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Anti-Inflammatory Effect of Exercise Mediated by Toll-Like Receptor
Regulation in Innate Immune Cells –A Review
Nicolas Collao
a
, Isabel Rada
a
, Marc Francaux
b
, Louise Deldicque
b
, and Hermann Zbinden-Foncea
a,c
a
Exercise Science Laboratory, School of Kinesiology, Faculty of Medicine, Universidad Finis Terrae, Santiago, Chile;
b
Institute of
Neuroscience, UCLouvain, Louvain-la-Neuve, Belgium;
c
Centro de Salud Deportivo, Cl
ınica Santa Mar
ıa, Santiago, Chile
ABSTRACT
Over the last three decades, the combination of a sedentary lifestyle and excessive food
intake has led to a significant increase in the prevalence of obesity. The latter favors a
chronic low-grade inflammatory state and an over-activation of the innate immune system,
which contribute to insulin resistance and type 2 diabetes. Physical exercise is a powerful
preventive tool and treatment for several diseases as it induces metabolic and immune
effects that provide health benefits. Exercise is known to reduce inflammation; however, the
underlying mechanisms responsible are not fully elucidated. One proposed mechanism is a
reduced expression and/or activation of pro-inflammatory toll-like receptors (TLRs) on innate
immune cells after exercise, which could contribute to the protective effect of exercise
against insulin resistance and the prevention of the development of metabolic diseases. The
aim of the present study is therefore to review the current evidence about the anti-inflam-
matory effects of exercise and toll-like receptors regulation on immune cells in humans.
KEY POINTS
1. Obesity leads to a low-grade chronic inflammatory state and an over-activation of the
innate immune system that is directly involved in the develop metabolic syndrome.
2. The anti-inflammatory effect of exercise has been previously suggested through the
reduction of the expression and/or activation of pro-inflammatory toll-like receptors
(TLRs) in innate immune cells, which represent one of the main inflammatory responses
triggered by obesity
3. The underlying mechanisms in which toll-like receptors expression modulate the reduc-
tion of chronic inflammation are not fully elucidated.
Abbreviations: TLRs: Toll-like receptors; MetS: Metabolic syndrome; T2D: Type 2 diabetes;
ROS: Reactive oxidative species; RNS: Reactive nitrogen species; PBMCs: peripheral blood
mononuclear cells; IjB-a: Nuclear factor of kappa light polypeptide gene enhancer in B-cells
inhibitor alpha; IL: Interleukin; TNF-a: Tumor necrosis factor alpha; IFN-c: Interferon gamma;
PAMPs: Pathogen-associated molecular patterns; PRRs: Pattern recognition receptors; DAMP:
Damage-associated molecular pattern; LPS: Lipopolysaccharides; HSPs: Heat shock proteins;
lcSFAs: Long-chain saturated fatty acids; MyD88: Myeloid differentiation factor 88; NF-jB:
Nuclear transcription factor kappa B; MCP-1: Monocyte chemotactic protein-1; BMI: Body
mass index; MAPKs: Mitogen-activated protein kinases; ERK: protein kinase 1/2; JNK: c-Jun
amino-terminal kinase; IRS-1: Insulin receptor substrate (IRS)-1; miRNAs: MicroRNAs.
ARTICLE HISTORY
Received 5 April 2019
Accepted 8 October 2019
KEYWORDS
Physical activity;
inflammation; obesity;
cytokines; metabolic
syndrome; diabetes
Background
Over the last three decades, changes in human lifestyle
such as diet, physical activity and pollutant exposure
have led to worldwide epidemic levels of overweight
and obesity [1]. Obesity is an important global problem,
since it is one of the major public health concerns due
to its elevated prevalence that reaches pandemic
characteristics. In 2013, it was estimated that 2.1 billion
adults worldwide were overweight (BMI 25 kg/m
2
), of
which 600 million were obese (BMI 30 kg/m
2
).
Therefore, we may now state that the prevalence of
overweight and obesity has reached epidemic propor-
tions [2]. This condition is a major risk factor for the
development of the metabolic syndrome (MetS) and
CONTACT Hermann Zbinden-Foncea hzbinden@uft.cl Exercise Science Laboratory, School of Kinesiology, Faculty of Medicine, Universidad Finis
Terrae, 1509 Pedro de Valdivia Av. Providencia, Santiago, Chile; Centro de Salud Deportivo, Cl
ınica Santa Mar
ıa, Santiago, Chile.
Color versions of one or more of the figures in the article can be found online at www.tandfonline.com/iiri.
