Quantifying the Effects of Different Treadmill Training Speeds and Durations on the Health of Rat Knee Joints
Rios et al. Sports Medicine - Open
Quantifying the Effects of Different Treadmill Training Speeds and Durations on the Health of Rat Knee Joints
Jaqueline Lourdes Rios 0 1 2
Kevin Rudi Boldt 1
James William Mather 1
Ruth Anne Seerattan 1
David Arthur Hart 0 1
Walter Herzog 0 1
0 McCaig Institute for Bone and Joint Health, University of Calgary , Calgary, AB , Canada
1 Faculty of Kinesiology, Human Performance Laboratory, University of Calgary , 2500 University Drive NW, Calgary, AB T2N 1N4 , Canada
2 CAPES Foundation , Brasilia, DF , Brazil
Background: Walking and running provide cyclical loading to the knee which is thought essential for joint health within a physiological window. However, exercising outside the physiological window, e.g. excessive cyclical loading, may produce loading conditions that could be detrimental to joint health and lead to injury and, ultimately, osteoarthritis. The purpose of this study was to assess the effects of a stepwise increase in speed and duration of treadmill training on knee joint integrity and to identify the potential threshold for joint damage. Methods: Twenty-four Sprague-Dawley rats were randomized into four groups: no exercise, moderate duration, high duration, and extra high duration treadmill exercise. The treadmill training consisted of a 12-week progressive program. Following the intervention period, histologic serial sections of the left knee were graded using a modified Mankin Histology Scoring System. Mechanical testing of the tibial plateau cartilage and RT-qPCR analysis of mRNA from the fat pad, patellar tendon, and synovium were performed for the right knee. Kruskal-Wallis testing was used to assess differences between groups for all variables. Results: There were no differences in cartilage integrity or mechanical properties between groups and no differences in mRNA from the fat pad and patellar tendon. However, COX-2 mRNA levels in the synovium were lower for all animals in the exercise intervention groups compared to those in the no exercise group. Conclusions: Therefore, these exercise protocols did not exceed the joint physiological window and can likely be used safely in aerobic exercise intervention studies without affecting knee joint health.
Osteoarthritis; Cyclical loading; Aerobic exercise; Joint health; Histology; Animal model
A stepwise increase in the speed and duration of
exercise did not lead to osteoarthritis-like changes in
Chronic exercise appeared to produce a protective
effect on the knee.
Working within an optimal physiological exercise
window is beneficial for joint health across the life
Exercise has been used to promote health and fitness as
far back as 2500 BC [
]. However, in the past years,
exercise has been postulated to act like a drug [
] and, as
such, provides advantages and risks to an individual’s
health and fitness. Specifically, aerobic exercise has been
shown to lead to increased bone mineral density, insulin
sensitivity, high-density lipoprotein cholesterol levels,
resting and maximal stroke volume, maximal oxygen
uptake, and basal metabolism [
]. Exercise has also been
shown to reduce body fat, fasting blood insulin levels,
low-density lipoprotein cholesterol levels, resting heart
rate, and systolic and diastolic blood pressure [
However, the effects of exercise on joint health remain
In a prospective survey, aimed at examining the
relationship of self-reported physical activity and
physiciandiagnosed osteoarthritis (OA) from 1970 to 1995, a
positive association between OA incidence and physical
activity was found. Running was suggested to lead to an
increased risk for developing OA in some studies [
but not in others [
]. Chakravarty et al. [
] did not
find an increased risk for knee OA in masters runners
compared to controls over an 18-year period [
Miller et al. [
] suggested that running does not increase
the risk for knee OA compared to walking. However,
Cheng et al. [
] reported a positive association between
running more than 32 km per week and
cliniciandiagnosed knee OA.
If exercise is thought to act like a drug preventing joint
disease, the dosage may be critical to its success. It has
been shown that moderate exercise, such as walking and
running, exerts added loading to the knee joint, and
cyclical loading is thought to be vital for maintaining
cartilage integrity/homeostasis and healthy joints [
contrast, in vivo and vitro studies with excessive
repetitive joint loading have been implicated with the
development of OA [
]. Joints are thought to be designed
to operate within a “physiological window” to maintain
proper function and allow for positive adaptations .
Loading outside this window may put the joint, and
specifically the cartilage, at risk for degeneration [
In rodent model of exercise, for example, Galois et al.
