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Piperine enhances carbohydrate/fat metabolism in skeletal muscle during acute exercise in mice

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Background: Exercise promotes energy metabolism (e.g., metabolism of glucose and lipids) in skeletal muscles; however, reactive oxygen species are also generated during exercise. Various spices have been reported to have beneficial effects in sports medicine. Here, we investigated the effects of piperine, an active compound in black pepper, to determine its effects on metabolism during acute endurance exercise. Methods: ICR mice (n = 18) were divided into three groups: nonexercise (CON), exercise (EX), and exercise with piperine (5 mg/kg) treatment (EP). Mice were subjected to enforced exercise on a treadmill at a speed of 22 m/min for 1 h. To evaluate the inflammatory responses following exercise, fluorescence-activated cell sorting analysis was performed to monitor changes in CD4(+) cells within the peripheral blood mononuclear cells (PBMCs) of mice. The expression levels of metabolic pathway components and redox-related factors were evaluated in the soleus muscle by reverse transcription polymerase chain reaction and western blotting. Results: There were no changes in the differentiation of immune cells in PBMCs in both the EX and EP groups compared with that in the CON group. Mice in the EX group exhibited a significant increase in the expression of metabolic pathway components and redox signal-related components compared with mice in the CON group. Moreover, mice in the EP group showed greater metabolic (GLUT4, MCT1, FAT/CD36, CPT1, CS) changes than mice in the EX group, and changes in the expression of redox signal components were lower in the EP group than those in the EX group. Conclusion: Our findings demonstrate that piperine promoted beneficial metabolism during exercise by regulating carbohydrate/fat metabolism and redox signals. Therefore, piperine may be a candidate supplement for improvement of exercise ability.
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R E S E A R C H Open Access
Piperine enhances carbohydrate/fat
metabolism in skeletal muscle during
acute exercise in mice
Jisu Kim
1
, Kang-Pa Lee
2
, Dae-Won Lee
3
and Kiwon Lim
1,4*
Abstract
Background: Exercise promotes energy metabolism (e.g., metabolism of glucose and lipids) in skeletal muscles;
however, reactive oxygen species are also generated during exercise. Various spices have been reported to have
beneficial effects in sports medicine. Here, we investigated the effects of piperine, an active compound in black
pepper, to determine its effects on metabolism during acute endurance exercise.
Methods: ICR mice (n= 18) were divided into three groups: nonexercise (CON), exercise (EX), and exercise with
piperine (5 mg/kg) treatment (EP). Mice were subjected to enforced exercise on a treadmill at a speed of 22 m/min
for 1 h. To evaluate the inflammatory responses following exercise, fluorescence-activated cell sorting analysis was
performed to monitor changes in CD4
+
cells within the peripheral blood mononuclear cells (PBMCs) of mice. The
expression levels of metabolic pathway components and redox-related factors were evaluated in the soleus muscle
by reverse transcription polymerase chain reaction and western blotting.
Results: There were no changes in the differentiation of immune cells in PBMCs in both the EX and EP groups
compared with that in the CON group. Mice in the EX group exhibited a significant increase in the expression of
metabolic pathway components and redox signal-related components compared with mice in the CON group.
Moreover, mice in the EP group showed greater metabolic (GLUT4, MCT1, FAT/CD36, CPT1, CS) changes than mice in
the EX group, and changes in the expression of redox signal components were lower in the EP group than those
in the EX group.
Conclusion: Our findings demonstrate that piperine promoted beneficial metabolism during exercise by regulating
carbohydrate/fat metabolism and redox signals. Therefore, piperine may be a candidate supplement for improvement
of exercise ability.
Keywords: Acute endurance exercise, Piperine, Carbohydrate metabolism, Fat metabolism, Antioxidant
Background
The western diet generally includes excessive caloric in-
take. Moreover, many individuals have adopted a seden-
tary lifestyle, including little or irregular physical activity,
leading to increased rates of obesity and mortality [1].
