DOI: 10.1002/JLB.

3MIR0220-270R

SLB MEMBER INSIGHT REVIEW

Complement: Bridging the innate and adaptive immune

systems in sterile inflammation

Martin W. Lo Trent M. Woodruff

School of Biomedical Sciences, Faculty of

Medicine, The University of Queensland,
St Lucia, Brisbane, Queensland, Australia
Correspondence
Prof Trent Woodruff, Laboratory for
Neuroinflammation, School of Biomedical
Sciences, The University of Queensland, St Lucia
4072, Australia.
Email: t.woodruff@uq.edu.au
Abstract
The complement system is a collection of soluble and membrane-bound proteins that together act
as a powerful amplifier of the innate and adaptive immune systems. Although its role in infection is
well established, complement is becoming increasingly recognized as a key contributor to sterile
inflammation, a chronic inflammatory process often associated with noncommunicable diseases.
In this context, damaged tissues release danger signals and trigger complement, which acts on a
range of leukocytes to augment and bridge the innate and adaptive immune systems. Given the
detrimental effect of chronic inflammation, the complement system is therefore well placed as an
anti-inflammatory drug target. In this review, we provide a general outline of the sterile activators,
effectors, and targets of the complement system and a series of examples (i.e., hypertension, can-
cer, allograft transplant rejection, and neuroinflammation) that highlight complement’s ability to
bridge the 2 arms of the immune system.

KEYWORDS
C1q, alloimmunity, hypertension, cancer, anaphylatoxins, neuroinflammation

1 INTRODUCTION 1.2 Danger signals

1.1 Sterile inflammation
Noncommunicable diseases (NCDs) account for 70% of all deaths
worldwide and are a group of chronic conditions that include cardio-
vascular disease, cancer, and neurologic disorders.1 Many are asso-
ciated with powerful sterile inflammatory responses that strive to
remove, contain, and repair dying tissue and are driven by a num-
ber of key biologic processes including necrosis, danger signaling, and
complement activation.2,3 Paradoxically, in the event of excess or pro-
longed inflammation, this well-intentioned response can cause collat-
eral damage and initiate a positive feedback loop that exacerbates the
underlying pathology.3 Thus, sterile inflammation remains an attrac-
tive target for therapeutic intervention in NCDs.
Abbreviations: BBB, blood–brain barrier; DAMPs, damage associated molecular patterns;
DCs, dendritic cells; EAE, experimental autoimmune encephalomyelitis; HMGB1, high
mobility group box 1; IR, ischemia-reperfusion; KO, knockout; MAC, membrane attack
complex; MASPs, MBL-associated serine proteases; MBL, mannose-binding lectin; MDSCs,
myeloid-derived suppressor cells; MS, multiple sclerosis; NCDs, noncommunicable diseases;
ROS, reactive oxygen species; RRMS, relapsing-remitting multiple sclerosis; TILs, tumor
infiltrating lymphocytes; Treg, T regulatory cell; WT, wild-type.
Sterile inflammation can be defined as inflammation in response to
necrotic tissue, which releases danger signals that recruit and acti-
vate the immune system.4 These danger signals or damage-associated
molecular patterns (DAMPs) are ubiquitously expressed molecules
such as ATP, high mobility group box 1 (HMGB1), and hyaluronan,
which under homeostatic conditions are hidden intracellularly or in
hydrophobic regions of the extracellular matrix.3 However, tissue dam-
age exposes these DAMPs, which are detected by a number of widely
expressed receptor classes including: Toll-like, NOD-like, RIG-I-like,
C-type lectin and absence in melanoma 2-like receptors.3 Through
these receptors, DAMPs stimulate monocytes, macrophages, den-
dritic cells (DCs), neutrophils, mast cells, NK cells, and eosinophils
to release proinflammatory mediators, which in turn leads to the
recruitment of inflammatory cells and the activation of the adap-
tive immune response5 Thus, DAMPs are the universal initiators of
sterile inflammation.
1.3 Complement system: Activation pathways
In concert with DAMPs, the complement system is a powerful collec-
tion of over 40 serine proteases, receptors, and regulators that amplify

Received: 5 November 2019 Revised: 7 February 2020 Accepted: 19 February 2020

J Leukoc Biol. 2020;1–13. www.jleukbio.org ○
c 2020 Society for Leukocyte Biology 1
2 LO AND WOODRUFF

C2/C4 MBL/Ficolin/ Prekallikrein

C1q/C1r/C1s Collectin/MASP
FXII
Renin
Classical Mannose- Kallikrien
binding Extrinsic
lectin Thrombin/FXIa/FXa/
C3
Extrinsic FXIa/Plasmin
C2bC4b (C3b)Bd
FXII
Alternative
C3a Altered surface/
C3b Factor B/Factor D/
Properdin
Alternative