] suggested that different levels of treadmill training
may have different influences on the severity of chondral
lesions in anterior cruciate ligament transection (ACLT)
model of osteoarthritis in Wistar rats; while slight to
moderate levels seemed to be beneficial to the knee
cartilage health, more strenuous exercise may be
detrimental. Controversially, Yang et al. [
] showed that
treadmill exercise up to 1 h per day, 5 days a week, for
8 weeks, in Sprague-Dawley rat model of monosodium
iodoacetate-induced OA, has a chondroprotective effect,
and this effect was more prominent in rats that fulfill in
three times per day the 1 h treadmill exercise, suggesting
that an adaptive phase training may be an important
factor in protecting the knee joint from OA-like changes.
Wistar rats, without previous lesion in the knee, have
also been randomly assigned to a sedentary control
group, a low-intensity running, a medium-intensity
running, and a high-intensity running [
]. As a result, rats
in the high-intensity running demonstrated OA-like
changes in their knees, while rats in the other running
groups did not. Moreover, in a study in rodent models
where the animals were allowed minimum [
] or not
allowed an adaptive phase [
], they have demonstrated
OA-like changes in their knees. However, it is unknown
how much exercise is too little or too much. Therefore,
the purpose of this study was to determine the effects of
a stepwise increase in speed and duration of treadmill
training on knee joint integrity and to identify the
potential treadmill training threshold for the development of
joint damage in the rat knee. We hypothesized that
excessive chronic treadmill training leads to the
development of OA-like changes in the knee, while moderate
levels of treadmill training maintain cartilage integrity
and induce positive adaptive responses. The key
outcome measures were joint integrity, body composition,
gene expression patterns, and blood-based and synovial
The aim of this study was to determine the effects of a
stepwise increase in speed and duration of treadmill
training on knee joint integrity and to identify the
potential treadmill training threshold for the development of
joint damage in the rat knee.
Twenty-four 10- to 14-week-old male Sprague-Dawley
rats were housed individually and randomized into four
groups: moderate duration exercise (MD, n = 6), high
duration exercise (HD, n = 6), extra high duration
exercise (EHD, n = 6), or no exercise (control, n = 6). Rats
were fed (ad libitum) a standard chow diet (Diet #5001,
Lab Diet, USA). A minimum sample size of five rats per
group is based on the ability to detect a minimal
meaningful difference in histological scoring of the knee joint
to provide an α = 0.05 and a power of 80%. Calculation
of sample size was performed using G*Power Software
(version 3.0.10, Germany) [
]. Data for sample size
calculations were obtained from a previous study [
experiments were approved by the University of Calgary
Life and Environmental Sciences Animal Care
Committee, and all methods were conducted in accordance with
the animal welfare regulations and guidelines at the
University of Calgary.
Exercise Training Protocol
Following 1 week of acclimatization to the housing
environment, rats were exposed to their respective exercise
programs (see Table 1 for details of the exercise programs) on
a Columbus Instruments Exer-3R treadmill (Columbus,
OH, USA) for 12 weeks. The moderate duration group
progressively built up to 30 min of treadmill training each
day, five times per week at 25 m/min. The high duration
group built up to 60 min of treadmill training per day for
5 days per week at 25 m/min. Rats in the extra high
duration protocol reached 60-min training sessions 7 days
per week at 25 m/min. In weeks 10, 11, and 12, these rats
trained twice, three times, and four times for 1 h daily,
respectively. This last training intervention has previously
been used to elicit overtraining in Wistar rats [
Animals in the control group were placed on the
treadmill 5 days a week and completed 15 min of
exercise at 10 m/min once per week. This was done to
account for the stress of handling and avoiding
confound results. A shock grid at the back of the
treadmill was used to prevent animals from falling behind
the pace of the treadmill.
Body mass was measured at the beginning of each week.
Body mass for each animal was normalized to that of
week 1 (familiarization week) and was expressed as the
percent increase in body mass from that initial value.
One week after completing the 12-week training
protocol, and immediately prior to sacrifice, rats were lightly
anesthetized with isoflurane and body composition was
measured using dual X-ray absorptiometry (DXA) with
software for small animals (Hologic ODR 4500; Hologic,
MA, USA). An average of three scans for each animal
was used for analysis.
