Akt Signaling Pathway in Macrophage

Activation and M1/M2 Polarization

This information is current as
of May 19, 2020.
Eleni Vergadi, Eleftheria Ieronymaki, Konstantina Lyroni,
Katerina Vaporidi and Christos Tsatsanis
J Immunol 2017; 198:1006-1014; ;
doi: 10.4049/jimmunol.1601515
http://www.jimmunol.org/content/198/3/1006

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Brief Reviews
Akt Signaling Pathway in Macrophage Activation and
M1/M2 Polarization
Eleni Vergadi,*,† Eleftheria Ieronymaki,* Konstantina Lyroni,* Katerina Vaporidi,† and
Christos Tsatsanis*
Macrophages become activated initiating innate immune
responses. Depending on the signals, macrophages obtain
an array of activation phenotypes, described by the broad
terms of M1 or M2 phenotype. The PI3K/Akt/mTOR
pathway mediates signals from multiple receptors includ-
ing insulin receptors, pathogen-associated molecular pat-
tern receptors, cytokine receptors, adipokine receptors,
and hormones. As a result, the Akt pathway converges
inflammatory and metabolic signals to regulate macro-
phage responses modulating their activation phenotype.
Akt is a family of three serine-threonine kinases, Akt1,
Akt2, and Akt3. Generation of mice lacking individual
Akt, PI3K, or mTOR isoforms and utilization of RNA
interference technology have revealed that Akt signaling
pathway components have distinct and isoform-specific
roles in macrophage biology and inflammatory disease
regulation, by controlling inflammatory cytokines, miR-
NAs, and functions including phagocytosis, autophagy,
and cell metabolism. Herein, we review the current
knowledge on the role of the Akt signaling pathway in
macrophages, focusing on M1/M2 polarization and
highlighting Akt isoform–specific functions. The Jour-
nal of Immunology, 2017, 198: 1006–1014.
M
acrophages respond to pathogens and other nox-
ious stimuli, such as apoptotic or necroptotic cell
debris, and initiate inflammatory responses. Acti-
vation of macrophages is mediated by signaling cascades
downstream of TLR and cytokine receptors. Upon activation,
transcriptional and epigenetic changes lead to production of
proinflammatory cytokines and chemokines, migration, and
elimination of the insult. At later stages of inflammation,
macrophages contribute to the resolution of inflammation,
and prohibit prolonged inflammatory responses that may lead
to tissue injury. The diminished response following pro-
longed activation has been described as endotoxin tolerance
when the inflammatory stimulus derives from Gram-negative
bacterial LPS.
*Laboratory of Clinical Chemistry, School of Medicine, University of Crete, Heraklion
71003, Greece; and †Laboratory of Intensive Care Medicine, School of Medicine, Uni-
versity of Crete, Heraklion 71003, Greece
ORCIDs: 0000-0003-4026-0328 (E.V.); 0000-0002-8441-298X (E.I.); 0000-0001-
7446-0999 (K.L.); 0000-0003-1214-4151 (C.T.).
Received for publication August 30, 2016. Accepted for publication September 26,
2016.
This work was supported by Greek national and European Union funds under the
Aristeia Grant (2071).
www.jimmunol.org/cgi/doi/10.4049/jimmunol.1601515
The Journal of
Immunology
The different stages of macrophage activation are broadly
described as M1 and M2 polarization, both consisting of an
array of states that depend on converging signals from in-
flammatory stimuli and the cellular environment (1). M1 and
M2 status can be defined according to the activation stimulus.
The M1 spectrum includes M(IFN-g), M(LPS+IFN-g), and
M(LPS) depending on whether the signal is IFN-g or LPS,
whereas the M2 spectrum includes M(IL-4), M(Ic), M(IL-
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10), M(GC+TGF-b), and M(GC) when the stimulus is
from IL-4, immune complexes (Ic), or glucocorticoids (GC)
in combination or not with TGF-b (1). These phenotypes
describe the different states of macrophages and their func-
tions, which range from elimination of diverse pathogens to
phagocytosis of apoptotic cells and tissue remodeling. Ac-
cordingly, M(IFN-g) macrophages are associated with Th1-
related pathologies where IFN-g is abundant, and M(LPS) as
well as M(LPS+IFN-g) macrophages are found in Gram-
negative bacterial infections before and after production of
IFN-g from Th1 cells. The M2 spectrum of macrophages is
broader and includes macrophages that contribute to parasitic
infections or other pathologies with abundant IL-4 such as
asthma (2, 3), activated by immunocomplexes, termed M(Ic),
such as those found in rheumatoid arthritis (4) and sclero-
derma (5) patients, activated by IL-10, termed M(IL-10),
found in the tumor stroma or in inflamed tissues (6, 7), or
induced by abundant glucocorticoids, termed M(GC), such as
late stages of sepsis (8, 9) or in cancer (9, 10). Macrophage
activation is a tightly regulated phenomenon, determined by
several signaling cascades elicited by receptor stimulation or
intracellular regulatory proteins. Many of the aforementioned
activation signals use common and distinct signaling cascades,
some of which are used for defining macrophage phenotype.