The World Health Organization (WHO) recommends
that individuals exercise daily to alleviate obesity and
improve health [2]. Therefore, appropriate research is
needed to fully elucidate the effects of exercise on health
and the appropriate type of exercise that should be
adopted to ensure a healthy lifestyle.
Endurance exercise demands consumption of energy
through increased metabolism, leading to reduction of
body weight [3, 4]. Moreover, endurance exercise involves
generation of reactive oxygen species (ROS) during energy
synthesis. Excessive ROS generation from high-intensity
endurance exercise can lead to muscle rupture and im-
mune system over-reaction [5]. Additionally, continuous
exercise increases immune system function by activating
immune cells, including T cells, natural killer (NK) cells,
* Correspondence: exercise@konkuk.ac.kr
1
Physical Activity & Performance Institute, Konkuk University, 120
Neungdong-ro, Gwangjin-gu, Seoul 05029, Republic of Korea
4
Department of Physical Education, Laboratory of Exercise Nutrition, Korea
University, 120 Neungdong-ro, Gwangjin-gu, Seoul 143-701, Republic of
Korea
Full list of author information is available at the end of the article
© The Author(s). 2017 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0
International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and
reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to
the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver
(http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
Kim et al. Nutrition & Metabolism (2017) 14:43
DOI 10.1186/s12986-017-0194-2
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
and T helper (Th) cells [6, 7]. Despite these findings, add-
itional work is still needed to determine the effects of im-
mune system activation and exercise on the generation of
ROS. Low levels of ROS augment metabolism, thereby en-
hancing cell proliferation and differentiation, whereas the
presence of high levels of ROS induces cell death [8, 9].
Enzymes within the redox signaling cascade function to
maintain homeostasis despite ROS generation. For ex-
ample, nicotinamide adenine dinucleotide phosphate
(NADPH)-oxidase (NOX), which is encoded by NOX
family genes [10], is a key enzyme involved in ROS gener-
ation. NOX1 is expressed in a variety of cells and pro-
motes ROS generation [10, 11]. Oxidative stress from the
generated ROS is eliminated by antioxidant signaling and
enzymes involved in redox homeostasis, such as manga-
nese superoxide dismutase (Mn-SOD), catalase, zinc
superoxide dismutase (Zn-SOD), and Ape/Ref-1 [12, 13].
In particular, Ape/Ref-1 is a multifunctional protein with
endonuclease, transcription factor, and antioxidant func-
tions. However, the effects of Ape/Ref-1 on the mainten-
ance of redox homeostasis and immune system function
in the context of excessive exercise have not yet been
elucidated.
Skeletal muscles have important metabolic energy
functions based on the generation and utilization of en-
ergy sources from the phosphagen system, glycolysis,
and the anaerobic system. Carbohydrate and fat metab-
olism occur in the muscles during exercise [14]. Skeletal
muscles generate energy using energy sources source
such as glucose, lactate, and fatty acids [4, 15]. These
components are transported into muscle cells via spe-
cific metabolic transporters, including glucose trans-
porter type 4 (GLUT4), monocarboxylate transporter 1
(MCT1), and FAT/CD36 [16, 17]. The energy required
for transport is generated through the tricarboxylic acid
(TCA) cycle in the mitochondria [18, 19]. In particular,
carnitine palmitoyltransferase 1 (CPT1), which is present
in the mitochondrial outer membrane and plays an im-
portant role in fat metabolism, transports long-chain
acetyl-CoA, which is generated from fatty acids in the
inner mitochondrial membrane [20, 21]. In addition, cit-
rate synthase (CS), which converts acetyl-Co-A to citrate
before entering the TCA cycle, is also important in the
production of energy [22, 23].
In both athletes and individuals who are exercising for
the first time, strenuous exercise can cause various types
of damage in the body. Therefore, an effective exercise
routine for each person is necessary, taking into account
nutritional and physiological changes. Accordingly, re-
cent research has focused on the role of health supple-
ments in promoting the beneficial effects of exercise.
