Article

Complement System Activation in Cardiac and Skeletal Muscle Pathology: Friend or Foe?

Center of Basic Research, Biomedical Research Foundation, Academy of Athens, Athens, 11527, Greece.
Advances in Experimental Medicine and Biology (Impact Factor: 1.96). 02/2013; 734(VIII):207-18. DOI: 10.1007/978-1-4614-4118-2_14

ABSTRACT

A major goal in current cardiology practice is to determine optimal strategies for minimizing myocardial necrosis and optimizing cardiac repair following an acute myocardial infarction. Temporally regulated activation and suppression of innate immunity may be critical for achieving this goal. Extensive experimental data in various animal models have indicated that inhibiting complement activation offers protection to cardiac tissue after ischemia/reperfusion. However, the results of clinical studies using complement inhibitors (mainly at the C5 level) in patients with acute myocardial infarction have largely been disappointing.
In cases in which complement activation participates in the initial events of muscle cell destruction, as in autoimmune myocarditis or autoimmune muscle disorders, inhibition of complement activation is expected to prove a successful treatment. In other pathologic conditions in which complement is recruited by degenerating or dying muscle cells, as in ischemia, the ideal approach is probably to modulate rather than abruptly blunt complement activation. Beneficial effects of complement action with regard to waste disposal, recruitment of stem cells, regeneration, angiogenesis, and better utilization of energy sources under hypoxic conditions may also prove important for successful disease treatment. Patient outcome after myocardial infarction almost certainly depend upon the combined activation of several distinct but potentially interrelated signaling pathways, suggesting that a combination of treatments targeted to different pathways should be the therapy of choice, and modulation of complement could be one of them.

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Available from: Manolis Mavroidis, Feb 09, 2016
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J.D. Lambris et al. (eds.), Complement Therapeutics, Advances in Experimental Medicine and Biology 735,
DOI 10.1007/978-1-4614-4118-2_14, © Springer Science+Business Media New York 2013
M. Syriga M. Mavroidis , Ph.D. (*)
Center of Basic Research , Biomedical Research Foundation, Academy of Athens , Athens 11527 , Greece
e-mail: emavroeid@bioacademy.gr
Abstract A major goal in current cardiology practice is to determine optimal strategies for minimizing
myocardial necrosis and optimizing cardiac repair following an acute myocardial infarction.
Temporally regulated activation and suppression of innate immunity may be critical for achieving this
goal. Extensive experimental data in various animal models have indicated that inhibiting comple-
ment activation offers protection to cardiac tissue after ischemia/reperfusion. However, the results of
clinical studies using complement inhibitors (mainly at the C5 level) in patients with acute myocardial
infarction have largely been disappointing.
In cases in which complement activation participates in the initial events of muscle cell destruc-
tion, as in autoimmune myocarditis or autoimmune muscle disorders, inhibition of complement acti-
vation is expected to prove a successful treatment. In other pathologic conditions in which complement
is recruited by degenerating or dying muscle cells, as in ischemia, the ideal approach is probably to
modulate rather than abruptly blunt complement activation. Bene cial effects of complement action
with regard to waste disposal, recruitment of stem cells, regeneration, angiogenesis, and better utiliza-
tion of energy sources under hypoxic conditions may also prove important for successful disease
treatment. Patient outcome after myocardial infarction almost certainly depend upon the combined
activation of several distinct but potentially interrelated signaling pathways, suggesting that a combi-
nation of treatments targeted to different pathways should be the therapy of choice, and modulation of
complement could be one of them.
14.1 Complement Activation in Cardiac Muscle
14.1.1 Myocardial Infarction and Innate Immunity
Myocardial infarction is the most frequent cardiovascular event in the western world and is respon-
sible for a large fraction of all cardiovascular deaths. Improved clinical management of acute myocar-
dial infarction has signi cantly lowered immediate mortality over the past two decades (Lloyd-Jones
et al. 2009 ) , but surviving patients still face another major complication: the development of heart
failure resulting in part from inadequate healing of their original infarct. Heart failure is a major health
problem that has reached almost epidemic proportions in the U.S., in that it affects 2% of the American
Chapter 14
Complement System Activation in Cardiac and Skeletal
Muscle Pathology: Friend or Foe?