Among the different signaling cascades, the PI3K/Akt path-
way and its downstream targets have emerged as central reg-
ulators of activation phenotype in macrophages.
PI3K/Akt signaling regulates macrophage activation
Since the discovery of Akt 25 y ago (11) and identification of
PI3K as its upstream regulator (12), the contribution of Akt
Address correspondence and reprint requests to Prof. Christos Tsatsanis, Laboratory of
Clinical Chemistry, School of Medicine, University of Crete, PO Box 2208, Heraklion
71003, Crete, Greece. E-mail address: tsatsani@uoc.gr
Abbreviations used in this article: ApoE, apolipoprotein E; DSS, dextran sodium sulfate;
EAE, experimental autoimmune encephalomyelitis; LDLR, low-density lipoprotein re-
ceptor; mTORC, mTOR complex; RA, rheumatoid arthritis; TAM, tumor-associated
macrophage; TSC, tuberous sclerosis complex.
Copyright Ó 2017 by The American Association of Immunologists, Inc. 0022-1767/17/$30.00
The Journal of Immunology
in multiple cell types and functions has been widely recog-
nized. The PI3K/Akt pathway regulates macrophage survival,
migration, and proliferation but also orchestrates the response
to different metabolic and inflammatory signals in macro-
phages (13). The PI3K/Akt pathway is activated by TLR4 and
other pathogen recognition receptors, cytokine and chemo-
kine, and Fc receptors (14–17), modulating downstream sig-
nals that control cytokine production. Activated PI3K type I
phosphorylates phosphatidylinositol 4,5-bisphosphate (PIP2)
to generate phosphatidylinositol 3,4,5-triphosphate (PIP3) at
the plasma membrane; PIP3 further recruits Akt and the
mechanistic target of rapamycin complex (mTORC) 2, and
facilitates Akt activation by mTORC2 (13, 14). Activated
(phosphorylated) Akt subsequently phosphorylates and inac-
tivates the tuberous sclerosis complex (TSC) 1/2. TSC1/2
inhibition by Akt leads to mTORC1 activation, via Ras ho-
molog enriched in brain (termed Rheb) suppression (14, 16).
Activation of the PI3K/Akt pathway is critical in restricting
proinflammatory and promoting anti-inflammatory responses
in TLR-stimulated macrophages (18), and has been consid-
ered as a negative regulator of TLR and NF-kB signaling in
macrophages (15, 16) (Fig. 1). Activation or overexpression of
PI3K or Akt kinases resulted in reduced macrophage activa-
tion by LPS, whereas nonspecific chemical inhibition of PI3K
signaling in TLR-activated cells augments NF-kB activation
and inducible NO synthase expression (19–21) promoting
M1-type macrophage responses. PI3K activation has been
reported as an essential step toward M2 activation of mac-
rophages in response to surfactant protein A or IL-4 (22, 23).
A crosstalk between the STAT6 and PI3K activation is re-
quired for IL-4–induced M2 macrophage activation in SHIP-
deficient macrophages (22). Akt activation is required for M2
activation, because Akt inhibition abrogates the upregulation
of M2 genes (14, 24, 25). Several signals such as TGF-b, IL-
10, and BMP-7 promote M2 polarization via PI3K/Akt sig-
naling (26–28). Furthermore, induction of IL-10 in macro-
phages by proinflammatory signals requires activation of Akt
(17, 29). Nevertheless, individual PI3K and Akt isoforms also
contribute to M1 polarization (30–32).
Signaling cascades affected by PI3K/Akt contribute to macrophage
polarization
TLR signaling regulation by PI3K/Akt can be both direct and
indirect. The PI3K/Akt1 axis has been found to upregulate
FIGURE 1. Stages of macrophage activation and the
contribution of Akt signals.
