptors remains elusive. Macrophages in white adipose tissue
in obese versus lean mice differ in phenotype, with features of
CAMs in the former and of AAMs in the latter. By contrast,
lipid-laden foam cells in atherosclerotic plaques have a distinct
phenotype, not necessarily corresponding to the binary classification of M1- and M2-polarized cells. AAMs could play a role in
smooth muscle cell migration and plaque rupture.
IL-4 enhances uptake and oxidation of fatty acids and biogenesis of mitochondria, via STAT6, regulating programs controlled
by PPARg and the coactivator protein PGC-1b. PPARg is
dispensable for induction of AAM but required to sustain it. It
plays a more complex role in limiting proinflammatory gene
expression by LPS, preventing turnover of NCoR corepressor
complexes and keeping such genes in the ‘‘off’’ position in
AAMs, thus controlling the initiation, magnitude, and duration
of inflammation.
PPARg regulates primarily the metabolic phenotype of AAMs,
whereas PPARd regulates the immune repertoire. PPARd is
a sensor of triglyceride VLDL and contributes to macrophage
numbers and phenotype in liver as well as adipose tissue.
Mono-unsaturated fatty acids, such as oleic acid, synergize
with IL-4 to enhance expression of AAM signature genes; PPARd
may also play a role in the proliferative and anti-inflammatory
effects of IL-4 on macrophages.
Role of AAMs in Tumorigenesis
Macrophages are a prominent component of the stroma and
leukocyte infiltrate in human and experimental mouse tumors
(DeNardo et al., 2009; Gocheva et al., 2010; Grivennikov et al.,
2010; Hallam et al., 2009; Joyce and Pollard, 2009; Mantovani
et al., 2008; Sinha et al., 2007). They interact with malignant
epithelial, blood vessels and lymphatics, lymphocytes, innate
cells, and extracellular matrix. They have been implicated in
carcinogenesis, the angiogenic switch, local invasion, and
metastasis. Systemic features of malignancy have also been
ascribed to macrophages, locally and outside the actual tumor
bed. It is established that they contribute trophic functions to
the emergence of nascent tumor clones, phagocytose apoptotic
tumor cells, influence the tissue response to hypoxia and,
together with regulatory T lymphocytes, suppress Th1 cell and
cytotoxic T lymphocyte antitumor responses.
The nature of tumor-associated macrophages (TAMs) has
become of great interest, though confusing because of phenotypic heterogeneity. TAMs have many properties of AAMs;
myeloid-derived suppressor cells (MDSC) are less defined and
Immunity
Review
in mouse contain infiltrating GR-1 intermediate CD11b+ monocytes, as well as GR-1 high CD11b+ granulocytic cells (Gabrilovich and Nagaraj, 2009). Some MDSC express F4/80 in the
mouse and IL-4Ra, enabling them to respond to both IL-4 and
IL-13. Innate and adaptive T cells as well as tumor cells can
produce these cytokines, and some tumors also express the
IL-4 and IL-13 alpha receptor (Mandruzzato et al., 2009).
Several recent studies with experimental tumors in mice
(transplantable as well as oncogene-induced) provide striking
evidence that TAMs have much in common with AAMs. They
contribute to local invasion through cathepsin B and S, promote
angiogenesis and tumor growth through VEGF, and induce
recruitment of other hemopoietic cells, through chemokines
(Gocheva et al., 2010). AAMs as well as tumor cells themselves
orchestrate tumor growth and spread through M-CSF, TNF,
IL-10, TGF-b, and other products. The evidence that alternative
activation of macrophages is important in promoting tumorigenesis includes expression of murine signature markers and
dependence on the IL-4 and IL-13 common receptor alpha
chain. Blockade of this receptor or its genetic ablation may
therefore shift the host-tumor balance toward tumor rejection
by Th1 cell-dependent effector mechanisms, such as IL-12
versus IL-10 production. A recent study (Weiss et al., 2009)
combined treatment with IL-2 and anti-CD40 to induce a switch
from M2 to M1 macrophage functions, depending on CCR2 and
IFN-g. The contribution of TAM to this balance varied in different
tumor models and could affect either primary or secondary
growth. B cell-dependent mechanisms have been demonstrated
in selected tumors (Andreu et al., 2010), with possible interactions with AAM Fc receptors.