C5
C2bC4bC3b (C3b)2Bd

Thrombin/FIXa/FXa/
FXIa/Plasmin

Extrinsic

C5a C5b-9

F I G U R E 1 Complement activation pathways. Inactive molecules and cleavage products at tail and heads of arrows respectively; intersecting
lines denote proteolytic cleavage; pathways are given in bold; and DAMP detection mechanisms are shown in orange

danger signals and augment the inflammatory response. We provide
here a short summary of complement activation and regulation, how-
ever detailed information can be found elsewhere.6-8 These serine pro-
teases are inactive in their native state, but once triggered, undergo
proteolytic cleavage and take part in a number of chain reactions that
are organized into 3 major pathways (Fig. 1), of which the classical and
mannose-binding lectin (MBL) pathways play a key role in danger signal
detection. The classical pathway is canonically triggered by IgM or IgG
immune complexes, which bind to C1q of the C1 complex (C1qC1r2 s2 )
to promote cleavage of C1r2 s2 . This complex then goes on to cleave
C2 and C4 to form the C3 convertase, C2bC4b,9,10 and may also be
triggered by a diverse set of molecules including the DAMP HMGB1,
C-reactive protein, pentraxin 3, and apoptotic cell membranes.11
Homologous to the classical pathway, the MBL pathway is activated by
carbohydrate moieties found on apoptotic and necrotic cells.12 These
sugars are detected by a number of proteins including MBL, collectin-
10/11, and ficolin H/L/M,13 which go on to activate the 3 MBL-
associated serine proteases (MASPs). MASP-1 is a C2 convertase that
can also cleave MASP-2, which is a C2 and C4 convertase, and MASP-3
can cleave factor D to activate the downstream alternative pathway.13
The MBL pathway can also be triggered by the DAMPs HMGB1,14
ATP,15 and S100a914 ; C-reactive protein; and pentraxin 3.13 Thus, the
classical and MBL pathways are powerful sensors for danger signals,
dying cells and acute phase reactants.
Further downstream, the alternative pathway is an amplification
loop that can contribute up to 80% of complement activation from
upstream pathways, and in itself can be triggered by “altered” sur-
faces such as damaged or foreign tissue.16 Unlike the other systems,
the alternative pathway undergoes autoactivation in which C3 spon-
taneously hydrolyses to form C3(H2O) and binds to factor B.17 Fac-
tor D is then able to cleave factor B into Ba and Bb, of which the
latter remains associated with the complex (“initiation” C3 conver-
tase). Subsequent cleavage of C3 into C3a and C3b allows the forma-
tion of C3bBb (“amplification” C3 convertase) and C3bBb and C2bC4b
from the upstream pathway can incorporate additional C3b to adopt
C5 convertase activity. Conversion of C5 to C5a and C5b then facili-
tates the formation of C5b9 or the membrane attack complex (MAC).18
C3a, C3b, C5a, and MAC are potent inflammatory mediators and thus
the pathways downstream of C3 are tightly regulated by a range of
inhibitors including factor I, carboxypeptidase N, and C4BP19 ; which
interestingly can activate B cells and promote IgE isotype switching.20
Damaged tissues often lack these inhibitors and can thus trigger the
alternative pathway, a process further promoted by properdin, which
stabilizes C3bBb and is secreted by activated neutrophils.17,21 Super-
imposed over these 3 canonically defined cascades, the extrinsic path-
way (Fig. 1) is a more recently recognized collection of proteases that
have isolated convertase activity. Many of these are derived from
the cardiovascular system and include factor IXa, Xa, XIa, throm-
bin, and plasmin, which are C3 and C5 convertases22 ; renin and
kallikrein, which cleave C323,24 ; and Hageman factor which cleaves
the C1 complex.25 Thus, a host of sterile stimuli can trigger the
complement system.
LO AND WOODRUFF 3

A C1q–antigen signalling B C1q–antigen–apoptotic body signalling

C1q–antigen complex C1q–antigen–apoptotic body
complex
cC1qR–
SIGNR1
IL-12 IL-12R IL-10 IL-10R1/2
? ?
STAT3/4 ?
NF-KB MAPK Th1 Th1/17
cC1qR–LRP
effector (CD91) effector
PI3K
status status
IL-12 & TNFa IL-10
TRAFs
TNFa TNFRs

Dendritic cell Naive T cell Dendritic cell Naive T cell

C C1q–antigen–apoptotic body signalling D B cell CR2 signalling

B cell receptor
IL-27 IL-27R
? PI3K
STAT1 CR3 (CD11b+CD18)-
Vav Memory B/
cC1qR–LRP iC3b-antigen
Plasma cell
(CD91) Th1/17
status
effector
status
IL-27
B cell
coreceptor
CR2 (CD21)-CD19

Macrophage Naive T cell Follicular Naive B cell

dendritic cell

E Autocrine and paracrine C3a/C5a signalling

Proteolysis C3 C5 Proteolysis
TLR FB FD C3a
C5a
C3b
C3a
C3 C5
C5a C3 C3 FB FD
C3b FB FD