1007
IL-1 receptor–associated kinase M (IRAK-M), a suppressor of
TLR4 signaling via TRAF6 inactivation (33). Activated Akt
also phosphorylates and inactivates the transcription factor
FOXO1 (34), resulting in suppression of its target genes that
include TLR4 (35). Akt also regulates let-7e, which in turn
suppresses TLR4 expression (36) and miR-146a, which sup-
presses the TLR signaling mediator TRAF6 (32).
In most studies, the PI3K/Akt/mTOR pathway was studied
as a single entity, and the effect of different isoforms of PI3K or
Akt kinases was not defined. However, studies on knockout
mice and studies using small interfering RNAs and chemical
inhibitors provided evidence on the differential function of
PI3K/Akt kinase isoforms on macrophage activation. The Akt
family of serine-threonine protein kinases consists of three
isoforms expressed from independent genes (Akt1/PKBa,
Akt2/PKBb and Akt3/PKBg) (12, 37). Akts are activated by
PI(3,4,5)P3 and to a lesser extent by PI(3,4)P2 that are generated
by PI3K class I activation (12, 37, 38). Class IA PI3K includes
three members, namely, p110a, p110b, and p110d, which in-
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teract with an SH2 domain–containing p85 regulatory subunit
(31, 39). PI3Kg belongs to class IB PI3K with its catalytic
subunit p110g coupled with either the p101 or the p84 regu-
latory subunit. Among PI3K members, PI3Kd and PI3Kg are
the predominant isoforms expressed in cells of the hematopoietic
lineage (39). Whether individual Akt isoforms are activated by
specific PI3K members in macrophages remains to be elucidated.
Nevertheless, isoform-specific effects on macrophage function
have been reported both for Akt and PI3K.
Akt upstream signals in macrophage polarization: the role of PI3K
isoforms, PTEN, and PDK-1. PI3K, the upstream regulator of
Akt, is shown to mediate M2 macrophage phenotypes (19, 23,
40). Accordingly, induction of Arg1 activity and M2 phe-
notype in SHIP-deficient macrophages depends on PI3K
activity, and specifically on class IA PI3K p110d isoform
(22). In addition, upregulation of miR-24 in macrophages,
a negative regulator of M1 activation that promotes M2
polarization, results in reduced expression of the class IA
PI3K subunit p110d (41). Activation of PI3Κg and PΙ3Kd
has been implicated in alveolar macrophage activation by
immune complexes via Fcg receptor (42), and is thus
involved in proinflammatory signaling, whereas the anti-
inflammatory effect of magnesium sulfate and naloxone
are mediated by PI3Kd and PI3Kg kinase isoforms (43, 44).
In the human monocytic cell line THP-1, the p110a PI3K
1008
subunit was required for phagocytosis, oxidative burst, and
secretion of proinflammatory mediators, whereas it nega-
tively regulated the production of TNF-a and IL-10 (45).
Furthermore, macrophages deficient for PI3K p85a regulatory
subunit have impaired NO and IL-12 production in response
to LPS and IFN-g stimulation, becoming vulnerable to
intracellular bacterial infection (46). Despite the distinct
functions of PI3K subunits, a degree of redundancy among
them has been described in several aspects of macrophage
activation (31, 47). The PI3K regulator lipid phosphatase,
phosphatase and tensin homolog (PTEN), also contributes
to macrophage polarization because deletion of PTEN
results in elevated levels of Arg1 and M2 polarization (48,
49), suggesting that regulation of PI3K/Akt signaling is a
central node for controlling macrophage polarization. PTEN
antagonizes PI3K by converting PI(3,4,5)P3 into PI(4,5)P2,
both recruiting Akt to the cell membrane (50), and loss of
PTEN results in increased Akt activity accompanied by
suppressed LPS responses in macrophages (19). Generation
of PI(4,5)P2 augments LPS responses (51), and loss of SHIP,
which converts PI(3,4,5)P3 to PI(4,5)P2, results in M2-skewing
of macrophages (52). Upon recruitment to the cell membrane,
Akt is phosphorylated and activated by phosphoinositide-
dependent kinase 1 (PDK-1). Loss of PDK-1 in myeloid
cells results in increased responses to LPS and susceptibility
of mice to endotoxin shock (53).