Maro Syriga and Manolis Mavroidis
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208 M. Syriga and M. Mavroidis
population. It is also an economic problem: The cost of hospitalizations related to heart failure is
currently twice that for all forms of cancer and myocardial infarctions combined. Therefore, a major
goal in current cardiology practice is to determine optimal strategies for minimizing myocardial
necrosis and optimizing cardiac repair following acute myocardial infarctions.
Myocardial infarction results in decreased oxygen tension within cardiomyocytes and a subse-
quent loss of oxidative phosphorylation and decreased generation of high-energy phosphates. The
mammalian heart cannot produce enough energy under anaerobic conditions to sustain cardiac func-
tion and viability, and therefore, irreversible cardiomyocyte injury develops after 40–60 min of sus-
tained severe ischemia (Jennings et al. 1990 ) . The predominant mechanism of cardiomyocyte death in
the infarcted heart is coagulation necrosis, although apoptosis is also likely to contribute to cardio-
myocyte loss. Dying cells activate innate immune mechanisms, initiating an in ammatory reaction
that has a dual effect, primarily repairing the trauma but also producing adverse ventricular remodel-
ing, with detrimental consequences if activation is enhanced or uncontrolled. Innate immunity is a
primordial system that was evolved in the lower phyla as a host reaction system primarily for repair-
ing cutaneous trauma. The critical goals of this reaction system were to prevent an infection; dispose
of apoptotic, necrotic cells and damaged tissue; and, in these and other ways, facilitate wound repair.
The size of the eventual scar was not the key issue. On the other hand, scar tissue is largely acellular
and lacks the normal biochemical properties of the host cells, and in cardiac tissue, this situation leads
to electrical uncoupling, mechanical dysfunction, and loss of structural integrity. A compensatory
hypertrophy of the noninfarcted area, accompanied by chamber dilation and upregulation of fetal gene
expression, also develops at a later point, and ultimately, these changes lead to the development of
heart failure (Jennings et al. 1995 ) . We have put forward the hypothesis that temporally regulated
activation and suppression of innate immunity may be critical for minimizing cardiac tissue injury and
achieving effective cardiac repair and regeneration (Jiang and Liao 2010 ) .
14.1.2 Complement System Activation in Cardiac Ischemia/Reperfusion
The complement system is an important arm of innate immunity whose essential role in ischemia/
reperfusion damage was rst described by Hill and Ward in the early 1970s (Hill and Ward 1971 ) .
After ischemia and reperfusion, myocardial levels of C3 and C9 have been shown to exceed the levels
of production of these complement components in the liver in a rabbit model (Yasojima et al. 1998 ) .
Extensive experimental data from a variety of animal models have indicated that inhibiting comple-
ment activation offers protection to cardiac tissue after ischemia/reperfusion [for a review, see
Diepenhorst et al. ( 2009 ) ]. In patients with acute MI, however, the results of clinical studies using
complement inhibitors (four studies of more than 10,500 patients using pexelizumab, an anti-C5 anti-
body, and studies using C1 esterase inhibitor or soluble complement receptor 1) have largely been
disappointing [for a review, see Oksjoki et al. ( 2007 ) and Diepenhorst et al. ( 2009 ) ]. Over the past 30
years, the use of speci c strategies that modulate the in ammatory response have led to dramatic
reductions in infarct size and attenuation of adverse remodeling in numerous experimental animal
models [reviewed in Frangogiannis ( 2008 ) ]. Nevertheless, attempts to mitigate the in ammatory reac-
tion in human patients after MI have yielded disappointing results. At present, no speci c immuno-
modulatory approach is used in patients after MI. Established therapeutic strategies such as b -adrenergic
blockade, ACE inhibition, and anticoagulation treatment may exert their bene cial effects in part by
interfering with the in ammatory cascade (Kilgore et al. 1994 ; Chen et al. 1998 ) .