ß2019 Taylor & Francis Group, LLC
INTERNATIONAL REVIEWS OF IMMUNOLOGY
https://doi.org/10.1080/08830185.2019.1682569
other associated health complications including insulin
resistance, type 2 diabetes (T2D), hyperglycemia, hyper-
tension, and dyslipidemia, which are themselves risk
factors for cardiovascular disease [3].
Exercise improves metabolic health, being a front-
line therapy for the prevention and treatment of MetS
[4] amongst others due to its anti-inflammatory
effects [5–7]. Some of the anti-inflammatory effects of
regular exercise are likely attributed to a reduction in
the size of adipose tissue [8]. However, growing evi-
dence suggest that acute exercise could directly impact
immune cells by regulating systemic inflammatory
mediators independently of weight or fat loss [6].
Individuals with overweight and obesity are an excel-
lent target group for lifestyle interventions as insulin
resistance is reversible at this stage [9].
The innate immune cellular responses are initiated
by the recognition of diverse molecular components
from pathogens. These pathogen-associated molecular
patterns (PAMPs) are recognized by pattern recognition
receptors (PRRs) located either at the cell surface or at
intracellular compartment surfaces and expressed on
various types of leukocytes. Toll-like receptors (TLRs)
are part of the pattern recognition receptors family and
have been implicated in the pathogenesis of obesity and
T2D [10,11]. Their roles will be presented hereafter.
Therefore, in the present study, we will review the
anti-inflammatory effects of exercise on innate immune
cells since a link between TLRs down-regulation, reduc-
tion in the production of pro-inflammatory cytokines
and insulin sensitivity improvement has been suggested
after exercise.
Obesity-induced inflammation
The first insight on the inflammatory origin of obesity
and diabetes came from studies in the early 1990s.
Hotamisligil et al. (1993) reported that insulin resistance
was related to the expression of the pro-inflammatory
cytokine TNF-ain adipose tissue of obese mice [12]. A
few years later, the same group confirmed those results
in obese individuals [13]. The key evidence linking
immunity to metabolism came from an obese mouse
model lacking TNF-afunction. These mice displayed
improved insulin sensitivity and glucose homeostasis,
indicating that the inflammatory response had a critical
role in the regulation of insulin action in obesity [14].
Activation of pro-inflammatory pathways as well as
inflammatory cytokine production have been found in
adipocytes and diverse types of immune cells, such as
macrophages, highlighting the capacity for adipose tis-
suetocontributetoinflammatoryprocesses[15]. White
adiposetissuemacrophagesinmicearebonemarrow
derived, suggesting that macrophages result from circu-
lating monocytes that infiltrate white adipose tissue,
rather than from differentiation from pre-adipocytes
[16]. Of note, approximately 60% of TNF-aexpression
is derived from the macrophages within adipose tissue
[16]. Therefore, the study of monocytes might be a
good approach for the estimation of macrophage activ-
ity and subsequent modulation of metabolism in adi-
pose tissue. Diet-induced obesity seems to promote the
polarization of macrophages in adipose tissue to a pro-
inflammatory (M1-polarized) phenotype that contrib-
utes to insulin resistance [17,18].
In addition to adipose tissue, the first evidence that
peripheral blood mononuclear cells (PBMCs) dis-
played a pro-inflammatory state in obese individuals
was reported more than a decade ago [19]. A
decreased IjB-aexpression and a higher NF-jB
DNA-binding was found in PBMCs of obese com-
pared to lean individuals [19]. Similarly, PBMCs from
obese individuals secreted more TNF-a, IFN-cand IL-
2 and less IL-10 compared to healthy controls [20].
As it plays a critical role in both the innate
immune function and inflammatory response to obes-
ity [11], it has been suggested that one of the specific
roles of the TLR family could be to mediate obesity-
induced inflammatory response.
Toll-like receptors
Emerging evidence suggest that TLRs may be involved
in the link between a sedentary lifestyle, inflammation
and disease [6]. TLRs are evolutionary preserved pat-
tern-recognition receptors that are primarily expressed
on immune cells [21]. Up to now, eleven TLR members
have been discovered in human and thirteen in mice
[22]. TLRs initiate an immune response after recogni-
tion of diverse exogenous signals, amongst which
PAMPs, such as endotoxin like lipopolysaccharides
(LPS), and endogenous ligands like damage-associated
molecular pattern molecules (DAMP) involved in sterile
inflammation induced by tissue damage and cellular
stress, such as heat shock proteins (HSPs) [23]. TLR
activation triggers a pro-inflammatory response, which
results in cytokines production [24].