Akt downstream signals in macrophage polarization: the role of
mTOR. The TSC/mTORC1 pathway, a downstream
effector of PI3K/Akt, is a key sensor of nutrient status that
is also activated by inflammatory stimuli, playing a role in
coordination of metabolic and inflammatory signals to
determine macrophage activation phenotype (14, 54). There
are two complexes of the mTOR: mTORC1 (Raptor) that is
negatively regulated by TSC1/2 and mTORC2 (Rictor) that
phosphorylates and activates Akt (55–57). Akt promotes
mTORC1 activation and activated mTORC1 mediates
feedback inhibition to suppress mTORC2 and Akt activity
(14, 24, 58).
Current evidence on the role of TSC1/2 mTOR in the
regulation of macrophage activation supports that deletion
of TSC1 promotes LPS-induced proinflammatory cytokine
production, M1 polarization, and suppression of IL-4–
induced M2 polarization. The hypersensitivity of TSC1-
deficient macrophages to LPS stimulation depends on
mTORC1-induced attenuation of Akt signals. Attenuated
Akt signaling results in increased inflammatory responses in
TSC-deficient macrophages (14, 24, 58, 59). In contrast,
another group showed that deletion of TSC1 enhances both
M2 and M1 polarization (60). In an independent study in
Table I. PI3K/Akt signaling components and their contribution in M1 or M2 phenotype
Molecules Contributing to
M1 Polarization References
Akt2 (30, 32)
p110d (41, 42)
PTEN (48, 49)
TSC1 (60)
TSC2 (61)
p85a (46)
List of individual PI3K/Akt signaling components that contribute to M1 and M2 polarization.
BRIEF REVIEWS: Akt SIGNALING IN MACROPHAGES
human macrophages, TSC2 deletion decreased the M1 in-
flammatory response in an mTOR-dependent manner (61).
Using rapamycin to suppress mTORC1 has also produced
contradicting results. Rapamycin treatment has been found to
promote a shift to an M1-like profile, enhancing M1 and
reducing M2 polarization in human and murine macrophages
(61–63) and to abrogate the anti-inflammatory effects of
glucocorticoids in monocytes (64, 65) via enhancing NF-kB
and blocking STAT3 activity. Similar effects were shown
when mTORC2 was blocked. mTORC2-deficient, bone
marrow–derived macrophages showed increased M1 activa-
tion in response to LPS stimulation and higher mortality in
sepsis (66, 67). A summary of the contribution of individual
PI3K/Akt signaling components in M1 and M2 polarization
phenotypes is provided in Table I. It remains to be investi-
gated whether isoform-specific crosstalk between PI3K, Akt,
and mTORC components dictate M1 or M2 polarization of
macrophages, which may also partly explain the differences
observed within previous reports.
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Role of individual Akt isoforms in macrophages
Recent studies have shown that Akt1 and Akt2 kinase iso-
forms hold a key role in the regulation of macrophage acti-
vation (30, 36, 68). Deletion of Akt1 promotes upregulation
of inducible NO synthase and IL-12b (M1 activation) and
enhanced bacteria clearance (30, 69), increased responses to
LPS, and abrogation of endotoxin tolerance (30, 36). The
effect of Akt1 kinase in murine macrophages has been shown
to be modulated via induction of miR-155 and suppression of
C/EBPb, a transcription factor that is a master regulator of
M2 differentiation (30, 36, 68, 70, 71). Induction of miR-
155 in Akt12/2 macrophages sustains and amplifies M1 ac-
tivation, and enhances their bactericidal capacity (72–76),
partly via targeting suppressor of cytokine signaling 1 (72,
73). Akt1 deficiency and miR-155 induction in murine
macrophages activate the autophagic machinery by reducing
mTORC1 kinase activity and Rheb levels, two negative reg-
ulators of autophagy and key innate defense mechanisms
against many invading pathogens (77–82). Akt12/2 macro-
phages have impaired mitophagy and reduced expression of
TGF-b1, and are shown to be protective in experimental
pulmonary fibrosis (83). Akt1 also mediates suppression of
TLR4-induced macrophage activation by adipokines and
neuropeptides. Vasoactive intestinal peptide suppresses TLR4
responses via Akt1 (84). Similarly, induction of IRAK-M and
suppression of LPS responses by the adipokine adiponectin
are abrogated in Akt12/2 macrophages (85).
Akt2 deficiency in macrophages highlights opposing roles of
Akt isoforms in macrophage polarization, because Akt22/2
Molecules Contributing to
M2 Polarization References
Akt1 (30, 36)
p110a (45)
p110b (43)
p110g (44)
TSC1 (24, 58–60)
Rictor/mTORC2 (66, 67)
The Journal of Immunology
macrophages possess an M2-like phenotype. Akt22/2 mac-
rophages express elevated levels of the M2 transcription factor
C/EBPb and the M2 markers Arg1, Ym1 and Fizz1 (30).
