Exploratory Study on Exercise Induced-
Plasmatic Follistatin Kinetics: Proof of Concept
Valeria Pérez del Toro-Cuaik1#, Jessica Saba-Behar1#, Paola Domínguez-Oropeza2, Álvaro
Antonio Martínez-Saleh2, Israel Ramírez-Sánchez3, Guillermo Ceballos-Reyes3 and Gabriela
1Facultad de Ciencias de la Salud, Universidad Anáhuac México (Campus Norte)
2Escuela de Ciencias del Deporte, Universidad Anáhuac México (Campus Norte)
3Laboratorio de Investigación Integral Cardiometabólica. Escuela Superior de Medicina, Instituto Politécnico
Gabriela Gutiérrez-Salmeán, Facultad de Ciencias de la Salud. Universidad Anáhuac México (Campus Norte),
Av. Universidad Anáhuac #46. Lomas Anáhuac, Huixquilucan, Estado de México. 52786. México, Tel:  5627 0210, Ext. 7230; E-mail:
Received: November 18, 2016; Accepted: December 06, 2016; Published:December 08, 2016
del Toro-Cuaik VP, Saba-Behar J, Domínguez-Oropeza P, Martínez-SalehÁA, Gutiérrez-Salmeán G, et al. (2016) Exploratory Study on Exercise Induced- Plasmatic Follistatin Kinetics: Proof of Concept. Obes Control Ther 3(2): 1-3. DOI: http://dx.doi.org/10.15226/2374-8354/3/2/00126
Background and objective: Follistatin is a protein that has been
positively associated with increased myogenesis and decreased fat
accumulation hence may be a potential target for preventing/treating
obesity. Its expression is induced by exercise –which may, partly,
explain the anti-obesity underlying metabolic pathways of physical
activity-; however, not many studies have described its kinetics after
performing validated exercise protocols that simultaneously evaluate
the aerobic capacity. Thus we aimed to determine the plasmatic levels
of exercise induced-follistatin and to correlate such with aerobic
Materials and methods: After performing a treadmill stress test
(Bruce protocol; with 3-minutes incremental velocity and inclination,
until fatigue or reaching 85% maximal cardiac frequency), blood
samples were drawn (at 1, 2, and 4 hours post-exercise) in order
to asses follistatin kinetics. Results were correlated with maximum
oxygen volume, calculated from protocol-validated equations.
Results: 5 moderately active women volunteers were included
(aged: 21 ± 0.7 years, weight: 54.7 ± 10.5 kg, VO2max: 57.4 ± 16.1 mL/
kg/ min). After stress test, a progressive decrease in the expression
of follistatin during the first 2 hours was observed (m = -892); levels
were found again increased at 4 hours. A negative correlation between
aerobic capacity and exercise-induced follistatin levels were found.
Conclusions: After exercise, follistatin levels exhibit a descending
slope which, at 4 hours, again rises. The most relevant finding in
this study was the negative correlation between aerobic capacity
and follistatin expression, thus allowing the assumption that their
metabolic and exercise-induced pathways may not be interrelated.
Keywords: Exercise; Follistatin; Aerobic capacity
Follistatin is an autocrine glycoprotein whose main role refers
to the antagonism and inhibition of molecules belonging to the
super family of Transforming Growth Factor Beta (TGF-β), whose members are, in turn, involved in processes of cell proliferation
and differentiation, inflammation and immunity .
Traditionally, follistatin has been studied as part of the
pituitary-gonadal axis as it occurs within the anterior pituitary
folliculostellate cells and, after secretion, follistatin has been
shown to inhibit the synthesis of Follicle Stimulating Hormone
(FSH). Complementarily, follistatin binds to –thus inhibitsactivin;
hence it stimulates gonadotropic function, not in terms
of uncontrolled cell proliferation but instead, enhances cell
The study of follistatin has gained increasing interest
because of the role of the hormone in myogenesis: it has been
shown to antagonize the function of myostatin -member of the
aforementioned TGF-β- both animal models and humans [3,4].
This phenomenon results in a significant increase in muscle mass
–as myostatin is a negative regulator of muscle growth potent
because it activates activin, hence inhibits protein transcriptionand
muscle strength  which, in turn, may be reflected in athletic
performance . Moreover, in recent years, the relationship
between muscle mass and obesity has been widely described: in
brief, muscle metabolism plays a key role in the prevention and
chronic diseases associated to excess fat body mass (i.e., obesity).
The previous are only a few reasons why skeletal muscle is no
longer seen as merely responsible for the movement, but is now
to be considered a highly active endocrine organ which, in fact,
synthesizes and releases substances -called myokines- into the
circulation after various stimuli produced by exercise and whose
action can be both para and endocrine. Within said myokines is
follistatin. Interestingly, some studies have reported that gene
expression of this protein is regulated by exercise and muscle
stretching; in fact, it has been shown that expression of follistatin
increases after the completion of a single session of exercise;
however, the specific molecular mechanisms have not yet been clarified (although it has been suggested that the expression
is rather hepatic vs. muscular-, which could be linked to the
depletion of macronutrients that occurs during exercise) nor
described the duration of such increase once the physical activity.
The objective of the present study was to therefore determine
the plasmatic levels of exercise induced-follistatin during the
recovery period and to correlate such with aerobic capacity.
Materials and methods
We conducted a proof of concept, including volunteer healthy
young adults (20 - 25 years old), with normal weight (body
mass index > 18 y < 25 kg/ m2), who performed moderately
intense physical exercise regularly and signed the corresponding
informed consent. Anthropometric assessment (weight and
height) was performed according to Lohman’s technique.