The two main TLRs involved in chronic low-grade
inflammation associated with altered metabolism are
TLR2 and TLR4 [25], which can be activated by ele-
vated free fatty acid (FFA) levels to induce pro-
inflammatory cytokines expression in adipocytes, and
liver [26]. However, the activation of TLRs by lipids
has recently been challenged [27]. Long-chain
2 N. COLLAO ET AL.
saturated fatty acids (lcSFAs) were found not to be
TLR4 agonists in monocytes/macrophages. Instead,
Lancaster et al. suggested that TLR4 indirectly regulates
lcSFA-induced inflammation by altering macrophage
lipid metabolism [27].
The recognition of ligands by TLRs activates mye-
loid differentiation factor 88 (MyD88)-dependent or
-independent pathways causing subsequent inflamma-
tory responses (Figure 1). The nuclear transcription
factor kappa B (NF-jB) is a key transcriptional factor
for the expression of inflammatory cytokines, such as
IL-6, TNF-a, monocyte chemotactic protein-1 (MCP-
1) or IL-1 [23], amongst which IL-6 and TNF-aare
more specifically involved in obesity-induced insulin
resistance [28]. Compared to lean controls, the mRNA
levels of IL-6 and TNF-ain PBMCs of obese subjects
were higher and correlated with TLR2 and 4 expres-
sion [29]. Additionally, a positive correlation was
found between the expression of TLR2, TLR4, MyD88
and body mass index (BMI).
Once activated, TLRs trigger a variety of signal trans-
duction pathways, leading to the production of inflam-
matory cytokines and initiation of downstream
inflammatory cascades [30]. In macrophages, evolution-
arily conserved signal transduction pathways have been
shown to mediate inflammatory processes including
those activated by the mitogen-activated protein kinases
(MAPKs) [31,32]. The MAPK family is composed of
three major members: the extracellular signal-regulated
protein kinase 1/2 (ERK), the p38 MAPK (p38), and the
c-Jun amino-terminal kinase (JNK), which differentially
regulate numerous cellular functions, including inflam-
mation [33]. The activation of the MAPKs results in
downstream substrates phosphorylation and activation
of several transcription factors, amongst which NF-jB
has been widely studied. Increased NF-jB activity will
Figure 1. TLR activation in immune cells and downstream signaling.
Obesity state promotes the activation of a pro-inflammatory pathway triggered by TLR2/4, leading to the production of cytokines
as TNF-aand IL-6 which impairs the insulin downstream signaling inducing insulin resistance in skeletal muscle, adipose tissue
and liver. LPS: Lipopolysaccharides; TLR2/4: Toll-like receptors 2/4; TIRAP: TIR Domain-Containing Adaptor Protein; MYD88 myeloid dif-
ferentiation factor 88; IRAK: Interleukin-1 receptor-associated kinase 1; TRAF6: TNF Receptor-Associated Factor 6; TAK1: Transforming
growth factor-b-Activated Kinase 1; IKK: IjB Kinase; IkBa: alpha Nuclear factor of kappa light polypeptide gene enhancer in B-cells
inhibitor alpha; NFkB: Nuclear transcription factor kappa B; ERK1/2: protein kinase 1/2; JNK c-Jun amino-terminal kinase; MAPKs:
Mitogen-activated protein kinases; P38: p38 mitogen-activated protein kinases; AP1: activator protein 1; P65: nuclear factor NF-kappa-B
p65 subunit; P50: nuclear factor NF-kappa-B p50 subunit; TNF alpha: Tumor necrosis factor alpha; IL6: Interleukin–6; IRS- 1/2: Insulin
receptor substrate 1/2; PI3K:phosphatidylinositol 3-kinase; AKT: Protein kinase B (PKB); GLUT2/4: glucose transporter type 2/4; IR: insulin
resistance; TG: Triglycerides; KCs: Kupffer cells
INTERNATIONAL REVIEWS OF IMMUNOLOGY 3
activate the transcription of pro-inflammatory genes,
such as IL-6 and TNF-a[34,35]. The upregulation of
MAPKs activity and pro-inflammatory cytokines pro-
duction results in the direct serine phosphorylation of
the insulin receptor substrate (IRS)-1, which inhibits
phosphorylation on its tyrosine residues and down-
stream insulin signal transduction [36,37]. The inhib-
ition of signaling downstream of the insulin receptor is
a primary mechanism through which inflammatory sig-
naling leads to insulin resistance [38].