Why have anti-in ammatory strategies that yield very promising results in animal models of car-
diac tissue injury been unsuccessful in human patients? Although fundamental differences between
animal models and their respective human diseases may provide an explanation in some cases, what
is becoming evident from the studies conducted thus far is that a deeper understanding of the biology
of this complex network of molecular and cellular interactions is necessary before any speci c inter-
ventions can be pursued in therapeutic trials. In addition, in the clinical reality, variables such as age;
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20914 Complement System Activation in Cardiac and Skeletal Muscle Pathology
the presence of comorbid conditions such as diabetes, obesity, and hyperlipidemia; the timing of
reperfusion; and genetic variations between individuals greatly complicate the prediction of the
potential effects of a therapeutic intervention. Finally, in the various animal models of MI, the out-
come of the cardiac injury may also differ according to various conditions, such as whether the MI is
small or large, whether a small or large amount of myocardial regeneration occurs, and whether the
ischemia is permanent or transient. If the duration of the ischemia is limited (e.g., 30 min, as in many
of the MI/reperfusion animal models used to assess the effects of complement system inhibition), the
main determinants in the outcome of the cardiac injury are the extent of the initial injury of the vessel
endothelium after the reperfusion and the degree of activation of the complement system. Complement
activation stimulates neutrophils by inducing chemotactic migration, aggregation (Crawford et al.
1988 ) , and the release of cytotoxic products such as proteases, elastases, and reactive oxygen species
that can compromise tissue integrity (Engler 1987 ; Hori and Nishida 2009 ) . Neutrophils activated by
C5a or by iC3b deposited within vessel walls (Hirahashi et al. 2006 ) can also contribute to microvas-
cular obstruction by promoting brin deposition and thrombosis; thus, the complement system can
contribute to a vicious cycle of vasoconstriction, microvascular hypoperfusion, and cell death (Saraste
et al. 1997 ; Rezkalla and Kloner 2002 ) that further compromises cardiac tissue integrity. Thus, in the
case of ischemia of limited duration with subsequent reperfusion, there is wide evidence that oxygen-
derived free radicals, neutrophils, and activated complement components are crucial players in the
destruction of cardiac tissue (Kilgore et al. 1994 ; Diepenhorst et al. 2009 ) . If it were possible to
restore the oxygen supply to cardiomyocytes without inciting the harmful acute in ammatory reac-
tion, then the restoration of cardiac function could be maximized.
In longer-lasting ischemia, when irreversible cardiomyocyte injury develops as a direct result of
oxygen deprivation, the extent of cardiomyocyte death determines the outcome of the cardiac dys-
function. Damaged and destroyed cardiomyocytes in the ischemic area can be roughly divided into
two categories: (a) cardiomyocytes under severe and extended stress whose death is inevitable and
(b) those that, while under stress, will eventually manage to survive through the upregulation of cyto-
protective mechanisms. The intrinsic cytoprotective mechanisms of cardiomyocytes can involve the
upregulation of small heat shock proteins, antioxidant enzymes, antiapoptotic molecules, and/or a
fetal gene expression program. As previously mentioned, dying cardiomyocytes can trigger an
in ammatory reaction, activating reparative pathways that ultimately result in the formation of a scar
(Frangogiannis 2008 ) . Enhanced or uncontrolled activation can lead to adverse ventricular remodel-
ing, with detrimental consequences. Timely resolution of the in ammatory in ltrate and spatial con-
tainment of the in ammatory and reparative response to the infarcted area are essential for optimal
infarct healing.
What is the role of complement activation in this milieu of interactions among secreted factors,
resident cells, and cells that have in ltrated into the site of injury? Although the complement system
often contributes to the tissue damage described in the previous paragraph, it is not easy to separate
this process from the bene cial effects of complement with regard to waste disposal, recruitment of
stem cells (Ratajczak et al. 2006 ) , regeneration (Markiewski et al. 2009 ) , and angiogenesis (Nozaki
et al. 2006 ) , as has been recognized in other tissue injury models. The ideal approach is likely to be
modulating rather than abruptly blunting complement activation.