Following 12 h of fasting, rats were anesthetized with
isoflurane and a blood sample was collected by cardiac
puncture. Blood was centrifuged at 3000 rpm for
15 min at 4 °C and serum stored in aliquots at − 80 °C
until analyzed. Rats were sacrificed by heart excision.
Serum cytokines and adipokines were quantified using
a Rat 27 Multiplex Discovery Assay with Luminex®
xMAP technology (eotaxin, EGF, fractalkine, IL-1α,
IL-1β, IL-2, IL-4, IL-5, IL-6, IL-10, IL-12(p70), IL-13,
IL-17A, IL-18, IP-10/CXCL10, GRO/KC, IFN-γ,
TNFα, G-CSF, GM-CSF, MCP-1, leptin, LIX, MIP-1α,
MIP2, RANTES, VEGF; Eve Technologies, Calgary, AB,
Knee joints were collected from both hind limbs. The left
knee was harvested by cutting the femur and tibia/fibula
2 cm above and below the joint line. Excess muscles were
removed, and joints were fixed in a 10% neutral buffered
formalin solution (Thermo Fisher Scientific, MA, USA)
for 14 days at room temperature. Knees were then
decalcified at room temperature, using Cal-Ex II solution (10%
formic acid in formaldehyde, Thermo Fisher Scientific).
The decalcification solution was changed daily. The end of
decalcification was determined by chemical testing with a
5% ammonium oxalate solution (Thermo Fisher Scientific)
until no precipitate was detected for 5 days (on average
21 days). Samples were then processed using an automatic
paraffin processor (Leica TP 1020, Leica Microsystems
Inc., Concord, Ontario, Canada). They were dehydrated in
a graded series of alcohols, cleared in xylene, and
infiltrated with 50% Paraplast X-TRA® wax (Thermo Fisher
Scientific) and 50% Paraplast Plus® wax (Thermo Fisher
Scientific). Further, the left knee joints were embedded in
paraffin wax and stored at room temperature until
sectioning. Serial, sagittal plane sections of 10-μm thickness
were obtained using a Leica RM 2165 microtome (Leica
Biosystems, Nussloch, Germany). Sections were mounted
onto Super Frost plus slides (Thermo Fisher Scientific)
and allowed to dry at 40 °C for 7 days. Alternate slides
were stained sequentially with hematoxylin, fast green,
and safranin-O (Thermo Fisher Scientific) using an auto
stainer (Leica ST 5010, Leica Biosystems). Sections were
then dehydrated in a graded series of alcohols, cleared in
xylene, and mounted with Cytoseal 60 mounting media
(Thermo Fisher Scientific) using an auto cover slipper
(Leica CV 5030, Leica Biosystems). Slides were dried at
room temperature for 7 days before being evaluated using
a light microscope (Zeiss Axiostar plus, Carl Zeiss Inc.,
Ontario, Canada). Two independent graders scored all
histological sections in a blinded manner using a modified
Mankin Histology Scoring System [
Research Society International (OARSI) histologic [
subscores for bone changes, synovial thickening, and
meniscus were also determined for each joint. The total
modified Mankin score for each animal represents the
sum of all Mankin scores and OARSI subscores. Tibial
and femoral cartilage thickness were determined from
The right knee joints were opened to collect the
synovium, menisci, fat pad, and patellar tendon. The tissues
were snap-frozen in liquid nitrogen and stored at − 80 °C
for RNA isolation and subsequent RT-qPCR analysis.
Genes analyzed for the patellar tendon and synovium were
Col-1, Col-3, iNOS, COX-2, IGF-1, IL-1, IL-6, and TGF-β.
Genes analyzed for the fat pad were iNOS, PPAR-γ,
COX2, IL-1B, IP-10, leptin, TF, TFPI, and VEGF. Genes analyzed
for the menisci were Col-1, Col-3, iNOS, PPAR-γ, COX-2,
IGF-1, IL-1, IL-6, TGF-β, IP-10, leptin, TF, TFPI, VEGF, and
PRG4 (see Table 2 for details). Synovial fluid was also
collected from the right knee shortly after sacrifice using the
Whatman chromatography paper method [
were weighed, diluted 1:50, and stored at 4 °C overnight.