IL-13 and its receptors have been implicated in a different
model of suppression of host immune effector responses to
tumors (Fichtner-Feigl et al., 2008a). IL-4 or IL-13, acting through
the IL13Ra1, is able to upregulate expression of the IL-13Ra2
polypeptide. TNF and NF-kB contribute in parallel to the
STAT6 pathway. IL-13 acts on GR1 intermediate CD11b high
monocytic cells, through variant AP1 transcription factor, to
produce TGF-b. This pathway is responsible for suppression of
tumor rejection by Th1 cell-type immune response. Treatment
with anti-TNF or siRNA to block IL-13Ra2 reduces metastasis
and promotes survival.
In summary, AAMs are important candidates for therapeutic
modulation of the tumor-host inflammatory interaction. However, AAM markers are urgently required to establish their contribution in naturally occurring human malignancy.
Therapeutic Interventions
As knowledge of the pathophysiologic pathways of IL-4 and
IL-13 in human disease has grown, attempts are being made
to achieve therapeutic blockade. In a well-controlled phase 2a
trial (Wenzel et al., 2007), recombinant human pitrakinra showed
promising results in ameliorating late-phase asthmatic response
to allergen challenge in patients, by aerosol and subcutaneous
routes. This agent, previously tested in cynomolgus monkeys,
is a potent inhibitor of binding of both IL-4 and IL-13 to IL-4R
receptor complexes, offering an advantage over previous
attempts to inhibit IL-4 alone, IL-5, or IgE.
Selective blockade of IL-13 signaling could offer further
advantages, e.g., in a subpopulation of asthma patients (Wood-
ruff et al., 2009). Various strategies have been employed to target
IL-13 through receptor-specific antibodies, chimeric proteins,
or soluble receptor (Blease, 2008). Proof-of-concept animal
and early human studies have been undertaken, to ameliorate
asthma and idiopathic lung fibrosis. Species differences need
to be borne in mind, because in contrast with the mouse, human
decoy IL-13 receptor is not secreted from cells (O’Toole et al.,
2008). Novel strategies include functional variants based on
analysis of human SNPs that demonstrate altered affinity for
IL-13Ra2 and cell penetrant proteins or small molecules that
target the signaling pathway through STAT 6. Structure-function
studies should aid in the development of IL-13-specific
therapeutic agents (LaPorte et al., 2008; Mitchell et al., 2010;
Lupardus et al., 2010).
Conclusion and Further Questions
Although there has been striking progress in understanding the
mechanisms and functions of AAM, much remains to be discovered. We have argued in favor of a restricted definition, shown
the value of in situ analysis in mouse and man, and pointed
to the possibility of selective targeting of pathogenic pathways
through IL-13. Major human diseases, including infection,
inflammation, fibrosis, and malignancy, demonstrate the twoedged nature of AAM function at different stages of disease.
The need for validated human markers is apparent. Perhaps
less obvious is a clearer understanding of what determines the
spectrum of innate, classical, alternative, and other activation
phenotypes in macrophage populations and individual cells.
The trophic, antigen-presenting, and killing functions of AAMs
need better characterization. Finally, does alternative activation
of macrophages extend to other innate cells, in particular dendritic, NKT, and even nonhematopoietic cells? As in other fields
of immunobiology, the alternative may indeed be the antecedent
path in evolution.
ACKNOWLEDGMENTS
We thank colleagues for sharing unpublished results, Stefanie Vogel and Keri
Ann Shirey for Figure 3, and Thiru Ramalingam for Figure 2. This review was
prepared while S.G. was a visiting NCI/NIAID scholar and he thanks Giorgio
Trinchieri and Alan Sher for their hospitality. F.O.M. was supported by the
British Heart Foundation.
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