14.1.3 Complement Activation as a “Friend” in Cardiac Tissue Injury
Complement activation leads to the deposition of iC3b at the site of necrotic cardiomyocytes; the iC3b
then binds to complement receptors CR3 and CR4 and mediates phagocytosis, thereby participating
in the removal of cell debris and containment of the in ammatory reaction. CR3/4-mediated phago-
cytosis, per se, does not elicit proin ammatory signals in phagocytes nor does it provoke a respiratory
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210 M. Syriga and M. Mavroidis
burst. Considering that CR3 has primarily evolved to support the physiologic functions necessary to
balance tissue homeostasis, i.e., through clearance of apoptotic cells, the lack of in ammation is not
surprising. In order to promote leukocyte adhesion and transmigration through the endothelium as
well as in ammatory responses, C3 needs to be preactivated to a high-af nity state in order for liga-
tion to result in an in ammatory response. Signaling pathways that induce this high-af nity status
include those downstream of activating immunoglobulin G receptors (Fc g Rs) and the chemoattractant
G-protein-coupled receptors (GPCR).
Secreted in ammatory mediators play an important role in mobilizing progenitor cells and may
regulate their homing to the infarcted myocardium (Vandervelde et al. 2005 ) . Most of the positive
effects reported thus far for cell-based therapy have been attributed to paracrine effects rather than to
the direct differentiation of engrafted cells to cardiomyocytes (Choi et al. 2011 ) . Thus, although it has
not yet been clearly established for cardiac tissue injury, C3 cleavage fragments may exert a bene cial
effect by participating in the chemoattraction and tethering of circulating CXCR4+ stem cells to
injured cardiac tissue, as has been reported in other tissue injury models (Ratajczak et al. 2006 ) . All
of the players (activated complement components, CRXR4-positive cells, SDF1) in this interaction
are present in the injured myocardium ((Askari et al. 2003 ; Abbott et al. 2004 ) ; see also Fig. 14.1 ).
The prorepair role of complement is apparent in liver regeneration, in which anaphylatoxin-induced
IL-6 and TNF prosurvival signaling promote hepatocyte growth and proliferation, which are dramati-
cally impaired in C3–/–, C3ar–/–, and C5ar–/– mice (Markiewski et al. 2009 ) . The role of TNF- a in
myocardial ischemia/reperfusion injury is ambivalent. Excessive TNF- a expression and subsequent
stimulation of cardiomyocyte TNF receptor type 1 induce contractile dysfunction, hypertrophy, brosis,
and cell death, whereas lower TNF- a concentrations and subsequent stimulation of cardiomyocyte
TNF receptor type 2 are protective (Schulz and Heusch 2009 ) . TNF- a can increase protective signals
such as the wingless-type integration site family member 1 (WNT1) in cardiomyocytes and cardiac
Fig. 14.1 Activated complement components and CRXR4-positive cells are present in the myocardium of the desmin-null
cardiac tissue injury/heart failure model. ( a ) CRXR4-positive cells are present in areas of cardiac tissue injury and
neovascularization in the desmin-null myocardium. ( b ) Extended iC3b deposition ( red ) is observed in the desmin null
cardiac tissue, in areas of cardiomyocyte degeneration. Actinin ( green ) is a cardiomyocyte marker (bar = 25 m m).
Extended cardiomyocyte injury and acute in ammatory reaction are observed in the desmin-null mice (Mavroidis and
Capetanaki
2002 ) , together with a 7.5-fold upregulation of CR3 RNA levels (Psarras et al. 2011 )
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21114 Complement System Activation in Cardiac and Skeletal Muscle Pathology
broblasts (Venkatachalam et al. 2009 ) . It has been shown in in vitro experiments that complement
activation can induce TNF- a synthesis in cardiac myocytes (Zwaka et al. 2002 ) , so a reasonable
hypothesis could be advanced that, depending on the timing, the concentration, and the cardiac injury
model chosen, induction of TNF- a by C3 activation could be protective. In liver injury, it has been
shown that there is a threshold of complement activation for optimal liver regeneration (He et al. 2009 ) ,
and this threshold is associated with increased early hepatic production of IL-6 and TNF- a and dimin-
ished systemic levels of the in ammatory cytokines by 48 hours after partial hepatectomy.