Piperine, one of the main components of pepper, has
been shown to have diverse biological activities, includ-
ing anti-inflammatory, antioxidant, anti-atherosclerotic
anti-obesity effects and blood lipid improvement in a
variety of cell types and animal models. However, the ef-
fects of piperine on endurance exercise and changes in
skeletal muscles have not yet been established.
Accordingly, in this study, we determined the effects
of piperine on muscles during endurance exercise in a
mouse model.
Methods
Animal experiment
Six-week-old male ICR mice (n= 18) were purchased
from Orient Bio Inc. (Seongnam, Korea). All mice were
housed in standard plastic cages under controlled condi-
tions of humidity (50%) and temperature (23 ± 1 °C)
with an alternating 12-h light/dark cycle. Mice were ac-
climated to the laboratory housing conditions for 7 days.
Mice were randomized into three groups: nonexercise
(CON), exercise (EX), and exercise with piperine treat-
ment (EP). Piperine was dissolved in dimethyl sulfoxide
(DMSO; via oral gavage, in a volume of 50 μL) and ad-
ministered to the EP group at 5 mg/kg; the CON group
and EX groups were treated with distilled water via oral
administration at 30 min before acute endurance exer-
cise. All experimental procedures were performed at the
Animal Experiment Research Center of Konkuk Univer-
sity. This study was conducted in accordance with the
ethical guidelines of the Konkuk University Institutional
Animal Care and Use Committee. All mice were adapted
to treadmill training (Daejong Systems, Korea) at a fixed
intensity (10 m/min, 8° slope, 10 min) for 3 days. The
following protocols were used: 22 m/min, 8° slope,
60 min (approximately 70% maximal oxygen consump-
tion [VO
2
]) for acute exercise. Blood samples and soleus
muscles were collected immediately after exercise.
Fluorescence-activated cell sorting (FACS) analysis
FACS assays were performed as previously reported [24].
To determine the expression of T-cell-related surface
molecules, peripheral blood mononuclear cells (PBMCs)
were isolated using a Ficoll-Hypaque gradient (Sigma-
Aldrich, UK). For analysis of cytokine production in dif-
ferentiated Th1, Th2, and Th17 cells, isolated PBMCs
were stained with anti-CD3-PerCP, anti-CD4-APC, and/
or anti-CD8-APC-Cy7 antibodies (BioLegend, USA).
After washing with FACS buffer (0.1% bovine serum al-
bumin [BSA] in phosphate-buffered saline [PBS]), the
cells were then stimulated with 50 ng/mL phorbol 12-
myristate 13-acetate (PMA; Sigma-Aldrich) and 1 μg/mL
ionomycin (Sigma-Aldrich) in the presence of Golgistop
(BD Biosciences, USA) for 4 h at 37 °C. The stimulated
cells were washed with FACS buffer and fixed for
10 min with 4% paraformaldehyde. After fixation, the
cells were permeabilized with FACS Perm 2 according to
the manufacturers instructions (BD Biosciences, USA)
Kim et al. Nutrition & Metabolism (2017) 14:43 Page 2 of 8
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and stained with appropriate fluorochrome-conjugated
antibodies, including anti-mouse interferon (IFN)-γconju-
gated with fluorescein isothiocyanate (FITC; BD Biosciences,
USA) and anti-mouse interleukin (IL)-17A conjugated with
phycoerythrin (eBioscience, Germany) antibodies using iso-
type antibodies as controls. All data were collected on a
FACSCalibur flow cytometer (BD Biosciences) and analyzed
using FlowJo software (Tree Star Inc., USA).
Total RNA isolation and reverse transcription (RT)-
polymerase chain reaction (PCR)
Total RNA isolation and RT-PCR analyses were per-
formed as previously reported [25]. The soleus muscles
of each mouse were harvested, and total RNA was iso-
lated from cells using TRI-reagent (GenDEPOT, Korea).