These clearly indicated that inflammatory pathways
are major contributors in the induction of insulin resist-
ance [22]. Since TLRs play an essential role in the
inflammatory pathways, then it would be conceivable to
assume that TLRs may participate in the induction of
insulin resistance; large numbers of evidences support-
ing this view have been published [26,39–41].
It has long been established that exercise has anti-
inflammatory effects, and therefore, can help prevent
chronic inflammatory diseases [5,6,42]. Regular
physical activity has been considered as a non-
pharmacological therapy to treat obesity and lead a
reduction in the signal activation of the inflammatory
pathways that can lead to insulin resistance [7,43].
Anti-inflammatory effects of exercise
Given the important role of innate immune cells in
diverse inflammatory states, the relationship between
inflammation and chronic illness and the anti-
inflammatory properties of chronic physical activity, it
is now recognized that physical activity is efficient to
prevent, or at least delay, the onset of metabolic disor-
ders [42]. Regular low- and moderate-intensity exercise
has been associated with reduction of circulating pro-
inflammatory markers and improved immune function
[5]. The main mechanisms by which exercise exerts
functional changes in the immune system resulting in
anti-inflammatory effects are (Figure 2): (1) a reduc-
tion in adipose tissue mass; (2) the development of an
anti-inflammatory environment through the release of
anti-inflammatory cytokines; and (3) changes in TLR
expression/activity in innate immune cells.
Reduction of adipose tissue mass
Physical activity may decrease the secretion of pro-
inflammatory adipokines to counteract chronic low-
grade inflammation state, which seems more related
to the reduction of abdominal fat storage [5] than to
global body weight loss [44,45]. This is not surprising
given the central role of adipose tissue in the develop-
ment of chronic low-grade inflammation as men-
tioned previously and the well-known effect of
physical activity on fat loss.
Anti-inflammatory environment development
During and after exercise, circulating IL-6 levels
increase and contracting skeletal muscles contribute to
this increase, which seems gradual with the duration
and intensity of the exercise session [46]. The tempor-
ary increase of muscle-derived IL-6 has been associated
with the activation of an anti-inflammatory response,
resulting in higher levels of IL-10 and IL-1 receptor
antagonist in the blood [47]. In vitro, IL-6 suppressed
LPS-induced production of TNF-aand IL-1bby
PBMCs [48]. However, while the development of an
anti-inflammatory environment is most pronounced fol-
lowing long term continuous exercise, anti-inflammatory
Figure 2. Anti-inflammatory effects of exercise.
Exercise counteracts the systemic inflammation through 1. The reduction in adipose tissue mass which favors the polarization of
macrophages to an anti-inflammatory state limiting cytokine production. 2. The down regulation of TLR receptors associated to
decreased inflammatory monocytes. 3. In response to exercise stimulation, the inflammatory state is modulated by increased levels
of IL-6 derived from muscle and preventing the cytokine release sustained in obesity state. M1: type 1 macrophages; M2: type 2
macrophages; IL6: Interleukin–6.
4 N. COLLAO ET AL.
effects have also been observed after short duration exer-
cise with low to moderate intensity, which does not elicit
IL-6 secretion [6]. The latter indicates that there should
be other mechanisms besides IL-6 muscle production to
reduce inflammation after exercise.
Changes in TLR regulation in innate immune cells
In the next sections, we will present the effects of acute
and chronic exercise on TLR expression and activity in
innate immune cells. Amongst the TLRs that have been
identified, TLR2 and TLR4 have received the most
attention in the exercise science field [49–53].
Acute exercise
The first report in CD14þmonocytes described a
reduction in TLR2 and TLR4 cell surface expression
following a single bout of 1.5 h endurance exercise
(65% VO
2
max) performed at 34 C in healthy sub-
jects [52]. Similarly, a reduction in TLR4 expression at
the cell surface of CD14þmonocytes was found after
45 min of endurance exercise at 75% VO
2
max [54]
and reduced CD14þmonocyte TLR4 expression in
healthy men after 1.5 h endurance exercise at 75%
VO
2
max, while TLR2 expression remained unchanged
in the latter study [55]. TLR4 expression returned to
baseline values after 4 h post exercise, which highlights
the acute effect of exercise on TLR4 expression [55].