On the other hand, in TNF- a -overexpressing mice, analysis of gene expression pro ling during the
transition to heart failure (Tang et al. 2004 ) has demonstrated the development of autoimmune myo-
carditis, with C3 and MHC class II antigen expression being strongly upregulated (see Fig. 14.2 ).
Given that complement activation is critical for the induction of experimental autoimmune myocardi-
tis (Kaya et al. 2001 ; Eriksson et al. 2003 ) , depletion of C3 may be essential to preventing TNF- a -
induced heart failure.
14.1.4 A Potential Role for the Connection of Complement to Fatty Acid
and Glucose Metabolism in the Adaptation of the Heart to Stress
A common feature of a variety of cardiac pathophysiological conditions, including hypoxia, ischemia,
and hypertrophy, is a return to a pattern of fetal metabolism. This adaptation is associated with a
whole program of cell survival under stress, and there is evidence that, at the same level of stress,
survival is greater in the fetal-adapted heart than in the adult heart (Rajabi et al. 2007 ) . A hallmark of
fetal metabolism is the predominance of carbohydrates as substrates for energy provision. Under nor-
mal conditions, the preferred substrate for the heart is fatty acids (80–100%). During ischemia/
reperfusion, the diminished O
2
supply for respiration and oxidative phosphorylation leads to a rapid
decline in ATP levels and stimulates anaerobic ATP generation, with an increase in glycolysis and
Fig. 14.2 ( a ) Activated C3 ( red ) is seen in aggregated form and perinuclearly in the cardiomyocytes of cardiac-speci c
tumor necrosis factor (TNF- a )-overexpressing transgenic mice. In TNF- a -overexpressing mice there is a mislocaliza-
tion of the intercalated disc components to the lateral sarcolemma of the cardiomyocytes, as shown by b -catenin ( green )
staining. ( b ) A cardiac tissue section of wild-type mice is negative for activated C3 staining, and b -catenin is mainly
localized to the bipolar ends of the rod-shaped cardiomyocytes. Frozen cardiac tissue sections; blue denotes nuclear
staining with DAPI, and the bar = 25 m m. TNF- a -overexpressing mice are from Li et al. (
2000 )
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212 M. Syriga and M. Mavroidis
lactate production (Oram et al. 1973 ; Neely and Morgan 1974 ) . Although the fetal-adapted heart can
survive better than the adult under hypoxic stress, this adaptation cannot meet the high cardiac pump-
ing workload (and, consequently, high energy demand) of an adult organism. At a certain point, the
fetal gene program is no longer suf cient to support cardiac structure and function, and the heart suc-
cumbs to maladaptation, cell death, and ultimately organ failure (Rajabi et al. 2007 ) .
An emerging role of complement system in the regulation of lipid metabolism has become appar-
ent in the last decade. Acylation-stimulating protein (ASP), which has been identi ed as C3adesArg
(produced after cleavage of the COOH-terminal arginine of C3a by carboxypeptidase B), has lipo-
genic activity. In adipocytes, ASP interacts with its cell-surface receptor C5L2, resulting in increased
nonesteri ed fatty acid (NEFA) uptake and triglyceride synthesis (MacLaren et al. 2008 ) . Conversely,
in muscle, ASP has the opposite effect and increases lipolysis, indicating a reduction in NEFA trap-
ping within muscle (Faraj and Cian one 2004 ) . ASP also increases glucose uptake in a number of cell
models, including human adipocytes and L6 myotubes, via the translocation to the cell surface of the
glucose transporters GLUT4 and GLUT3 (in myotubes), independently of, but additively with, insu-
lin. Thus, the generation of C3adesArg at the site of cardiac tissue injury through complement activa-
tion could offer a potential means of improving the energy utilization of glucose and lipids when
under hypoxic stress.