T effector
NF-KB
status
Cytokine Lysosome
expression
Nucleus
MHC-TCR and
B7-CD28
complex

Dendritic cell Naive T cell

F I G U R E 2 Mechanisms by which complement bridges the innate and adaptive immune system. (A) C1q-opsonized antigen activates an
unknown pathway in DCs, potentially via cC1qR–SIGNR1, that triggers NF-𝜅B nuclear translocation and up-regulates proinflammatory cytokine
IL-12 and TNF𝛼 production. These cytokines then stimulate their respective receptors on T cells to promote Th1 polarization. (B) C1q-opsonized
antigen with apoptotic cells are phagocytosed by DCs, potentially via the cC1qR–LRP receptor complex. This phagocytosis then promotes immuno-
suppressive cytokine IL-10 production through currently unknown mechanisms and IL-10 then down-regulates Th1/17 induction in naive T cells.
(C) Similarly, phagocytosis of C1q-opsonized antigen with apoptotic cells in macrophages promotes the production of IL-27, which inhibits Th1/17
polarization in naïve T cells. (D) CR2 signaling at the APC-B cell immune synapse lowers the threshold for B cell activation and promotes germinal
center formation. (E) APC-T cell interaction triggers auto- and paracrine C3a/C3b/C5a signaling that drives effector T cell differentiation and DC
cytokine production. Stimulation is represented by normal arrowheads and inhibition represented by blunt arrowheads