Furthermore, Akt2 deficiency in macrophages results in
negative regulation of TLR4 signaling pathway via upregu-
lation of miR-146a, an miRNA targeting IRF5, TRAF6 and
IRAK1 and associated with M2 polarization (86–88). In ac-
cordance with their M2-like phenotype, Akt2-deficient mac-
rophages express elevated levels of LPS-induced IL-10 (89).
Akt signaling in phagocytosis and autophagy
A major attribute of macrophages is active elimination of
pathogens through phagocytosis. To initiate phagocytosis
macrophages respond to chemokines, which induce their re-
cruitment to the site of inflammation or tissue damage.
Chemotaxis and migration precedes phagocytosis and involves
signals that prime macrophages to become activated and en-
gulf pathogens or cell debris. Chemotactic signals promote
actin polymerization and formation of filopodia, facilitating
movement to the site of inflammation or injury. The effect of
individual Akt isoforms on migration and actin polymerization
has been demonstrated in neutrophils, where, upon stimula-
tion with chemoattractants, Akt2 but not Akt1 translocates to
the membrane contributing to migration and formation of
filopodia and lamellipodia. The same signals promote secretory
granule release facilitating innate immune responses (90). In
macrophages, absence of Akt2 results in defective CSF-1 and
MCP1 signaling, impaired LIMK/cofilin phosphorylation
resulting in attenuation of chemokine-induced actin poly-
merization and migration (91). Phagocytosis requires reor-
ganization of actin filaments to engulf foreign bodies. Signals
that initiate phagocytosis are mediated by receptors recog-
nizing molecules that will be engulfed. CD300, a receptor
that recognizes outer membrane–exposed phosphatidylserine
present on apoptotic bodies, promotes F-actin polymerization
via Akt activation (92). Activation of Akt appears essential for
phagocytosis because suppression of Akt2 but not Akt1 or Akt3
inhibits phagocytosis of opsonized beads from macrophages
(93). Different signals upstream of Akt control macrophage
phagocytosis. Thus, Fc receptor–mediated phagocytosis is aug-
mented by constitutively active Akt by activating p70S6 kinase
(94). Deletion of PTEN results in enhanced Akt activity and
increased Fcg receptor–mediated phagocytosis (95). At the
same time Fc receptor signals, via Akt, activate transcription of
IL-10 and IL-12 (17), orchestrating responses to immune
complexes. Engulfment of apoptotic cells triggers signals that
activate Akt and suppress ERK1/2 pathways, and at the same
time promote macrophage survival.
Pathogens or cell debris are engulfed into the phagocytic
bodies which are fused with lysosomes to be eliminated by
autophagy, a process primarily regulated by the mTOR
pathway (96). Autophagy is widely known as the process that
provides an endogenous source of nutrients to secure cell
survival under starvation conditions or during cell stress. The
role of Akt in autophagy reveals its significance in both
metabolism and survival. Thus, activation of Akt, which co-
incides with presence of nutrients and glycolytic metabolism,
suppresses autophagy. Accordingly, M1 macrophages that have
active glycolysis exhibit reduced autophagy (97). In phagocytes,
the mechanisms of autophagy are used for the elimination of
engulfed or intracellular pathogens. The process of elimination
1009
requires formation of active autophagosomes. Formation of
autophagosomes is characterized by the processing of LC3
protein to the LC3-II type and its recruitment on their surface,
termed LC3-associated phagocytosis. LC3-II induction and
autophagosome formation depend on PI3K/Akt/mTOR sig-
nals. In cancer cells, activation of Akt suppresses autophagy and
promotes proliferation and survival. Similarly, in Salmonella
enterica infection, focal adhesion kinase is recruited on the cell
membrane, activates Akt, and inhibits autophagy (98). LPS
stimulation induces expression of GADD34, a sensor of nu-
trient depletion and DNA damage, which is involved in sup-
pressing Akt signaling and autophagy (99). Autophagy is
primarily mediated by signals that activate ULK1/2 and re-
cruitment of Atg and LC3-II proteins on the autophagosomes.
mTORC1 activation suppresses autophagy by blocking ULK1/2
and LC3-II recruitment. Thus, activation of Akt and subsequent
induction of mTORC1 results in inhibition of autophagy. Sig-
nals that activate Akt result in reduced autophagy and pathogen
or apoptotic body clearance. For example, in Pseudomonas-
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infected macrophages, annexin a2 initiates signals that result in
suppression of Akt1 and mTOR-mediated induction of auto-
phagosome formation (79, 100). Thus, activation or inhibition
of Akt can modulate phagocytosis and autophagy, contributing
to pathogen elimination but also to macrophage survival and
inflammatory disease progression.