After 24 h, samples were centrifuged and stored at − 80 °C
until analysis. Synovial fluid cytokines and adipokines were
quantified using a Rat 27 Multiplex Discovery Assay with
Luminex® xMAP technology (eotaxin, EGF, fractalkine,
IL1α, IL-1β, IL-2, IL-4, IL-5, IL-6, IL-10, IL-12(p70), IL-13,
IL-17A, IL-18, IP-10/CXCL10, GRO/KC, IFN-γ, TNF-α,
GCSF, GM-CSF, MCP-1, leptin, LIX, MIP-1α, MIP-2,
RANTES, VEGF; Eve Technologies).
The right tibia was stored at − 20 °C until assessment of
the biomechanical properties of cartilage was performed.
A spherical indenter (r = 175 ± 2.5 μm) made of a stainless
steel 316 L shaft and a spherical glass bead was installed
under the multiaxial load cell (force resolution: Fz =
3.5 mN and Fx = Fy = 2.5 mN) of a three-axis mechanical
tester (Mach-1 v500css, Biomomentum, QC, Canada).
The tibia was fixed in a sample holder using dental
cement. The sample was then immersed into a testing
chamber that contained PBS and was equipped with a
camera registration system (Biomomentum). A position
grid was superimposed on the image of the tibia articular
surface for a mechanically controlled surface mapping
]. Stress relaxation tests for cartilage properties were
performed on 11 sites each for the medial and lateral tibial
plateau using the automated mapping system, and Young’s
modulus were calculated for each test site.
Non-parametric Kruskal-Wallis testing was used to
determine differences between the four animal groups
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CTA ACC AAG GCT GCA AGA TG
CAG TAC ACT ACA TCC TGA CC
GCA TTG TGG ATG AGT GTT GC
CAG GCA TCC TCA GCA GCA GA
AAC CTG CTG GTG TGT GAC GTT C
TCA CAG AAG GAG TGG CTA AG
AAG GCA CAA GAC TCT GAC AC
CAA GGC TTC CCA ATT CTC
CCT GTG GCT TTG GTC CTA TCT G
CTT GGC CAT ATT TAT AGC TGT CAT TAT T
AGG GCG TTG CAT CCA AGA A
GAC AAT CTT GGA GTG GCA AC
CCG AGG AAG CTA TGT GTA AG
ACG GCA GCT GTA CAT TGA CT
CGC CTG CTG CCT TCC TTG G
GAC ATG TAG ACT CTT TGC GGC
ATC TGT CCA CCA GTG CTT CC
CGT CAA CAC GTA TCT CAT GG
GGT CTT GTT TCC TGC ACT TC
GGC TCC TAA GAA CAA GAA TG
CAG CAC GAG GCT TTT TTG TTG T
ACC ACA GTG AGG AAT GTC CA
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ATA ATC ACA GTT GCA GGT GGC
GCT TCA TAG GTC CAG TTC AC
AGC CAG TGT AGG TGA AGG TC
CAG GAG CGC ACG ATC ATG TT
for all variables (RT-qPCR analyses, body fat, body
mass, cartilage thickness, Mankin/OARSI score,
equilibrium Young’s modulus, and protein assays). If
significant (p < 0.05), post hoc testing using the Mann-Whitney
U test was used to indicate differences between groups.
Further, estimates of effect size were calculated using
the ANOVA univariate approach on SPSS (version 22).
Partial eta squared (partial η2) were reported to
provide small (partial η2 = 0.01), medium (partial η2 =
0.06), and large (partial η2 = 0.14) effects [
Rats in the control group traveled a distance of 150 m
per week from week 2 to week 12, with a total distance
of 1.65 km at the end of the experimental protocol. Rats
in the MD and HD groups had a gradual increase in
speed and duration from week 2 to week 5 (4 weeks of
adaptation), and then, they ran in a constant speed and
duration from week 6 to week 12, leading to a total
distance traveled of 37.9 and 69.9 km, respectively (Fig. 1,
Table 1). Rats in the EHD group have a stepwise increase
in distance traveled from week 2 to week 5, then a
constant distance traveled from week 6 to week 9, and
another stepwise increase in distance traveled from week
10 to week 12, leading to a total distance traveled of
162.8 km by the end of the exercise protocol.