1.4 Complement system: Effector molecules phagocytes to tailor the inflammatory response to its antigen.19 When
bound to “regular” antigen, it acts as a proinflammatory mediator and
The major effector fragments of the complement system including
encourages DCs to present antigen, migrate to lymphoid tissue and
C1q, C3a, C3b, C5a, and C5b9/MAC are potent molecules that coor-
secrete Th1 polarizing cytokines such as IL-12 and TNF𝛼 26-29 (Fig. 2A);
dinate and link the innate and adaptive immune systems (Fig. 2). C1q
but when bound to apoptotic cells, C1q stimulates DCs to produce IL-
is an opsonin that is thought to signal through C1qR and SIGNR1 on
4 LO AND WOODRUFF
10, which suppresses Th1 and Th17 proliferation as a means of lim-
iting inflammation during apoptosis (Fig. 2B), a phenomenon that is
mirrored in macrophages (Fig. 2C).30-33 C3b and its proteolytic prod-
ucts iC3b, C3c, C3dg, and C3d are also opsonins of antigens and
apoptotic cells, but bind to CR1/CD35 (C3b and iC3b), CR2/CD21
(iC3b and C3dg), CR3/CD11b+CD18 (iC3b, C3dg, and C3d), and
CR4/CD11c+CD18 (iC3b); which are differentially expressed amongst
leukocytes but are all considered to be involved in phagocytosis and
antigen presentation.19,34 In particular, CR2 has a unique role in
humoral immunity (Fig. 2D). As part of the B cell coreceptor complex,
it can lower the threshold for B cell activation, prolong antigen dis-
play in germinal centers, and facilitate firm contact between follicular
DCs and B cells to initiate germinal center reactions and the forma-
tion of plasma and memory B cells.35,36 By contrast, CR3 acts in a sim-
ilar fashion to C1qR in that its ligation by opsonized apoptotic bodies
can reduce DC immunostimulatory function by reducing cytokine pro-
duction (e.g., IL-12).37,38 Thus, complement factors from the upstream
pathways act as dynamic opsonins that influence both the innate and
adaptive immune systems.
Further downstream, C3a and C5a are potent and versatile
complement peptides or “anaphylatoxins” that signal through C3aR
and C5aR1/2, respectively.39,40 In innate immunity, both have well
defined roles and multiple leukocyte targets in which 1 or both can
promote or inhibit chemotaxis, granulocyte degranulation, phago-
cytosis, respiratory burst formation, inflammasome activation, and
cytokine and eicosanoid production,41-44 though the function of
C5aR2 is controversial and has been reviewed elsewhere.39 In addi-
tion, C3a and C5a have been increasingly recognized as regula-
tors of the adaptive immune response. Both are produced at the
APC-T cell interface to promote T cell activation and APC mat-
uration (Fig. 2E) and production from intracellular stores known
as the “complosome” with C3b signaling through CD46 are nec-
essary for T cell survival, activation, and differentiation, in which
the anaphylatoxins and C3b cause polarization away from and
towards the T regulatory (Treg) phenotype respectively.45 Lastly,
C5b-9/MAC is a toroidal pore that depending on its form can
induce cell lysis and apoptosis, cytokine production,46 inflammasome
activation,47 and gene expression.48 Thus, the complement system is
a diverse group of molecules, each with a repertoire of innate and
adaptive capabilities.
2 HYPERTENSION
Hypertension is the leading global burden of disease risk factor and can
be defined as systolic and/or diastolic blood pressure above 130 and
80 mmHg, respectively.49,50
Mostly an idiopathic phenomenon, hypertension is driven by renal
sodium dyshomeostasis and reduced arterial compliance.50 The for-
mer is often caused by lifestyle factors and comorbidities, whereas the
latter is due to endothelial dysfunction, blood vessel fibrosis, and vas-
cular smooth muscle cell hyperplasia.51 Sterile inflammation in hyper-
tension is elicited by hypertensive stimuli that directly or indirectly
activate the immune system and affects both the vascular system and
target organs.52 Cytokines produced during inflammation promote
the formation of a neointima (an intimal thickening), which increases
vascular resistance and blood pressure, and may also increase renal
tubular production of angiotensin and salt and water retention, which
both increase blood volume and intravascular pressure.53 Additionally,
matrix metalloproteinases secreted by a variety of cells can degrade
the extracellular matrix of the vessel wall and allow immune cells to
infiltrate affected organs and inflict damage by promoting apoptosis
and enhancing collagen synthesis and matrix deposition.53
Regulating these processes, Treg cells are a specialized subpop-
ulation of T cells that suppress the immune response to maintain
homeostasis and self-tolerance. In an angiotensin II (Ang II)-induced
hypertensive mouse model, FOXP3+ Tregs that are known to express
the Ang II receptor type 1 exhibited increased C3aR and C5aR1
expression,54,55 which would imply that Ang II is able to prime CD4+
T cells for differentiation away from a FOXP3+ Treg phenotype.56 In
support of this, C3aR–C5aR1 double knockout (DKO) mice that under-
went Ang II hypertensive treatment had reduced CD4+ and CD8+
T cells and a mitigated decrease in FOXP3+ Tregs in their blood and
kidneys compared to wild-type (WT) control.54 DKO mice that under-
went Ang II hypertensive treatment also had reduced systolic and
diastolic blood pressure compared to WT mice and depletion of Tregs
abolished the protective effects against Ang II-induced hypertension
and target organ damage in DKO mice.54 Thus, at least in mouse
models, C3aR and C5aR exert a hypertensive effect in Ang II-induced
hypertension mediated by a down-regulation of the protective effects
conferred by Tregs (Fig. 3A). In addition, hypertension in humans is
associated with elevated plasma C3a and C5a concentrations and
reduced FOXP3+ Treg counts54,57 and thus, an analogous mechanism
may occur in humans. However, although this antihypertensive effect
is linked to anti-inflammatory cytokine production (e.g., IL-10 and TGF-
𝛽) and reduced blood vessel infiltration by macrophages, its full nature
is incompletely understood.58,59 This and other emerging studies sug-
gest complement is a key regulator of the innate and adaptive immune
system in hypertension.57,60–62
3 CANCER
Cancer is expected to rank as the leading cause of death and the sin-
gle most important barrier to increasing life expectancy in the world
in the 21st century.63 Though there are over 100 distinct types of
cancer, all tumors have defects in regulatory circuits that govern nor-
mal cell proliferation and homeostasis and to varying degrees are
characterized by the 8 hallmarks: (1) evasion of apoptosis, (2) self-
sufficiency in growth signals, (3) insensitivity to anti-growth signals,
(4) sustained angiogenesis, (5) limitless replicative potential, (6) tis-
sue invasion and metastasis, (7) reprogramming of energy metabolism,
and (8) evading immune destruction.64,65 In particular, cancer is associ-
ated with a distinct form of chronic inflammation that both predisposes
to cancer development (15–20% of all cancer cases) and advances
established malignancies.66,67 Cancer-associated inflammation has a
LO AND WOODRUFF 5

C3a IL-10
C3aR
A Hypertension B Tumor
CD8+ TIL

ANG C3a IL-10R

TGF-B Tumour
Cytolytic 1/2
production
AT1R C5a Tumor function
of C5a IL-10
Treg cell
effector
FOXP3
status C5aR1 (CD88) C5a
IL-10
C5a
C5a
C5a
Macrophage C5a
TAM Treg
vascular wall infiltration
effector
status
MDSC
Endothelial-dependent MDSC MDSC CD4+ TIL
vascular smooth muscle relaxation chemotaxis differentiation IL-10R1/2
into TAM and
TGF-BR
IL-10 and TGF-B
C Allograft transplant
Tumour evasion of
immunosurveillance
Host APC
Host APC migration in
donor antigen
afferent lymphatics Host APC
uptake

Immune synapse with

autocrine and paracrine
C3a and C5a
T cell migration in signalling
efferent blood vessel

Cytokine
production
Effector T cell
CD4+/CD8+ differentiation
T cell and proliferation
alloimmunity
Transplant tissue Lymph node