Contribution of Akt in endotoxin tolerance
Endotoxin tolerance is a state in which cells that have been
previously exposed to Gram-negative bacterial LPS are refrac-
tory to subsequent activation. Endotoxin tolerance is transient
and it protects the organism from excessive inflammation and
endotoxin shock. Endotoxin tolerance is established in several
disease contexts such as sepsis and trauma (101, 102) and the
main cell population that mediates is macrophages (103).
Endotoxin tolerance represents a state of M2-like macro-
phage activation, previously termed as M2c (8, 104). Thus,
endotoxin-tolerant macrophages express genes that are asso-
ciated with tissue repair yet have distinct features from IL-4–
induced or STAT6–driven M2 macrophages because absence
of IL-4Ra or STAT6 does not block endotoxin tolerance (1,
105). The Akt signaling pathway has been shown to play a
significant role in the establishment of endotoxin tolerance in
macrophages. Akt1 is indispensable for the establishment of
endotoxin tolerance as demonstrated in mice lacking Akt1.
Akt1-mediated endotoxin tolerance is mediated by suppres-
sion of miR-155 and subsequent enhancement of its target
suppressor of cytokine signaling 1, and induction of let-7e
followed by reduced expression of its target TLR4 (36). En-
dotoxin tolerance is also regulated by another miRNA, miR-
146a (106, 107). Akt signaling, via Akt1, regulates expression
of miR-146a and miR-155 by mediating signals that allow
colocalization and silencing of one of their genomic loci,
maintaining low levels of expression and suppressing induc-
tion upon a subsequent stimulus (108).
G protein–coupled receptor signaling also contributes to
the induction of endotoxin tolerance. Among Gai proteins the
subunits Gai1 and Gai3 participate in endotoxin tolerance.
Knockdown of Gai1/3 results in impaired activation of in-
flammatory response by LPS, and in macrophages lacking
Gai1/3 Akt is not phosphorylated in response to LPS. Because
Gai1/3 mediate association of Gab1 and the p85 subunit of
1010
PI3K, deletion of this subunit suppresses Akt activation and
promotes M2-like polarization and tolerance to LPS (109).
Whether individual PI3K or Akt isoforms are activated by
Gai1/3 is not known.
The p53 tumor suppressor was recently shown to be in-
volved in M1/M2 polarization and LPS tolerance involving
Akt signaling. Phosphorylation of Akt results in MDM2
activation and p53 inactivation upon M2 polarization of mac-
rophages. Nutlin-3a on the other hand activates p53 in IL-4–
treated macrophages by destabilizing the p53/MDM2 complex,
which results in downregulation of M2 genes. Utilizing this
mechanism, Nutlin-3a delays the establishment of tolerance to
LPS both in vitro and in vivo (110).
Contribution of macrophage Akt in inflammatory diseases
Macrophages are both positive and negative regulators of
inflammatory responses possessing a pivotal role in the de-
velopment of inflammatory diseases. Evidence from models of
disease in rodents as well as from analysis of human specimen,
support the contribution of Akt signaling in inflammatory
disease development and resolution. Rheumatoid arthritis
(RA) is a widely studied Th1 immune response–driven in-
flammatory disease in which macrophages are abundant in the
inflamed synovium. Akt activity in macrophages has been
associated with increased chemotaxis (111). In the mouse
model of collagen-induced arthritis, increased activation of
Akt1 correlates with increased migration in response to che-
motactic factors, an action that can be blocked either by dom-
inant, negative, mutant Akt1 or the PI3K inhibitor LY294002
(112, 113). In human monocyte-differentiated macrophages, the
PI3K/Akt pathway is constitutively active, suggesting that it
contributes to their homeostasis. Indeed, Akt mediates cell sur-
vival, and its activation in macrophages isolated from inflamed
synovium of RA patients was shown to confer resistance to ap-
optosis through upregulation of myeloid cell leukemia sequence
1 (termed MCL1) (114). Inhibition of the PI3K/Akt1 pathway
in synovial macrophages from RA patients leads to apoptotic cell
death (114), supporting the value of the PI3K/Akt pathway as a
therapeutic target in RA.