Rats in all groups exhibited a gradual increase in body
mass during the intervention period. However, body fat
was significantly reduced by 4.5% (p = 0.046), 6.9% (p =
0.032), and 9.4% (p = 0.003) in the MD, HD, and EHD
group rats, respectively, compared to that in the control
group rats at the end of the exercise intervention period.
Body fat was also significantly reduced by 4.9% (p =
0.025) in the EHD rats compared to that in the MD
group rats (Fig. 1b, Table 3).
Knee Joint Integrity and Inflammatory Markers
There were no differences in cartilage integrity (Fig. 1c,
Table 3), mechanical properties, and cartilage
thicknesses (Fig. 2a, Table 3) between groups across all joint
locations. There were also no differences between
groups in gene expression levels for the fat pad, patellar
tendon, and menisci. Cyclooxygenase 2 (COX-2) levels
in the synovium were lower for the three exercise group
animals than those in the control group animals (p =
0.010) (Fig. 2b). Epidermal growth factor (EGF) levels in
the synovial fluid were significantly greater in EHD and
HD group animals than those obtained in the control
group animals (p = 0.039, p = 0.014, respectively; Table 4,
Fig. 2c). Serum interleukin-12 (IL-12) levels were
significantly higher for the EHD group animals than for the
control (p = 0.015) group and the HD (p = 0.008) group
animals (Table 4).
Inactivity is the fourth leading risk factor for global
]. A physically inactive lifestyle is associated
with the development of non-communicable diseases
], including OA [
]. In contrast, exercise is reported
to be a strong factor in health promotion and the
prevention or delay of many non-communicable
]. However, poor health outcomes have been
reported in athletes exposed to extra high levels of
], and it has been reported that
toplevel athletes have an increased risk of developing
knee OA . Consequently, exercise can be beneficial or
detrimental to the general health. Mechanical loading of
joints due to exercise may be minimal but sufficient to
maintain tissue homeostasis, it may be within the optimal
physiological window and help maintain tissue
homeostasis and produce positive tissue adaptations, or it may be
strenuous, thus exceeding the physiological window and
lead to disruptions of tissue homeostasis, leading to joint
disease. In this study, we investigated the effects of a
stepwise increase in speed and duration of treadmill training
on knee joint health in Sprague-Dawley rats.