Conversely, an acute bout of resistance exercise (9
exercises, 3 sets, 10 repetitions, 80% of 1RM) did not
induce changes in CD14þmonocyte cell surface
TLR4 expression in either untrained or trained older
women [50], suggesting that the type of exercise and/
or the inflammatory state related to age might induce
different TLR regulation. As age does not influence
CD14þcell surface TLR4 expression and inflamma-
tory cytokines production after a submaximal tread-
mill test [51], it seems that the type of exercise is a
more critical parameter to modulate TLR expression
than age. In favor of this hypothesis is the lack of
effect of a high intensity interval training session on
TLR4 expression in type 2 diabetic patients [56]. In
the later study, only TLR2 expression was lowered in
CD16þpro-inflammatory monocytes and classical
monocytes after the exercise session. This may suggest
that high intensity interval training is not as efficient
as continuous endurance exercise at reducing TLR4
expression and that exercise duration is an important
parameter. An acute bout of aerobic exercise (30-min,
70% of VO2 peak) on patient with systemic lupus
erythematosus (SLE) an autoimmune disease charac-
terized by persistent systemic inflammation; led to the
down-regulation of TLR3 gene expression on leuko-
cyte followed by an up-regulation at recovery, suggest-
ing that an anti-inflammatory response occurred
immediately after exercise [57]. TLR3 is expressed in
several immune cells, such as monocytes, dendritic
cells and natural killers (NK) cells, which is associated
with inflammatory process that may participate in the
development of T2D and its complications [58].
Further studies are needed to assess the association of
exercise-induced cellular changes in TLR3 expression
with the potential benefits of exercise.
Chronic exercise training
Twelve weeks of combined resistance and endurance
training resulted in reduced CD14þmonocytes cell
surface TLR4 expression [53] and both TLR2 and
TLR4 expression [59] in sedentary subjects compared
to pre-training values. In addition, the intervention
lead to a reduction in CD16þ“pro-inflammatory”
monocytes/classical monocytes ratio, suggesting a
switch to a more anti-inflammatory monocyte profile
[59]. While a single resistance exercise session does
not seem to regulate TLR4 expression, a 10-week
intervention comprised of resistance exercises exclu-
sively down-regulated TLR4 mRNA monocyte expres-
sion in older women compared to their sedentary
counterparts [49]. Eight weeks of resistance training
decreased TLR2 and TLR4 expression in PBMCs of
healthy elderly subjects [25]. This down-regulation was
associated with an increase in HSP70 protein content
and a reduced NF-jB signaling and pro-inflammatory
cytokines production [25]. Therefore, training interven-
tions of at least 8 weeks seem to reduce TLR expression
and downstream inflammatory pathways in PBMCs.
Shorter training interventions have elicited conflicting
results. Six sessions of high intensity interval training
over 2 weeks increased monocyte TLR4 surface expres-
sion [60]. A daily aerobic exercise program for 15 days
did not induce changes in TLR2-4 protein content in
PBMCs of type 2 diabetic individuals [61]. Ten sessions
of high intensity interval training or of moderate-
intensity continuous training over 2 weeks both reduced
TLR4 surface expression in monocytes and TLR2-4 sur-
face expression in lymphocytes of pre-diabetic subjects
[62]. However, decreased TLR4 surface expression was
only reported in neutrophils after moderate-intensity
continuous training [62]. In an animal model, the
mRNA expression of TLR3, which is implicated in the
recognition of respiratory viruses was down-regulated
by exercise training in both blood monocytes and pul-
monary alveolar macrophages (PAMs) [63]. The
observed down-regulation of TLR3 has also been
INTERNATIONAL REVIEWS OF IMMUNOLOGY 5
reported in regularly exercising humans [52]; whereas
the expression of TLR6 was down-regulated only in
monocytes after the training period.
Altogether, it is difficult to get a clear picture on
TLR regulation after a 2-week intervention.
Possible TLR regulation mechanisms
One of the proposed mechanisms underlying the anti-
inflammatoryeffectsofexerciseisareductioninTLR
activation and expression in innate immune cells [6].
This reduction has been observed after both acute bouts
of exercise and longer duration training studies [53–55]
but the molecular mechanisms behind exercise-induced
TLR2 or TLR4 down-regulation are not fully understood.
The possible mechanisms are presented in Figure 3.
TLR tolerance
Reduced expression of TLR2 and TLR4 may occur as
a result of low-dose exposure to exogenous ligands
including LPS, peptidoglycan and double stranded
RNA as well as endogenous ligands such as HSP, all
of which may increase during and after exercise [64].
Exposure to these ligands may induce TLR tolerance,
which concretely translates into hypo-responsiveness
to subsequent agonism and potential decrease in
expression [62]. Important endogenous ligands that
may influence TLR expression are the HSPs. These
proteins are present in all cells and are up-regulated
during physiological stress. The HSPs act as chaper-
ones to guide the synthesis, transportation and deg-
radation of proteins [65]. In addition, it is suggested
that specific HSPs, e.g. HSP60, act as activators of
Figure 3. Possible TLR regulation mechanisms.