D Multiple sclerosis
CR4
CR3 (CD11c+CD18)
(CD11b+CD18)
?
Neuron
T cell T cell, B cell,
activation microglia
T cell autoimmunity

T cell CD46
receptor

Treg
effector
Immunosupression Oligodendrocyte
status

Autoimmune attack
T cells

F I G U R E 3 Complement in sterile inflammatory diseases. (A) In hypertension, C5aR1 and C3aR signalling inhibits Treg polarization and thus
down-regulates their anti-inflammatory and vasodilatory effects, of which only known mechanisms are shown above. (B) In tumorigenesis, C5a
acts as a chemokine for myeloid-derived suppressor cells (MDSCs), which differentiate into tumor-associated macrophages (TAMs) that produce
immunosuppressive and Treg polarizing cytokines. Additionally, autocrine C3a signaling in CD8+ tumor-infiltrating lymphocytes (TILs) inhibits
autocrine IL-10 signaling, which normally promotes cytolytic activity. (C) In allograft transplant rejection, indirect allorecognition is mediated by
host APCs that travel to the lymphatic system to interact with naive T cells. These T cells then form immune synapses and with the assistance of
autocrine and paracrine C3a and C5a signaling, proliferate and differentiate into effector T cells that return to the allograft to promote inflam-
mation and chronic rejection. (D) In MS, CR3 and CR4 drive T cell activation and autoimmune attack and defective CD46 O-glycosylation and
cosignaling with the T cell receptor impairs Treg effector status and IL-10 production
6 LO AND WOODRUFF
wide range of stimuli including gene mutations that promote the pri-
mary production of cytokines,68 infective agents (e.g., human papil-
loma virus in cervical cancer), and cell death secondary to chemo-
/radiotherapy, hypoxia, metabolic stress, or mutagenic insult.69 More-
over, unlike inflammation linked to infection, in which sterilizing immu-
nity is able to eradicate a pathogen and promote tissue repair, cancer-
associated inflammation is usually unable to completely remove all
malignant cells, which is in no small part due to their ability to evade the
immune system. This results in the continued release of danger signals
and positive feedback amplification that leads to the accumulation of
growth-promoting cytokines.69 Thus, inflammation in cancer is a com-
plex process in which malignant cells seek to evade and exploit the
immune system.
Complement has a diverse set of functions in the context of
cancer including directly promoting proliferation and metastasis,
stimulating angiogenesis, and regulating the immunosuppressive
microenvironment.70-72 In the latter, the complement system acts as
a bridge between innate and adaptive immune systems and plays a key
role in recruiting myeloid-derived suppressor cells (MDSCs) and mod-
ulating T-cell-mediated antitumor immunity.
Myeloid-derived suppressor cells are a diverse group of imma-
ture, immunosuppressive, and pathologically active leukocytes that are
the result of persistent myelopoiesis during chronic inflammation.73
To varying degrees, these cells secrete immunosuppressive cytokines
and eicosanoids, produce reactive oxygen species (ROS), facilitate
Treg recruitment, and rapidly differentiate into tumor-associated
macrophages that persist in the tumor microenvironment.74 Com-
plement peptide C5a acts as a chemokine for MDSCs and is pro-
duced by some cancers to facilitate their recruitment (Fig. 3B).71,75–77
Non-small cell lung cancer patients have elevated plasma C5a lev-
els and lung cancer cell lines are able to produce C5a in serum free
conditions.76 Moreover, C5aR1 antagonism and/or genetic knockout
(KO) in a lung cancer mouse model reduced tumor growth, MDSC
recruitment, and tumor expression of the immunomodulators ARG1,
CTLA-4, IL-6, IL-10, LAG3, and PDL1, which are associated with Treg
induction.76,78 Similarly, in a preclinical mouse model for breast can-
cer, C5a suppressed CD8+ and CD4+ T cell responses in lung tis-
sue to create a metastatic niche.79 In this instance, pharmacologic
blockade and genetic ablation of C5aR1 was sufficient to reduce
lung metastases with a concomitant increase in CD4+ and CD8+
T cell recruitment and a decrease in MDSC infiltration and IL-10
and TGF-𝛽 cytokine production.79 This suppression of T cell immuno-
surveillance is mirrored in an autocrine complement mechanism, in
which autocrine C3a suppresses the expression of IL-10, a potent
activator of CD8+ tumor-infiltrating lymphocytes (TILs) (Fig. 3B). In
this context, C3-deficient mice are resistant to tumor development
in a T cell- and IL-10-dependent manner and produce CD8+ TILs
that have enhanced effector function.80 By contrast, C3aR KO mice
with B16F0 melanoma had reduced CD4+ T lymphocyte infiltration
compared with their WT littermates due to a neutrophil-dependent
mechanism, which highlights the complex relationship between com-
plement and cancer immunity.81 Thus, the complement system is
a regulator of both the innate and adaptive immune systems with
direct effects on tumor progression and is a promising target for
therapeutic intervention.82
4 ALLOGRAFT TRANSPLANT REJECTION
Allograft organ transplants are transplantations to a recipient from a
genetically nonidentical donor of the same species and has emerged