We randomized 24 male Sprague-Dawley rats into
four groups: moderate duration exercise (MD), high
duration exercise (HD), extra high duration exercise (EHD),
or no exercise (control). We did not detect changes in
cartilage structure, mechanical properties, or thickness
between groups and across joint locations. Therefore,
the speed and duration of even the most strenuous
treadmill exercise protocol used here (up to 4 h of
exercise daily) was not detrimental to the knee joint cartilage
in these animals. Thus, the load applied to the knee
through the exercise intervention protocol provided here
was likely within the physiologic loading window that
has been suggested to allow for adaptation, remodeling,
and proper functioning of cells and tissues [
An aggressive endurance exercise protocol in 16- to
18-week-old Wistar rats has been shown to lead to
OAlike changes in the knee [
]. These rats were allowed
1 week of familiarization to the exercise protocol,
followed by exercising twice a day, 5 days per week, for
3 and 6 weeks, for a total of 30 and 55 km of distance
]. In our study, rats were allowed a
prolonged adaptation period and were introduced gradually
to the increasing speed and duration of the exercise
protocol, a factor which may be critical for joint health
outcomes. The gradual increase in speed and duration of
the exercise sessions may have allowed the cartilage to
adjust gradually to the increasing load requirements,
operating within the physiological window. The adaptive
training phase in our study consisted of 4 weeks, and
has been suggested to trigger increases in fitness and
health in Wistar rats [
], and may be an important
factor in the protection of the knee joint. It has been shown
that a gradual increase in running volume in Wistar rats
results in bone and cartilage remodeling by reducing
catabolic genes and increasing aggrecan expression [
However, to our knowledge, there is no study
systematically investigating adaptive training phase in order to
protect the knee joint, and this is a limitation of our
study. Additionally, in the present study, rats in the
EHD group doubled, tripled, and quadrupled the
amount of exercise in weeks 10, 11, and 12, respectively,
relative to weeks 5–9. If these levels of exercise were
sustained for an extended period, they may cause
damage to the knee. Additionally, the physiological window
for knee health is thought to be highly individual and
may be influenced by genetics, sex, lifetime loading
history, presence of prior injury/scar tissue, systemic
pathology, and local anatomy [
Rats from all groups exhibited a gradual increase in
body mass during the exercise intervention period. This
increase was consistent with the expected increase in
body mass of laboratory rats with age [
]. However, the
MD, HD, and EHD group rats were leaner than the
control group rats, and the EHD group rats were leaner
than the MD group rats at the end of the intervention
period. Body fat has been shown to decrease with
aerobic exercise in humans [
] and mice [
]. Since body
fat was reduced in the exercise group animals, but body
weight was similar across all groups, the lean body mass
(muscle mass) must have increased in the exercise group
animals compared to that in the control group animals
(not evaluated in our study). Results similar to ours have
been reported for human studies where strength and
*No statistics were computed due to extrapolated values and/or values out of
**Indicates p < 0.05.
SF: synovial fluid
endurance exercise did not produce differences in
body mass but resulted in a reduction in body fat for
individuals in an exercise program compared to
individuals in a non-exercising group [
]. A reduction in
body fat is considered a positive outcome for overall
health and has been shown to reduce risks for
diabetes, cardiovascular disease, metabolic syndrome, and
knee OA [
Exercise induces multiple biochemical changes that
may affect gene expression [
]. In the present study,
markers for oxidative stress (iNOS), collagen (Col-I and
Col-III), pro-inflammation (IL-1β, COX-2, IL-6, leptin,
IGF-1, IP-10), and anti-inflammation (TF, TFPI, VEGGF,
TGF-β) were evaluated for the knee joint tissues. Gene
expression in the fat pad, patellar tendon, and menisci
were similar across all groups. However, COX-2 mRNA
levels in the synovium were reduced for all animals in
exercise groups compared to those in the animals in the
control group. A reduction in COX-2 expression is
thought to be positive for the joint environment, since
drugs that inhibit COX-2 have been associated with a
reduction of OA-like histological changes and suppressed
chondrocyte apoptosis [
]. Indeed, it has been
demonstrated that exercise is a robust approach to preserve
healthy cartilage [
]. Specifically, low intensity treadmill
walking for 2, 5, or 15 days has been demonstrated to
regulate metabolic responses at the cellular and systemic
level, protecting cartilage against OA by altering gene
expression of markers involved in OA onset [
findings indicate that all levels of exercise used in our
study led to a reduction in COX-2 mRNA in the
synovium, suggesting a potential protective effect of exercise
for the knee, even in the lowest end highest exercise
programs. It is important to highlight that at the end of
the 12-week treadmill training, many molecular changes
may have occurred to adapt to the new loading
conditions and had already plateaued and normalized to the
new exercise threshold by the time tissue was taken.
Additionally, we did not study the protein and activity
levels; this needs to be done in future studies.
Synovial fluid EGF levels were significantly higher in
the EHD and HD group animals than those in the
control group animals. It has been suggested that EGF is
produced by the synovium in the initial stages of
rheumatoid arthritis [
] and that activation of EGF
receptor signaling may be a causal factor in OA [
However, more recent studies indicated anabolic effects
of EGF receptor signal activation in articular cartilage,
suggesting that EGF may promote the expansion and/or
activity of an endogenous EGF receptor responsive cell
population within the articular cartilage . Moreover,
the EGF receptor is an important signaling molecule in
bone development and remodeling and plays an anabolic
role in bone metabolism [
]. Overall, the finding of
elevated synovial fluid EGF levels in the EHD and HD
group animals in the absence of OA-like changes
suggests that EGF was not detrimental to knee joint health.
Serum IL-12 levels were significantly higher for the
EHD group compared to those for the control group
and HD group rats. This result agrees with the
findings from in vitro studies. Exercise has been reported
to increase IL-12 production by macrophages
following lipopolysaccharide stimulation [
]. IL-12 is
believed to be essential for the clearance of bacterial
] and is thought to be an important
pathogenetic factor in Th1-type-mediated autoimmune disease.
IL-12-deficient mice have been found to be resistant to
collagen-induced inflammatory arthritis [
transgenic over-expression of IL-12 exacerbates the course
of this disease [
]. However, the role of IL-12 in
maintaining knee joint integrity, and its role in the
pathogenesis of osteoarthritis, is poorly understood. It
should be noted that the over-expression of serum
IL-12 may represent responses to exercise unrelated
to the knee joint, as serum may represent input from
a number of sources, i.e. muscle and vasculature.