A reduced TLRs response to certain ligands may be related to repeated exposure which induce tolerance. Receptor sheeding medi-
ated by matrix metalloproteinases could lead to TLRs downregulation and increased soluble TLR. The internalization of TLRs might
promote its lysosomal degradation regulated by Rab7b. Exercise has elicited modulatory effects on TLR miRNA levels which is one
of the possible transcriptional mechanism involved. Exercise exerts positive effects on gut microbiota due to the reduction of endo-
toxinemia concentrations in favors to gut permeability and diversity that contributes to TLR downregulation. LPS:
Lipopolysaccharides; TLR2/4: Toll-like receptors 2/4; TIRAP: TIR Domain-Containing Adaptor Protein; MYD88 myeloid differentiation factor
88; MMP-9: Matrix metallopeptidase 9; sTLR2: soluble TLR2; Rab7b: Ras-related protein Rab-7b; miRNAs: MicroRNAs.
6 N. COLLAO ET AL.
TLR4 in a similar manner to LPS. Repeated monocyte
exposure to HSP60 induces a tolerance to HSP and a
‘cross-tolerance’to LPS stimulation [65], subsequently
reducing TLR4 activation [55]. HSP70 is one of the
numerous DAMPs recognized by both TLR2 and
TLR4 [66]. An increase of HSP70 concentrations after
exercise has been reported in both animals [67,68]
and humans [69,70] in an intensity- and frequency-
dependent way [71,72]. HSP70 decreases NF-jB
activity [73,74], thereby reducing the expression of
pro-inflammatory cytokines such as TNF-a, IL-6, or
IL-1b[75]. In line with the proposed mechanism of
TLR tolerance, exercise-induced decrease of TLR2 and
TLR4 was inversely correlated with the increase of
HSP70 in PBMCs [76].
Receptor shedding
Shedding of TLR is another proposed mechanism that
can explain the reductions in TLR at innate immune
cells surface after exercise [6]. Proteolytic cleavage of
transmembrane proteins is a common post-translational
mechanism, that can specifically occur at the ectodo-
main level [77]. Matrix metalloproteinases are a class of
enzymes that participate in ectodomain shedding and
their activation appears to be responsible for shedding
TLR2 from immune cells, which leads to an increase in
soluble TLR2 (sTLR2) and down-regulation of TLR2
activation [78]. Acute exercise increases plasma matrix
metalloproteinase-9 levels, which could contribute to the
decrease in TLR expression measured after exercise [79].
However, to date, sTLR2 levels have not been deter-
mined after exercise. The hypothesis of exercise-induced
TLR shedding remains to be tested.
Receptor internalization
>The internalization of TLRs could be another pos-
sible regulatory mechanism for the decreased expres-
sion at the cell surface of innate immune cells after
both acute exercise and exercise training. This mech-
anism involves Rab7b small GTPase, which localizes
to lysosome-associated subcellular compartments and
is selectively expressed in monocytic cells [80]. Like
its homologous Rab7, Rab7b regulates the later stages
of the endocytic pathway and is involved in the trans-
port and lysosomal degradation of several kinds of
receptors [81,82]. In macrophages, decreased Rab7b
expression by siRNA resulted in up-regulation of LPS-
induced pro-inflammatory cytokine production [83].
Reciprocally, overexpression of Rab7b suppressed
TLR4-mediated production of cytokines as well as
activation of intracellular signaling molecules. In
Rab7b-silenced cells, the expression of TLR4 was
higher than in control cells, and translocation of
TLR4 from early endosomes to late endosomes/lyso-
somes was delayed. This study clearly demonstrates
that Rab7b could serve as a negative regulator of
TLR4 signaling in macrophages by accelerating lyso-
somal degradation of TLR4 and decreasing TLR4
expression level at the plasma membrane, thus provid-
ing the first evidence for a role of Rab proteins in
TLRs signaling [83]. However, the detailed membrane
trafficking process mediated by Rab7b may need fur-
ther investigation as well as the possible role of phys-
ical activity could play in this mechanism of TLR
internalization. Tracking differences in cell surface
versus internalized TLRs using recently-developed
imaging flow cytometers may be one technique that
could also help determine if TLRs is internalized
after exercise.