as a viable therapeutic modality for a variety of ailments.83 Organs
including kidneys, livers, and hearts can be transplanted with impres-
sive short-term survival rates, but allograft rejection of which there are
3 classes (hyperacute, acute, and chronic) remains as a limiting factor to
long-term survival.84 Hyperacute rejection occurs in the first few min-
utes following transplantation and is triggered by recipient antidonor
antibodies that via the complement and coagulation cascades cause
vessel thrombosis and secondary infarction. In modern times, this type
of rejection is a rarity as a result of ABO compatibility checks and
human leukocyte antigen crossmatching.85 By contrast, acute rejec-
tion occurs between 1 week and several months after transplantation
and arises as a result of intense cellular and humoral attack, but with
current immunosuppressive regimens occurs in less than 15% of non-
sensitized transplant patients. Chronic rejection however occurs after
months to years and is mediated by insidious cellular and humoral pro-
cesses that cause obliterative vasculopathy, parenchymal fibrosis, and
tertiary lymphoid organ formation.85,86 This type of rejection responds
poorly to current immunosuppressive regimens and is now the leading
cause of graft rejection.87,88 Transplant organs are scarce and valuable
and thus development of novel therapeutics against acute and chronic
rejection is needed to maximize their utility.89
Whether a transplant undergoes acute or chronic rejection is
thought to depend in part on the nature of the recipient’s immune
response to donor antigens. It is hypothesized that host exposure to
intact MHC alloantigen displayed on donor APCs (direct allorecog-
nition) results in acute rejection because of substantial expansion of
T cells of multiple specificities.86 By contrast, host exposure to donor
alloantigen processed and presented by host APCs (indirect allorecog-
nition) leads to activation of a limited repertoire of T cells with a
restricted ability to recognize graft targets, but importantly act as
helper T cells for B cells that produce alloantibodies, and thus causes
chronic rejection instead.90-93 Other factors are also likely to have
a role in determining the nature of rejection and include ischemia-
reperfusion (IR) injury, which enhances effector T cell accumulation in
allografts.86 Thus, the APC-T cell relationship is not only a vital link
between the innate and adaptive immune system but also a key deter-
minant of transplant alloimmunity.
In the context of an allograft transplant, the complement system
is activated by IR injury and acts as a bridge between the innate
and adaptive immune systems. During the transplantation procedure,
interim cold ischemic storage causes hypoxia, ATP depletion and mito-
chondrial damage within the graft and subsequent reperfusion para-
doxically causes the formation of ROS, which further damages the
graft and promotes graft dysfunction in a process known as IR injury.
Injured tissue then releases neoantigens and DAMPs that activate the
LO AND WOODRUFF
complement system and initiate a sterile inflammatory response.94-98
This initial insult is then also propagated by complement, which acts
as a potent modulator of cellular and humoral immunity. In the former,
autocrine and paracrine C3aR and C5aR1 signaling at the APC-T cell
interface amplifies T cell effector responses and in numerous mouse
models has been shown to accelerate rejection (Fig. 3C).99 For exam-
ple, anti-C5,100 C5aR1 antagonism,101 and C3aR1 KO grafts102 reduce
T cell priming and improve allograft survival in cardiac and renal trans-
plant mouse models, whereas genetic ablation of decay accelerating
factor, which normally inhibits C3a and C5a production,103 augments
T cell immunity and accelerates rejection in murine cardiac grafts.104 In
addition, C3aR and C5aR1 signaling inhibits Treg polarization,56,105,106
and thus C5aR2, which in T cells does not interact with 𝛽-arrestin or
inhibit ERK1/2 signaling, can regulate T cell effector function by act-
ing as a decoy receptor.107 Indeed, in a Treg-dependent cardiac trans-
plant mouse model, absence of recipient C5aR2 accelerated rejection,
whereas C5aR2 overexpression prolonged graft survival.107 Interest-
ingly, like in cancer, complement also has an immunosuppressive effect
mediated by MDSCs, as seen in myeloid cell-specific C5aR1 condi-
tional KO mice with cardiac allografts, which have increased donor
T cell immunity, decreased Treg expansion and accelerated graft rejec-
tion despite costimulatory blockade.108 Additionally, complement has
an important role in humoral transplant alloimmunity, in which CR2 is
expressed on B and follicular DCs and recognizes the opsonins iC3b,
C3dg, and C3d.19 When activated, this receptor lowers the thresh-
old for B cell activation and promotes antigen retention by follicular
DCs,109,110 a process that is reflected in C3-deficient mice that fail to
produce high affinity IgG responses against major histocompatibility
antigens in skin grafts.111 Thus, the complement system is a power-
ful amplifier of sterile inflammation in allograft transplantation and is
currently being pursued in phase II/III clinical trials as a drug target for