Maximum Cardiac Frequency (MCF) and its correspondent 85%
were calculated (MCF = 220 – age) in attention to the exercise
protocol, as described in the following section.
Treadmill stress test
Bruce’s protocol was used; in brief, this test progressively
increases both velocity and inclination every 3 minutes (starting
at 1.7 miles/ hour; 10% elevation) in 7 stages (reaching up to 6
miles/ hour; 22% elevation). The test finalizes whenever: a) total
ergometry time (21 minutes) ends, b) subject refers “maximum
fatigue” and “physical inability to continue the stress test” or, c)
subject reaches his/ hers 85% of MCF. Whichever the case, total
test time was recorded and further used to calculate aerobic
capacity (i.e., VO2max) with the following formula:
VO2max (mL / kg/min) = 4.38 × (time in minutes) - 3.9
In addition, Heart Rate (HR) is also recorded immediately
after finishing the stress protocol, 1 and 3 minutes afterwards.
Follistatin plasmatic levels
Before (i.e., prior to treadmill test), 1, 2, and 4 hours after
completing stress test, venous blood samples were collected
into plastic tubes containing citrate as anticoagulant. These were
immediately centrifuged (3500 r.p.m., 15 minutes) and plasma
was further separated into cryotubes. Follistatin determination
was performed using an commercial immunocolorimetric kit
The present study complies with the ethical principles
stipulated in Helsinki Declaration, the Nuremberg Code, and
Mexican regulatory laws (Ley General de Salud en Materia de
Investigación). In accordance to the latter, this research implies
a “minimum risk” as procedures considered as “routinely”
(weight measurement, stress test, blood collection < 450 mL)
are employed. Moreover, participants were informed of all
procedures prior to any intervention and, finally, they could
withdraw consent whenever they want to.
Quantitative variables are presented as mean ± standard
deviation, unless otherwise stated; qualitative variables are
presented as frequencies. For inferential analysis, repeated
measures ANOVA with Bonferroni post hoc test were performed;
correlations were evaluated according to Pearson. A statistical
significance level of p < 0.05 was considered. All analysis was
performed with GraphPad Prism®, v.5.
After an open call, 5 subjects –all females, players of the
soccer team- were recruited from a local academic institution
(Universidad Anáhuac México, Campus Norte). Demographic and
anthropometric data are show in Table 1.
Total test time was 14 ± 3.7 minutes, yielding an overall
aerobic capacity of 57.4 ± 16.0 mL / min/kg. All but one of the
participants reached their corresponding 85% of MHR (161.6
± 21.0 bpm) thus stress test was ended. 3 minutes post-test,
finishing heart rate decreased, in average, almost 50% (83.0 ±
18.9 bpm) with a referred level of fatigue of 5 ± 1.7.
Plasmatic follistatin kinetics is shown in Figure 1. No
significant differences were found among test times, although a
decreasing slope (m = -892.0 ± 259.8, r2 = 0.92) presented during
the first two hours of recovery, whereas a perfectly fit increased
gradient (m = 970.0 ± 948, r2 = 1.0) was found in the late recovery
(2-4 hours post-exercise).
Correlation analyses yielded a non significant correlation
between between aerobic capacity and percent change in heart
rate from test finishing (r2 = 0.51), as shown in Figure 2. Finally,
aerobic capacity was negatively correlated (m = -273.6 ± 66.5, r2
= 0.85, p < 0.05) with basal plasmatic follistatin (i.e., before stress
test), but shown a positive (m = 1.5 ± 0.7, r2 = 0.65, ns) correlation
Table 1 : Descriptive data
Mean ± SD
21 ± 0.7
54.7 ± 10.5
Maximum Heart Rate (bpm)
199 ± 0.7
85% MHR (bpm)
169 ± 0.7
BPM: Beats Per Minute; MHR: Maximum Heart Rate; SD: Standard
to percent change (basal vs. 4 hours post-test) regarding
follistatin blood levels (Figure 3, A and B, respectively).
Discussion And Concluding Remarks
Although not the initial aim of the present study, one of the
main findings was that even though all volunteers were part of
the University soccer team hence claimed to perform a moderatehigh
level of physical activity (i.e., 2 hours of practice, 5 days a
week), surprisingly, not one of the were able to conclude the
stress test as they all reached 85% of MCF before completing
21 minutes of the Bruce protocol. But even more, standard
deviations showed that results were greatly disperse among the
recruited participants. This observation may be attributed to the
fact that cardio-respiratory fitness and heart rate recovery have
been associated with the in-field playing position of the study
subjects , a confounding variable we did not take into account.
In this same thought, follistatin levels did not significantly
differ from one time to another. This may be due to: a) small
sample size and, b) wide variation that may be attributed to the
previously mentioned basal cardio-respiratory fitness of each
soccer player. Nevertheless, we found an ascending trend in
follistatin plasmatic levels 4 hours after finishing the stress test.
As ours, other studies have reported that the hormone’s levels
peak at 3 hours of recovery post acute exercise bouts .
Finally, even though a positive strong and significant
correlation between VO2max and exercise was certainly expected,
the negative correlation with the prior with basal follistatin was
not. This is not widely reported within the literature, although
a study reported no significant changes in blood follistatin after
resistance exercise and, moreover, authors attribute such to the
fact that levels of plasmatic myokines are enhanced by moderate
intensity exercise, rather than by resistance (i.e., high intensity)
training . This, in turn, may correlate with the types of muscle
fibers (I or II): follistatin has been associated with type II (i.e.,
aerobic or slow twitching) fibers  thus may explain the
aforementioned association with moderate (aerobic) exercise.
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