Modulation of gene expression
It is vital for the body to regulate the expression of
genes in the process of adapting to changes in the
environment, such as exercise [84]. Thanks to the
microarray technology, it is nowadays possible to
measure genome-wide changes in gene expression in
response to exercise and to better understand the pos-
sible transcriptional mechanisms of exercise-related
transient anti-inflammatory process. Acute exercise-
induced changes in anti-inflammatory gene expression
may have the potential to modulate TLRs expression
and function [85]. Several studies quantified the gene
expression levels of TLRs in immune cells after exer-
cise [86–89]. After 1 h cycling at 70% VO2max, TLR2
mRNA levels, but not TLR4 or TLR6 mRNA, were
lower in PBMCs than before exercise [86]. After
marathon running, TLR7 mRNA levels were decreased
in PBMCs independently of body composition or
training status while TLR4 mRNA was decreased only
in lean non endurance trained individuals with no
modification of TLR2 mRNA [87]. The day after,
TLR4 and TLR7 mRNA levels were up regulated com-
pared to baseline. In two other studies, TLR4 mRNA
levels increased immediately after and 2 h after acute
eccentric exercise [88,89]. Importantly, those changes
in gene expression were not systematically followed by
similar changes in protein expression. Therefore, more
research is needed to understand the impact of the
observed changes in gene expression, and to elucidate
the complex interaction between immune cells and
the inflammatory response after exercise.
INTERNATIONAL REVIEWS OF IMMUNOLOGY 7
Gut microbiota
The modification of gut microbiota has emerged as a
new factor by which exercise may promote beneficial
health effects [90]. The gut microbiota is a set of
microorganisms living throughout the gastrointestinal
tract of mammals, and which increase in number and
diversity from the stomach to the colon. It has been
estimated that human microbiota consists of 1014
cells (10 times the total number of cells in the human
body) [91]. Changes in microbiota composition have
been associated with obesity as obese individuals have
different and altered gut microbiota composition com-
pared to lean individuals [92]. The microbiota also
impacts host immune status and dysbiosis-related
inflammation can augment insulin resistance, inde-
pendently of obesity [93]. It has recently been shown
that gut bacteria can initiate the inflammatory state of
obesity and insulin resistance through the activity of
LPS [94].
Exercise seems to exert positive effects by reducing
the level of endotoxinemia due to obesity-related gut
permeability and by increasing microbial diversity,
thereby reducing TLR signaling activation [95–98].
The microbiota per se modulates the expression of
TLRs through the microbe-associated molecular pat-
tern (MAMP), leading to the activation of the nuclear
factor-kappa B pathway and activation of T-cells [99].
Also, metabolic by-products of the microbiota can be
implicated in mucosal tolerance via induction of T-
regulatory cells [100].
Theimpactofphysicalexerciseongutmicrobiotahas
only few controlled studies on humans [98,101–103]
have been conducted in the attempt to confirm the find-
ings of studies on animals [95–97,104–106], which have
been carried out in greater numbers.
The potential mechanisms involved in the effects of
exercise on the gut microbiome include; suppression
of TLRs signaling pathway in the liver, muscle, and
adipose tissue by reducing lipopolysaccharide (LPS)
serum levels [107]; elevated production of SCFAs via
AMPK activation [108]; increase in fecal bile acids; as
exercise-amount and -intensity increase [109]; increase
of immunoglobulin A (IgA) [110] production and a
reduced number of B and CD4 þT cells; weight loss
[111]; myokines (IL-6, IL-10, IL-1ra, TNF-R) releasing
during exercise and reduction in intestinal transit
time influencing microbiota composition [112].
All together, these observations reveal that the
effects of exercise go beyond changes in host tissue
metabolic function, and that repeated exercise training
alters the gut microbial composition and diversity in a
way that opposes dysbiosis, indicative of obesity [113].
Manipulation of gut microbiota by modifying diet or
exercise habits could be a powerful tool in the future
to prevent or treat several diseases, where a complete
dose-response analysis between exercise levels and
their beneficial alterations in microbial composition is
yet to be fully explored.