Next, cDNA was synthesized using 1.0 μg of total RNA
with oligo dT(18mer). For PCR, 1 μLofcDNAwassub-
jected to predenaturation at 95 °C for 3 min, 35 cycles of
denaturation at 94 °C for 1 min, primer annealing at the
optimal temperature for 1 min, and extension at 72 °C for
1 min, followed by a final extension at 72 °C for 10 min.
The following primers were used for PCR: GLUT4 (60 °C;
sense primer, 5-AACTTGGCATTGTGGAAGG-3,and
antisense primer, 5-ACACATTGGGGGTAGGAACA-3),
MCT1 (60 °C; sense primer, 5-GCTGGAGGTCCTAT
CAGCAG-3, and antisense primer, 5-AGTTGAAAG
CAAGCCCAAGA-3), CD36 (60 °C; sense primer, 5-
GGCCAAGCTATTGCGACAT-3,andantisenseprimer,
5-CAGATCCGAACACAGCGTAGA-3), CPT1 (60 °C;
sense primer, 5-ATCATGTATCGCCGCAAACT-3,and
antisense primer, 5-CCATCTGGTAGGAGCACATGG-
3), CS (60 °C; sense primer, 5-CAAGTCATCTACGC
CAGGGAC3, and antisense primer, 5-CAAAGCGTCTC
CAGCTAACCA-3), and GAPDH,senseprimer,5-GG
CATTGCTCCTCAATGACAA-3,andantisenseprimer,
5-TGTGAGGGAGATGCTCAGTG-3.PCRproducts
were separated by 2.0% agarose gel electrophoresis and vi-
sualized by staining with ethidium bromide.
Tissue processing and immunoblotting assay
Immunoblotting was performed as previously reported
[26]. Tissue was homogenized in ice-cold cell lysis buffer
(50 mM Tris-HCl, pH 7.4, 150 mM NaCl, 0.25% deoxy-
cholic acid, 1% NP-40, 1 mM ethylenediaminetetraacetic
acid [EDTA], and protease inhibitors, including 1 mM phe-
nylmethylsulfonyl fluoride [PMSF], 1 g/mL aprotinin, 1 g/
mL leupeptin, 1 mM Na
3
VO
4
, and 1 mM NaF), and ho-
mogenates were agitated for 1 h at 4 °C. Homogenates were
then centrifuged at 13,000×gat 4 °C for 15 min, and the
supernatant was collected and stored at 80 °C until further
analysis. To analyze protein expression, we performed
western blotting using specific antibodies. Briefly, 30 μgof
protein from each group was boiled, separated by electro-
phoresis on 12% acrylamide gels, and then transferred onto
polyvinylidene difluoride membranes in transfer buffer
at 4 °C for 2 h. The membranes were blocked with 5%
BSA in Tris-buffered saline (TBS) at room temperature
for 1 h and then washed in TBS with 0.1% Tween 20
(TBS/T). The membranes were incubated overnight at 4 °C
with specific antibodies against NOX-1, Ape/Ref-1, Mn-
SOD, and β-actin (1:1000 dilution [Santa Cruz, USA]). The
membranes were washed with TBS/T, followed by incubation
with IgG secondary antibodies conjugated with horseradish
peroxidase (1:1000 dilution [Santa Cruz, USA]). The expres-
sion levels of the proteins were analyzed using chemilumin-
escence (ECL Plus Kit; Amersham Pharmacia Biotech).
Developed protein bands were visualized and quantified
using Image J software (NIH, Bethesda, MD, USA).
Data analysis
Data were expressed as the mean ± standard error of the
means (SEMs). Statistical evaluation of the data was per-
formed using GraphPad Prism, version 5.0 (GraphPad
Software, USA). Studentst-tests and one-way analysis of
variance (ANOVA) with Tukeys post-hoc tests were
used to compare the data. Differences with P< 0.05
were considered statistically significant.