acute transplant rejection.82
5 NEUROINFLAMMATION
Neurological disorders are the leading cause of disability-adjusted
life years and the second-leading-cause of death.112 Bar those con-
ditions with a relatively clear and isolated etiology and pathogen-
esis (e.g., extradural hematoma), this diverse group of conditions is
typically challenging from an interventional and drug development
standpoint, not least because of their profound clinicopathologic vari-
ability. Though typically defined by a small number of key features,
neurologic diseases tend to each contain distinct subgroups or a wide
spectrum of patients with varying symptoms, progressions, etiolo-
gies, and pathologic processes.113-117 Moreover, the CNS is particu-
larly sensitive to insult. As a hierarchical organ,118 the topology of its
neuronal connections is critical to its overall function and recapitu-
lating this during a repair process remains difficult.119 Lesions in the
CNS also tend to be propagated by excitotoxicity, an intrinsic neuronal
property mediated by the neurotransmitter glutamate.120 Found at
60–70% of all synapses in the CNS, this excitatory neurochemical is
released in excess upon neuronal breakdown and stimulates neighbor-
7
ing AMPA and NMDA receptors, the latter of which facilitates Ca2+
entry into the cytoplasm and drives cell death by activating proteases
(e.g., calpains) and lipases and increasing the production of ROS and
arachidonic acid.121,122 In severe cases, this can result in necrosis,
which triggers a self-perpetuating cycle of glutamate release and exci-
totoxicity , as seen in the expanding umbra of an ischemic stroke.123
From a drug development standpoint, these pathologic phenomena
necessitate a drug target that is ubiquitous, promptly expressed, and
highly efficacious; characteristics that are found in neuroinflammation.
Neuroinflammation is a broad term that encompasses a range of
context-driven inflammatory processes within the brain and spinal
cord that are distinct from “classic” sterile inflammation. These
processes can be grouped into 2 distinct categories, denoted here
as “primary” and “secondary” inflammation.124 “Primary” neuroin-
flammation refers to that in autoimmune diseases (e.g., multiple
sclerosis [MS]) and is associated with significant peripheral leukocyte
invasion and a strong adaptive immune response characterized by
heightened MHC antigen presentation and T cell co-stimulation.124
In contrast, “secondary” neuroinflammation refers to that in response
to primary neuropathology (e.g., neurodegeneration, cancer, and
epilepsy) and is a glial-driven process with comparatively low
chemokine production and T cell stimulatory capacity.124 Despite
these differences, neuroinflammation can be defined by a number of
standard features including macrophage/microglial activation and a
suite of cytokines (IL-1𝛽, IL-6, and TNF𝛼), chemokines (CCL2, CCL5,
and CXCL1), secondary messengers (NO and prostaglandins), and
ROS.125 These immune mediators may be neuroprotective (e.g., in
synaptic pruning, immune surveillance, neurotrophin production, and
euflammation) or neurotoxic (e.g., via cytotoxicity and excitotoxicity)
and the balance of their functions ultimately drives the overarching
effect of the inflammatory response.125,126 In line with this, neuroin-
flammation commonly has a biphasic nature characterized by an initial
neuroprotective reactive gliosis that may either resolve, or become
overwhelmed and transition into a self-perpetuating proinflammatory
reaction,127 the latter of which reflects the tendency of microglia to
become hyperactive in response to repeated immune stimulation.125
Many of these processes have been correlated with disease progres-
sion and their genetic and pharmacologic modulation have been able
to improve outcomes in animal models.43,128–139 Thus, neuroinflam-
mation has become an attractive drug target for neurologic disease
and is currently being pursued in phase I/II/III clinical trials.140-142
The complement system is a key driver of neuroinflammation and
is a promising therapeutic target given its universality, potency, and
timely reactivity. In neurologic conditions, the nature of complement
activation is heavily dependent on the permeability of the blood–brain
barrier (BBB). In response to minor insults, the BBB generally remains
intact and complement may be synthesized locally by astrocytes (C1,
C2, C3, C4, C5, C6, C7, C8, C9, FB),143,144 microglia (C1, C3, C4),143
neurons (C5),145 and oligodendrocytes (C1, C2, C3, C4, C5, C6, C7, C8
C9)146 at relatively low (nanomolar) concentrations over a period of
days, weeks and even months.147 By contrast, in response to severe
(e.g., neurotrauma) or chronic insults, the BBB may lose its integrity
and permit systemic and leukocyte-synthesized complement to enter
8 LO AND WOODRUFF
the CNS and reach relatively high (micromolar) concentrations in a rel-
atively short amount of time (24 h).147 In addition, the complement sys-
tem has a range of functions specific to the nervous system. In both
development and disease, complement opsonins (i.e., C1q and C3b)
facilitate microglia-mediated synaptic pruning and phagocytosis of cel-