In another model of Th1-mediated inflammatory disease,
this of experimental autoimmune encephalomyelitis (EAE),
the genetic deletion of Akt3 isoform results in increased CNS
damage. Bone marrow transplantation experiments revealed
the importance of Akt3 in both CNS (microglia) and pe-
ripheral immune cells in the development of the disease (115).
Table II. Involvement of Akt isoforms in inflammatory disease development
Akt Isoform Disease
2/2
Akt3 ↑ EAE
Akt22/2 ↓ DSS-induced colitis
Akt22/2 ↑ Salmonella enterica–induced gastroenterocolitis
Akt12/2 ↑ DSS-induced colitis
Akt12/2ApoE2/2 ↑ Atherosclerosis
Akt12/2 → LDLR2/2 No effect
Akt22/2 → LDLR2/2 ↓ Atherosclerosis
Akt32/2ApoE2/2 ↑ Atherosclerosis
Akt12/2Lyz2-cre ↓ Idiopathic pulmonary fibrosis
Utilization of Akt isoform–deficient mice in models of inflammatory diseases revealed their role in inflammatory disease development. The up arrow indicates augmented disease,
the down arrow indicates suppressed disease, and the dash indicates no mechanism.
BRIEF REVIEWS: Akt SIGNALING IN MACROPHAGES
In the same model, Akt activation in macrophages was asso-
ciated with reduced severity of EAE and M1 markers, an
activation that depended on immunity-related GTPase family
member 1 (termed Irgm1), a protein expressed in M1 mac-
rophages in early EAE (116).
M1 polarization of macrophages contributes to the pa-
thology of intestinal inflammatory diseases. In the model of
dextran sodium sulfate (DSS)-induced colitis, genetic ablation
of Akt1 isoform exacerbated the disease, whereas Akt22/2 mice
were protected. This difference was due to the M1 phenotype
of Akt12/2 and M2 phenotype of Akt22/2 macrophages.
Macrophage depletion and transfer experiments showed that
Akt kinases expressed in additional cell types also contribute to
the pathogenesis of inflammatory bowel disease (30). In a
different model of intestinal inflammation, Salmonella enterica
infection, Akt22/2 mice exhibited severe inflammation in the
intestine and increased bacterial load accompanied by higher
mortality compared with wild-type–infected mice, which was
due to defective bacterial killing (117).
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In noninfectious inflammatory diseases such as idiopathic
pulmonary fibrosis, a condition initiated by extensive tissue
damage, Akt1 is activated in alveolar macrophages and is as-
sociated with increased mitophagy and resistance to apoptosis.
Alveolar macrophages require Akt1 for the production of
mitochondrial reactive oxygen species and secretion of TGF-b,
which promote fibrosis. Deletion of Akt1 in macrophages re-
sults in apoptotic cell death of alveolar macrophages preventing
the onset of pulmonary fibrosis (83).
Tumor-associated macrophages (TAMs) are present in the
tumor stroma and contribute to tumor development and
progression. TAMs possess an M2-like phenotype supporting
tumor growth and metastasis. Efferocytosis of apoptotic tumor cells
polarizes macrophages toward an M2 type via PI3K/Akt activation
(118). ICAM-1 deficiency causes increased efferocytosis through
Akt activation leading to increased metastasis in the liver of a
mouse model of colon cancer liver metastasis (118). Among
Akt isoforms, Akt2 contributes to macrophage polarization
and trafficking toward tumor cells in response to CSF-1, as
demonstrated in coculture experiments of siAkt2-transfected
THP-1 cells and MCF7 breast cancer cells (91). Suppression
of Akt2 expression in THP-1 and mouse peritoneal macro-
phages resulted in reduced macrophage migration (91).