14.2 The Role of Complement in Skeletal Muscle
14.2.1 Skeletal Muscle Regeneration
Skeletal muscle is the most abundant tissue in the human body, contributing to motion, protein and
carbohydrate storage, and heat production. Given its super cial body distribution, it is constantly
exposed to mechanical insults caused by muscle contraction and to external physical trauma. As a
consequence of the signi cance of its role and its continuous exposure to mechanical stress, skeletal
muscle has developed the ability to regenerate and restore its original tissue architecture and func-
tional integrity after damage. Acute muscle injury and genetic de ciencies both lead to compromised
muscle function, along with rupture, myo brillar necrosis, and in ammation. The degenerative phase
is followed by tissue recovery and repair, which are characterized by the removal of necrotized tissue
and regeneration of the damaged myo bers (Tidball and Villalta 2010 ) . This phase depends on a
population of progenitor myogenic cells, the satellite cells, which are located in the basal lamina of
the muscle bers. A complex repertoire of cytokines, growth factors, and extracellular molecules
(ECM) triggers the activation, proliferation, and differentiation of satellite cells that result in the sub-
sequent fusion and formation of new, intact myo bers (Corti et al. 2001 ) . A key role in this process is
performed by the tightly regulated interplay between the injured skeletal muscle and the immune
system (Tidball 2005 ) .
Immediately after damage, neutrophils invade the muscle and promote the progress of the inflam-
matory reaction in the damaged area. The subsequent accumulation of macrophages continues the
proin ammatory phase of muscle injury (Nguyen and Tidball 2003 ) . The M1 phenotype dominates the
initial macrophage population, with the release of proin ammatory cytokines such as TNF- a and IL-6
(Collins and Grounds 2001 ) . TNF- a and the rest of the Th1 cytokines produced in this phase contribute
to muscle damage, but at the same time they also promote regeneration and repair by triggering the
activation, proliferation, and early differentiation stages of satellite muscle cells (Collins and Grounds
2001 ; Villalta et al. 2009 ) . The M1 phenotype is replaced by in ltrating M2 macrophages, which attenu-
ate the previous proin ammatory reaction and release anti-in ammatory cytokines such as IL-10, IL-4,
and IL-13, thus promoting tissue growth and regeneration (Tidball and Wehling-Henricks 2007 ) .
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14.2.2 Complement Activation in Skeletal Muscle
Initial studies have revealed the biosynthesis of complement by skeletal muscle cells in vitro. The
alternative pathway proteins C3, factor B, and factor H as well as the classical pathway components
C1, C2, and C4 have been shown to be produced by human myoblast cell lines (Legoedec et al. 1995,
1997 ) . Despite the activation of autologous complement, muscle bers are protected from destruction
and cell lysis by their abundant expression of the regulatory molecules MCP, CD59, and C4BP
(Gasque et al. 1996 ) . Complement synthesis by muscle cells is upregulated by in ammatory cytok-
ines, including INF- g and IL-1 (Legoedec et al. 1995 ) , suggesting a potent role for the local produc-
tion and activation of complement under in ammatory pathophysiological conditions in skeletal
muscle. A series of studies have pinpointed the involvement of the complement cascade in the com-
plete reconstruction and regeneration of body parts such as limbs and eye tissue in several species of
urodele amphibians (Kimura et al. 2003 ) . The central proteins C3 and C5 of the complement cascade
are speci cally expressed at both the transcriptional and translational (protein) levels in the areas
responsible for the repopulation and reformation of the old, amputated (or injured) structures (i.e., the
urodele blastema) into the appropriate differentiated, newly reconstructed body parts.
An important question is how complement proteins are related to the molecules responsible for the
establishment of cellular adhesion and communication. C3 interacts with molecules of the extracel-
lular matrix, such as laminin and bronectin (Leivo and Engvall 1986 ) , and with receptors of the
integrin family (Law et al. 1987 ) ; factor B and C2 share homologous domains with cartilage matrix
protein, von Willebrand factor, and the collagen-binding domain of alkaline phosphatase. Given the
upregulation of these molecules during the synthesis of new cell membranes in the regeneration phase
(Tsonis et al. 1996 ) , these interactions strongly suggest that complement participates in the progress
of remodeling and the differentiation of the regenerating parts. Recent reports have highlighted the
important role of complement in removing cellular debris during successful skeletal muscle remodel-
ing after chemical or ischemic injury in stem cell antigen 1 (Sca1)-de cient mice (Long et al. 2011 ) .