However, in combination with the other findings in
our study, IL-12 may be playing a protective role for
knee joint cartilage. Further mechanistic studies are
required to elucidate the role of IL-12 in this model
of exercise-induced changes in knee joint health, as
well as follow the EHD rats for longer period of time
than the 12 weeks of this protocol.
In summary, a stepwise increase in the speed and
duration of a 12-week chronic treadmill exercise program
did not lead to OA-like changes in the rat knee but
appeared to produce a potential protective effect through a
reduction in COX-2 mRNA levels in the synovium.
Further studies aimed at elucidating the preventive and
potentially harmful effects of repetitive chronic exercise
need to be performed to better understand the effects of
joint loading on joint health above or below the
physiological window. Working within an optimal physiological
exercise window is beneficial for general and for joint
health across the life span. It is also important in the
context of recreational and elite sports, where the
optimal window may be altered following joint injury or
disease and may thus affect the return to sport.
BC: Before Christ; Col-1: Type I collagen; Col-3: Type II collagen;
COX2: Cyclooxygenase-2; DXA: Dual X-ray absorptiometry; EGF: Epidermal growth
factor; EHD: Extra high duration; G-CSF: Granulocyte colony-stimulating factor;
GM-CSF: Granulocyte macrophage colony-stimulating factor; GRO/KC: CXCL1
chemokine, growth-related oncogene keratinocyte chemoattractant; HD: High
duration; IFN-γ: Interferon gamma; IGF-1: Insulin-like growth factor 1;
IL: Interleukin; iNOS: Inducible nitric oxide synthase; IP-10/CXCL10: Interferon
gamma-induced protein 10; LIX: CXCL5 chemokine; MCP-1: Macrophage
chemoattractant protein-1; MD: Moderate duration; MIP: Macrophage
inflammatory protein; mRNA: Messenger ribonucleic acid; OA: Osteoarthritis;
OARSI: Osteoarthritis Society International (Histology Scoring System);
PPARγ: Peroxisome proliferator-activated receptor gamma; RANTES: Regulated on
activation, normal T cell expressed and secreted; RNA: Ribonucleic acid;
RTqPCR: Real-time quantitative polymerase chain reaction; TF: Tissue factor;
TFPI: Tissue factor pathway inhibitor; TGF-β: Transforming growth factor-beta;
Th1: Type 1 T helper cells; TNF-α: Tumor necrosis factor alpha; VEGF: Vascular
endothelial growth factor
The authors thank Dr. Tannin Schmidt for providing access to the Mach-1
v500css for mechanical testing and Carolyn Hewitt for technical contributions
to this paper.
This work was supported by the Canadian Institutes of Health Research #
RT736475 and MOP 115076, the Canada Research Chair Programme, the
Alberta Innovates Health Solutions Osteoarthritis Team Grant, Alberta
Innovates Health Solutions, Killam Foundation, and the Ministry of Education,
Brazil (CAPES Foundation Grant 13157-13-2).
Availability of Data and Materials
The datasets supporting the conclusions of this article are included within
JLR was responsible for the design of the study, execution of the study, data
collection, data analysis, interpretation of data, drafting the manuscript,
revising the manuscript, and approving the final submitted version. KRB was
responsible for the design of the study, execution of the study, data
collection, interpretation of data, revising the manuscript, and approving the
final submitted version. JWM was responsible for the data collection, data
analysis, interpretation of data, and approving the final submission. RAS was
responsible for the data analysis, interpretation of data, and approving the
final submission. DAH was responsible for the data analysis, interpretation of
data, drafting the manuscript, revising the manuscript, and approving the
final submission. WH contributed to the study design, interpretation of the
data, writing the manuscript, revising the manuscript, and approving the
All experiments were approved by the University of Calgary Life and
Environmental Sciences Animal Care Committee (AC12-0139), and all
methods were conducted in accordance with the animal welfare regulations
and guidelines at the University of Calgary.
Consent for Publication
Jaqueline Lourdes Rios, Kevin Rudi Boldt, James William Mather, Ruth Anne
Seerattan, David Arthur Hart, and Walter Herzog declare that they have no
conflict of interest.
Springer Nature remains neutral with regard to jurisdictional claims in
published maps and institutional affiliations.
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