Results
Effects of piperine on acute endurance exercise-induced
immune responses
Physical exercise can enhance the activity of immune re-
sponses, including both the innate and adaptive immune
responses [27]. Therefore, we investigated the effects of
piperine on acute endurance excise by evaluating differen-
tiated Th1, Th2, and Th17 cells in PBMCs using FACS
analysis. The one-time acute endurance excise experiment
is depicted in Fig. 1b. As shown in Fig. 2, there were no
changes in IFN-γor IL-17 expression in mice in the EX or
EP groups compared with those in the control group.
Effects of piperine on carbohydrate/fat metabolism in
soleus muscles from mice after acute endurance exercise
The skeletal muscle system represents an accessible meta-
bolic energy source, providing small carbohydrates during
exercise [15]. Therefore, to determine the effects of piper-
ine on the energy source used during acute endurance ex-
ercise, we evaluated GLUT4,MCT1,FAT/CD36,CPT1,
and CS mRNA expression by RT-PCR. As shown in Fig.
3a, the expression levels of GLUT4 and MCT1 mRNAs
were higher in the EP group than in the EX and CON
groups. Moreover, as shown in Fig. 3b, FAT/CD36,CPT1,
and CS mRNAs were also significantly upregulated by ex-
ercise and piperine administration. Besides, we performed
the following additional experiments. We checked the pro-
tein (GLUT4, FAT/36 and CPT1) levels using the western
blot assay. As show in Additional file 1: Figure S1, these
results showed similar expression of genes and proteins.
Kim et al. Nutrition & Metabolism (2017) 14:43 Page 3 of 8
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Effects of piperine on antioxidant enzymes in soleus
muscles from mice after acute endurance exercise
High-intensity exercise-induced oxidative stress usually
involves elimination mechanisms and homeostasis sys-
tems [28]. Therefore, to examine whether piperine inhib-
ited ROS generation, we measured the expression levels of
antioxidant enzymes by immunoblotting. As shown in
Fig. 4, NOX-1, Ape/Ref-1, and Mn-SOD levels were
significantly increased in soleus muscles following exercise
as compared with those in the control group. However,
piperine administration significantly reduced the expres-
sion levels of NOX-1, Ape/Ref-1, and Mn-SOD compared
with those in the control group. In addition, we checked
the results of L6 cell in vitro as follows. To explore the
effect of piperine on H2O2-stimulated L6 cell, we per-
formed the three experiments such as cell observed assay
and reverse transcription polymerase chain reaction (RT-
PCR) analysis. Cell morphology change was observed by
inverted microscope for 1 h, respectively. RT-PCR analysis
was used to determine CAT, NOX-1, APE/REF1 and Mn-
SOD. As show in Additional file 2: Figure S2A, the cell
morphology for 1 h is not altered in all conditions (un-
treated, 100 μMH2O2,and100μMH2O2withpeprine
(10 uM). H2O2 significantly increases the expression
levels of mRNA CAT, NOX-1, APE/REF1 and Mn-SOD
whereas piperine significantly regulated those-signals.
Fig. 2 Effects of piperine on the immune response in peripheral blood mononuclear cells (PBMCs) following acute exercise. a,bImmune cells
were prepared from PBMCs. Cells were stimulated with PMA, ionomycin, and golgistop for 4 h and then stained with anti-IFN-γand anti-IL-17
antibodies for 1 h. Flow cytometry analysis was used to gate IFN-γ
+
T cells (T helper type 1; Th1) and IL-17
+
T cells (Th17). The graphs show the
frequencies of Th1 and Th17 cells
Fig. 1 Study design. aStructure of piperine (C
17
H
19
NO
3
; molecular weight: 285.34 Da). bStudy design showing the exercise protocol. Mice were
administered 5 mg/kg piperine orally 30 min prior to running on a treadmill for 1 h. After exercise, mice were sacrificed, and tissues were collected
Kim et al. Nutrition & Metabolism (2017) 14:43 Page 4 of 8
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Discussion
Although piperine has been shown to have anti-
inflammatory, anticancer, anti-atherosclerotic, and anti-
oxidant effects and to inhibit lipid synthesis, the effects
of piperine on carbohydrate/fat metabolism and skeletal
muscle damage during endurance exercise have not been
evaluated. Here, we provide evidence of the role of pip-
erine in mediating carbohydrate/fat metabolism in skel-
etal muscle in acute endurance exercise for the first
time. Our findings demonstrate that piperine treatment
enhanced fat/carbohydrate metabolism in skeletal
muscle and that piperine administration decreased the
expression of NOX1 and antioxidant enzymes, such as
Mn-SOD and Ape/Ref-1, after acute exercise. Moreover,
acute endurance exercise did not alter immune re-
sponses, with or without piperine treatment. These re-
sults imply that piperine regulated muscle damage and
energy metabolism during acute endurance exercise.