Recruitment of IgM and complement C3 has been shown to be essential for tissue regeneration since
the inability of damaged cells to be cleared from C3-de cient mice leads to a profound brotic condi-
tion (Mevorach et al. 1998 ; Markiewski et al. 2004 ) .
14.2.3 Complement-Mediated In fl ammatory Muscle Diseases
In ammatory myopathies are a heterogeneous class of autoimmune muscle disorders that are charac-
terized by the production of speci c autoantibodies and immune attack against muscle antigens, with
subsequent tissue malfunction. As an important link between innate and adaptive immunity, comple-
ment plays a key role in the pathogenesis of serious autoimmune diseases such as myasthenia gravis
(Vincent and Drachman 2002 ) and the in ammatory myopathies dermatomyositis and juvenile der-
matomyositis (Dalakas 2010b ) . In addition, complement contributes to the pathogenesis of a class of
in ammation-associated muscle disorders, the dysferlinopathies (Liu et al. 1998 ) , in which a defec-
tive repair of the plasma membrane produces constant leakage of cytoplasmic contents to the extracel-
lular environment.
The immune system recruits complement in the course of many other pathologic muscle condi-
tions, thus preserving the in ammatory “waves” in the local area of the affected skeletal muscle and
leading to either progressive or acute tissue destruction, as in the case of muscle ischemia/reperfusion
injury.
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14.2.3.1 Complement in Myasthenia Gravis
Myasthenia gravis (from Greek and Latin, literally meaning “grave muscle weakness”) is a chronic
autoimmune disease that is characterized by the production of autoantibodies that target the acetyl-
choline receptor (AchR) at the neuromuscular junction (NMJ) (Vincent and Drachman 2002 ) . Studies
conducted over the past few decades have shown that complement is a signi cant participant in the
destructive pathway that leads to neuromuscular disruption (Lennon et al. 1978 ; Tuzun et al. 2003 ) .
Initial evidence has revealed a reduction in circulating serum levels of several complement proteins in
myasthenia gravis patients, in parallel with the deposition and colocalization of IgG, C3, and C9 on
the NMJ membrane (Sahashi et al. 1980 ) . In agreement with the clinical evidence, in mice with exper-
imental autoimmune myasthenia gravis (EAMG), IgG and complement proteins are detected at the
degenerating NMJs (Tuzun et al. 2004 ) . Studies including the manipulation of complement activation
in EAMG animals, either through a genetic de ciency or pharmacologic blockade, have con rmed a
role for complement in the disease mechanism. C3 and C4 knockout mice immunized with the AchR
show an improved phenotype when compared to wild-type controls (Tuzun et al. 2007 ; Zhou et al.
2007 ; Soltys et al. 2009 ) . However, examination of the disease progress in C5aR knockout mice has
revealed that the incidence in EAMG mice is similar to that of wild-type mice, suggesting that the
anaphylatoxin C5a is not involved in the pathogenesis of myasthenia gravis; instead, C5 plays a role
in pathogenesis in the form of C5b, which contributes to the subsequent formation of the membrane
attack complex (MAC) (Chamberlain-Banoub et al. 2006 ; Qi et al. 2008 ) . Novel therapeutic reagents
speci cally targeting the activation of the classical pathway and preventing MAC formation at the
NMJs, such as monoclonal antibodies or even complement inhibitors conjugated to anti-AchR anti-
body fragments (Spitzer et al. 2004 ) , could potentially eliminate myasthenia gravis symptoms by
preventing complement activation speci cally at the site of pathology rather than systemically.
14.2.3.2 Complement in Dermatomyositis (DM)
In ammatory myopathies comprise a heterogeneous group of degenerative muscle diseases with the
common characteristics of mild to severe muscle weakness and the presence of in ammatory in ltrates
in muscle biopsies. This group consists of four clinically distinct conditions: dermatomyositis, poly-
myositis, necrotizing autoimmune myositis, and sporadic inclusion body myositis (Dalakas 2010b ) .
Although their general phenotypic characteristics are similar, the pathophysiological mechanisms of
these diseases differ and involve the recruitment of different branches of the immune system. Complement
plays a signi cant role in the pathogenesis of dermatomyositis, and therefore complement activity is
considered a useful diagnostic measure and a strong candidate as a future therapeutic target.