MicroRNAs
MicroRNAs (miRNAs) have emerged as key regulators
and as an essential part of the networks involved in
regulating TLR-signaling pathways during and after
exercise [114] miRNAs are a class of small noncoding
RNAs (about 22 nucleotides in length) that regulate
gene expression by binding to the 30-untranslated
regions of target messenger RNAs, typically resulting in
protein translation repression or mRNA degradation
[115]. Growing evidence indicates a crucial role for
miRNAs in modulating immune functions in response
to exercise. Radom-Aizik and colleagues subjected
healthy young men to cycle ergometer exercise to
investigate the response of miRNAs in circulating cell
populations. Blood sampling immediately after exercise
revealed differential expression for 38 miRNAs in neu-
trophils [116], 34 miRNAs in PBMCs [117], 23
miRNAs in natural killers cells [118] and 19 miRNAs
in monocytes [119], with many of them playing a role
to regulate TLRs and inflammatory processes [120,
121]. In addition, Tonevitsky et al. found that specific
miRNA-mRNA regulatory networks were dynamically
regulated during exercise and recovery in white blood
cells [122]. Exercise was able to modulate the levels of
miR-21, miR-24-2, miR-27a and miR-181a, all of them
regulating the mRNA levels of genes involved in proc-
esses relevant to exercise response, including apoptosis,
immune function, protein membrane trafficking and
transcription regulation [122].
In the last decade many reports have confirmed that
miRNAs enter the circulation system including blood,
plasma, serum and other body fluids. Extracellular/cir-
culating-miRNAs (c-miRNAs) have received attention
as potential biomarkers of physical fitness, performance
potential and training adaptation [123].
The first investigation of the effect of acute exercise
on plasma ci-miRNAs profile was in 2011 by Baggish
et al. [124] immediately after exercise, ci-miRNAs
-146a, -222, -21, and -221 increased in plasma. Also
linear correlation was observed between the expression
level of miRNA-146a and the aerobic performance par-
ameter VO2max, suggest its potential as a biomarker
for cardiorespiratory fitness. Immediately after a mara-
thon running, expression on c-miRNAs has been
8 N. COLLAO ET AL.
observed in different studies showing an increases in
miRNA-1 and miRNA-133a [124–126], where de
Gonzalo-Calvo et al. [127] describe no changes in these
miRNAs, however observed a downregulation of
inflammation-related ci-miRNA-106. Nielsen et al.
[128] demonstrated that acute endurance training
robustly modifies miRNA expression patterns, changing
expressions of ci-miRNA-188 in plasma. Wardle et al.
[129] investigated whether levels of c-miRNAs differ
between endurance-trained and strength-trained
cohorts of elite male athletes. Plasma levels of miR-21,
miR-221, miR-222 and miR-146a were significantly
higher in endurance athletes than in strength athletes.
Taken together, the miRNAs seem to be important
in response to exercise but their exact implication on
TLRs regulation needs further investigation.
Conclusion and future perspectives
Physical exercise is a powerful preventive tool and treat-
ment for several diseases as it induces metabolic and
immune effects that provide health benefits. Exercise is
known to reduce inflammation; however, the underlying
mechanisms responsible are not fully elucidated. Here,
weemphasizedtheroleofTLRsinobesityandMetS
and how exercise can specifically regulate TLRs expres-
sion and activation. A reduced expression of TLRs has
been found in innate immune cells after exercise, which
probably contributes to the protective effect of exercise
against insulin resistance and the prevention of the
development of metabolic diseases, although more
research is needed in individuals suffering from obesity
and/or MetS. For example, it is currently unclear what
type and intensity of exercise are the most effective in
those individuals. In addition, it remains to be deter-
mined whether exercise decreases TLRs cell surface
expression in innate immune cells by down-regulating
TLR gene expression, TLRs shedding from the cell sur-
face and/or internalization by the cell. Of particular
interest are miRNAs and gut microbiota as they both
can be modulated by physical activity, thereby constitut-
ing novel therapeutic targets for treating obesity
and MetS.
Authors’contributions
NC conducted the research protocol, assessed the data
of the “Review”independently from the other
reviewers, and wrote the manuscript. IR critically
revised and synthesizing the manuscript. MF critically
revised the manuscript. LD critically revised, edited
and synthesizing the manuscript. HZ critically revised
and approved the final manuscript.
Availability of data and materials
Data sharing is not applicable to this article as no
datasets were generated or analyzed during the cur-
rent study.
Ethics approval
Not applicable.
Consent for publication
Not applicable.
Competing interest
The authors, Nicolas Collao, Isabel Rada, Marc
Francaux, Louise Deldicque and Hermann Zbinden-
Foncea, declare that they have no competing interests.
Funding
This project was funded by the Chilean National Science
and Technology Fund, FONDECYT N11150576.
ORCID
Nicolas Collao http://orcid.org/0000-0001-6293-4018
Marc Francaux http://orcid.org/0000-0001-8182-1588
Hermann Zbinden-Foncea http://orcid.org/0000-0002-
9643-1037
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