Akt1 regulates miR-155, which in turn appeared to be es-
sential for antitumor macrophage responses (36). Myeloid-
specific deletion of miR-155 impaired M1 activation of
Mechanism Reference
Increased CNS damage, increased inflammation (115)
M2 macrophage polarization (30)
Increased inflammation and reduced ability of killing (117)
bacteria
M1 macrophage polarization (30)
Apoptosis of macrophages and endothelial cells, (126)
accelerated inflammation
— (128)
M2 macrophage polarization, impaired migration (128, 129)
ability, reduced foam cell formation
Foam cell formation (127)
Increased apoptosis of alveolar macrophages (83)
The Journal of Immunology
macrophages and reduced Akt activity leading to accelera-
tion of tumor growth in a model of spontaneous breast
cancer (119). The distinct roles of PI3K and Akt isoforms in
TAMs and antitumor responses is highlighted by the fact
that deletion of p110g (p110g2/2 mice) reduces tumor
growth, angiogenesis and metastasis via blockade of hypoxic
stabilization of hypoxia inducible factor 1a (termed HIF1a)
and decreases production of other proangiogenic factors by
macrophages (120). Myeloid-specific deletion of p110g
suppresses migration of macrophages to the tumor and in-
hibits metastasis (121). Thus, activation of the Akt pathway
can confer protumorigenic effects either by promoting M2
responses in the tumor microenvironment or by support-
ing inflammation (M1 phenotype). Given the opposing
roles of Akt1 and Akt2 in M1 and M2 polarization, Akt
isoform-–specific inhibition is imperative for modulating
antitumor responses. This is supported by the fact that
Akt22/2 mice bearing hepatic deletion of Akt1 develop
hepatocellular carcinoma exhibiting increased infiltration
of Akt1-expressing macrophages (122), potentially with an
M2/TAM phenotype.
Because Akt is a major mediator of insulin signals, it has also
been involved in mediating obesity and type 2 diabetes–related
inflammatory diseases. The defective Akt pathway in macro-
phages is protective against the development of atherosclero-
sis. Mice bearing macrophages that lack insulin receptor and
are apolipoprotein E (ApoE) deficient, have reduced Akt ac-
tivity and develop smaller atherosclerotic lesions after expo-
sure to a high cholesterol diet (123). In a different model of
atherosclerosis, this of low-density lipoprotein receptor
(LDLR) deficiency, transplantation of insulin receptor–
deficient bone marrow cells in LDLR2/2 mice showed decreased
Akt activity in macrophages and enhanced formation of
complex atherosclerotic lesions (124). Isoform-specific dele-
tion of Akt in macrophages highlights their differential role in
M1/M2 polarization through their contribution in athero-
sclerosis. Akt1 deficiency in macrophages in ApoE2/2 mice
resulted in severe coronary atherosclerosis and increased
macrophage and endothelial cell apoptosis in atherosclerotic
lesions, accompanied by increased infiltration of macrophages
and secretion of proinflammatory cytokines. Transplantation
of Akt12/2ApoE2/2 bone marrow to ApoE2/2 mice did not
increase atherosclerosis severity, indicating that macrophages
lacking Akt1 are not sufficient to worsen atherogenesis and
that other cells of vascular origin contribute to the severity of
the disease (125, 126). Akt32/2ApoE2/2 mice developed
more atherosclerotic lesions with increased macrophage in-
filtration, characterized by increased cholesterol accumulation
and foam cell formation (127). In the LDLR2/2 mouse
model of atherosclerosis, Akt2 deficiency in hematopoietic
cells provided protection from the development of athero-
sclerosis. This atheroprotective phenotype could be attributed
to the M2 phenotype of Akt22/2 macrophages leading to
reduced inflammation, CCR2, and subsequent reduced mi-
gration capacity (128) as well as increased cholesterol efflux,
leading to decreased foam cell formation (129). A summary of
the involvement of Akt kinases in inflammatory diseases is
provided in Table II. The underlying mechanisms are com-
plex and Akt kinases are involved in inflammatory diseases not
only through their role in macrophages but also in additional
cell types, rendering them promising therapeutic targets.
1011
FIGURE 2. Akt signaling converges extracellular signals to regulate mac-
rophage biology and M1/M2 polarization.
Downloaded from http://www.jimmunol.org/ by guest on May 19, 2020
Conclusions
Akt signaling converges multiple extracellular and intracellular
signals to regulate macrophage biology that includes pro-
duction of pro- and anti-inflammatory cytokines, phagocy-
tosis, autophagy, and homeostasis as determined by apoptosis
and metabolism. Through the regulation of these functions,
Akt signals determine the functional identity of macrophages,
broadly characterized as M1 and M2 polarization phenotypes
(Fig. 2). How Akt isoforms are regulated in macrophages and
whether their expression changes upon different activation
stimuli or in different subpopulations is not known. More-
over, whether PI3K/Akt isoform–specific interactions and
activation of downstream effectors provide specificity for the
outcome remains to be elucidated.
Disclosures
The authors have no financial conflicts of interest.
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