Dermatomyositis occurs in both adults and children (juvenile dermatomyositis) and predominantly
affects the skin and muscles. It is characterized by a humorally mediated autoimmune attack against
the vascular endothelium of the capillaries. Deposited autoantibodies activate the complement cas-
cade, with the subsequent formation of the MAC complex (Kissel et al. 1986 ; Emslie-Smith and
Engel 1990 ) . Immune attack and complement activation lead to extended capillary damage and necro-
sis, resulting in an insuf cient blood supply to the muscle. The detection of MAC offers high sensitiv-
ity and speci city for discriminating dermatomyositis from other in ammatory myopathies at both
the early and advanced stages (Jain et al. 2011 ) . The future unraveling of complement-mediated
destruction and the related cytokine milieu can potentially give us the ability to design targeted thera-
peutic strategies. Monoclonal antibodies against immune components, including complement (Basta
and Dalakas 1994 ) , cytokines, T- and B-cell activating factors, and adhesion molecules, are promising
candidates and have proven effective in some cases of dermatomyositis (Dalakas 2010a ) .
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14.2.3.3 Complement in Dysferlin Myopathy
Dysferlinopathies comprise a large group of autosomal recessive myopathies caused by the complete
or partial absence of dysferlin. Dysferlin is a 230-kDa protein that acts as a key player in the mem-
brane repair system (Cenacchi et al. 2005 ) . Membrane tears cause an increase in Ca
2+
in fl ux, which
triggers the accumulation of dysferlin-containing vesicles and fusion to the plasma membrane at the
site of the membrane disruption (Lennon et al. 2003 ) . Dysferlin mutations give rise to three types of
autosomal recessive muscle wasting diseases: limb-girdle muscular dystrophy type 2B, Miyoshi myo-
pathy, and a distal anterior compartment myopathy (Liu et al. 1998 ) .
The immune system plays a complex role in the pathogenesis of dysferlinopathy. Muscle biopsies
of dysferlin-de cient patients exhibit a prominent in ammatory in ltration (Gallardo et al. 2001 ) ,
while the lack of dysferlin in the SJL/J mouse strain results in an altered behavior of immune cells,
with an increase in phagocytic activity and upregulation of the in ammasome molecular pathway
(Rawat et al. 2010 ) . Complement activation with subsequent MAC deposition on the surface of non-
necrotic muscle bers has been detected on both human and murine dysferlin-de cient skeletal mus-
cles, suggesting a potential role in the pathology (Spuler and Engel 1998 ) . Microarray data have
indicated the upregulation of complement proteins and downregulation of complement regulators
(Suzuki et al. 2005 ) . In accordance with these microarray results, decay-accelerating factor (DAF) has
been found to be eliminated at both the RNA and protein levels in skeletal muscle from dysferlin-
de cient mice and patients diagnosed with dysferlinopathy (Wenzel et al. 2005 ) , and this loss of DAF
has been shown to result in an increased susceptibility to complement attack. Further investigation has
revealed that disruption of the C3 gene in dysferlin-de cient skeletal muscle attenuates muscle pathol-
ogy by reducing in ammatory in ltration, central nucleation, brosis, and fat replacement (Han et al.
2010 ) . Interestingly, ablation of the terminal complement component C5 has only a minimal effect on
muscle pathology in this mouse model, suggesting that it is not the terminal activation of the comple-
ment system but the activation of C3 that accelerates muscle injury.
In summary, dysferlinopathy causes a strong activation of the complement cascade with a simulta-
neous attenuation of regulatory control, making the muscle bers more susceptible to complement
attack. The additional involvement of dysferlin in cytokine release is another aspect of the compro-
mised muscle remodeling and healing. Based on these ndings, an inhibition of the complement
cascade at the C3 level could offer potential therapeutic options for this currently untreatable muscle
wasting disorder.
Acknowledgments We thank Dr. Yassemi Capetanaki for the continuous support, Dr. Douglas L. Mann
for providing the TNF- a overexpressing mice, and Dr. Deborah McClellan for the editorial assistance.
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