lular debris148-151 and C1q can affect microglial cytokine production
in a similar manner to that in DCs whereby apoptotic blebs modulate
the response, although whether or not microglia interact with T cells
in this manner is not known.152,153 Likewise, anaphylatoxins (i.e., C3a
and C5a) have a number of canonical effects on glial cells including
driving chemotaxis,154,155 promoting cytokine production,156,157 and
activating the inflammasome,158,159 but may also have a number of
direct effects on neurons including limiting excitotoxity,160 promot-
ing neurogenesis,161 and driving neuronal stem cell proliferation in
brain cancer.162 Such diverse capabilities are also exhibited by MAC,
which has canonical cytolytic effects that are driven in autoimmune
conditions by autoantibody activation of the classical pathway163-166
and immunomodulatory and protective effects on glia.167,168 These
immune pathways occur in an environment with relatively low comple-
ment regulator expression and thus complement is a powerful driver of
neuroinflammation.169-172
Unlike its effect on the innate immune response, complement’s
effect on the adaptive immune response in neuroinflammation is rel-
atively under-investigated. Nevertheless, emerging studies give some
insight into its potential role. MS is an autoimmune neurologic dis-
order characterized by the intermittent and progressive presentation
of a wide range of neurologic deficits.173 The condition is tradition-
ally viewed as a 2-stage disease with early inflammation responsible
for relapsing-remitting disease (RRMS) and delayed neurodegenera-
tion causing nonrelapsing progression, that is, secondary and primary
progressive MS.173 In the both instances, patients develop inflamma-
tory plaques that cause demyelination and eventually axonal damage.
However, in RRMS, the “classical active lesion” predominates and is
associated with profound T cell (mainly MHC class I restricted CD8+
T cells), B cell, and plasma cell infiltrates, whereas in progressive dis-
ease, lesions tend to have an inactive core surrounded by a narrow
rim of activated microglia and macrophages.173,174 In relation to the
former, CR3 and CR4 appear to play a critical role in T cell-mediated
disease. In the experimental autoimmune encephalomyelitis (EAE) MS
mouse model, CR3 and CR4 KO mice exhibited reduced spinal cord
T cell infiltration and IFN𝛾 production by these cells (Fig. 3D).175,176
Additionally, CR3 and CR4 KO T cells have an unusual cytokine pro-
file and limited EAE-inducing capacity when transferred to control
mice,175,176 though the signaling pathways that underlie these phe-
nomena are incompletely understood.177 The impact of complement
on MS adaptive immunity can also be seen in Tregs in which isolated
CD46 stimulation of these cells results in impaired IL-10 production
and reduced immunosuppression.178 Indeed, MS T cells have defec-
tive T cell receptors that are unable to glycosylate CD46 for recruit-
ment to the immune synapse, which is thought to drive Treg induction
(Fig. 3D).179 These findings suggest that complement plays a key role in
bridging the innate and adaptive immune system in neuroinflammation
and flag this area for future research.
6 CONCLUSION
The rates of NCDs continue to increase globally and with it a greater
need for disease modifying treatments. In these conditions, a variety of
danger signals trigger a sterile inflammatory response that chronically
exacerbates the underlying pathology, in which the complement sys-
tem acts as a potent amplifier with a wide range of innate and adaptive
leukocyte targets. In hypertension, studies indicate an intimate rela-
tionship between the cardiovascular and immune systems, in which
angiotensin II may act directly on T cells. Here, complement acts as
an autocrine T cell activator that increases blood pressure, promotes
leukocyte tissue invasion and exacerbates end organ damage. By con-
trast, cancer cells exploit complement as a means of avoiding immuno-
surveillance, in which C5a is a salient chemokine for MDSC recruit-
ment to malignancies. These dysregulated leukocytes secrete anti-
inflammatory and Treg polarizing cytokines that allow for unchecked
neoplastic proliferation. Likewise, this cellular immune response is
seen in transplant alloimmunity, in which complement activates T lym-
phocytes at the APC–T cell interface and promotes the production
of alloreactive antibodies, and in MS, in which CR3/4 and dysregu-
lated CD46 signaling influence T cell activation. Thus, complement is
a bridge between the 2 arms of the immune system and at this nexus is
well placed for drug development against chronic inflammation.
AUTHORSHIP
M.W.L. and T.M.W. coconceived the review. M.W.L. drafted the
manuscript and created the figures, with T.M.W. contributing to edit-
ing and revisions.
ACKNOWLEDGEMENTS
T.M.W. acknowledges support from the National Health and Medical
Research Council (NHMRC fellowship APP1105420).
DISCLOSURES
The authors declare no conflicts of interests.
ORCID
Martin W. Lo https://orcid.org/0000-0002-2736-5507
Trent M. Woodruff https://orcid.org/0000-0003-1382-911X
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