Regular exercise can have beneficial effects on health;
however, high-intensity physical exercise or excessive exer-
cise can induce muscle degeneration and inflammation
[28, 29]. Recently, sports science and nutrition experts
have suggested that short-term supplementation with
antioxidants may provide beneficial effects on health
and reduce damage and inflammation after exercise
[30]. Based on our data, there were no changes in
Fig. 3 Effects of piperine on the expression of glucose/fat metabolism-associated mRNAs in the soleus muscle after acute exercise. aTotal RNA
from soleus muscles was isolated, and the expression of glucose metabolism-related genes was evaluated using RT-PCR. The graphs were obtained
from the left bands and show GLUT4 and MCT1 mRNA expression (%). Data are expressed as the mean ± SEM (n= 6). mRNA expression in the exercise
(EX) group was set at 100%. *P< 0.05 versus the EX group. bThe expression of fat metabolism-related genes was analyzed by RT-PCR. The graph
shows FAT/CD36,CPT1,andCS mRNA expression (%). Data are expressed as the mean ± SEM (n= 6). mRNA expression in the exercise (EX) group was
set at 100%. *P< 0.05 versus the EX group
Fig. 4 Effects of piperine on the expression of reactive oxygen species (ROS)-regulated mRNA in the soleus muscle after acute exercise. aSoleus
muscles were collected after exercise, and protein expression was evaluated using immunoblotting. bThe graphs represent the intensities of
NOX1, APE/Ref-1, and Mn-SOD bands (%). Data are expressed as the mean ± SEM (n= 6). The band intensity in the exercise (EX) group was set at
100%. *P< 0.05 versus the EX group
Kim et al. Nutrition & Metabolism (2017) 14:43 Page 5 of 8
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immune responses in PBMCs among the three groups
(control, exercise, and exercise after piperine adminis-
tration). These results imply that the performance in
the EP group was not related to the innate immune
response during acute exercise after short-term ad-
ministration of piperine. Therefore, we suggest that
piperinemaybeadministeredsafelyduringexercise.
Exercise is tightly associated with the generation of ROS
in the skeletal muscle [30]. During exercise, ROS are pro-
duced through energy-generating metabolic processes,
which consume high levels of oxygen. We found that
NOX1, Mn-SOD, and Ape/ref.-1 expression levels were
decreased in mice administered piperine and subjected to
acute exercise compared with those in exercised mice
without piperine administration. These results imply that
piperine provided scavenging activity against superoxide
generation, consistent with a previous study [31].
Because of the increased metabolic rate and energy de-
mand associated with exercise, the oxidization of both
fat and carbohydrates must be activated simultaneously
[4]. Previous studies have reported that FAT/CD36,
CPT1, and CS are key components of the molecular ma-
chinery required for regulating fat oxidation in skeletal
muscle [32, 33]. During exercise, increased glycolysis
and subsequent production and accumulation of lactate
necessitate increased expression of GLUT4 and MCT1
[16]. Our data also indicate that the expression levels of
fat metabolism-related genes, such as FAT/CD36,CPT1,
and CS, were significantly increased in the skeletal
muscle following exercise with or without short-term
piperine administration. A previous study reported that
piperine was supplemented in different doses (20, 30
and 40 mg/kg) through administration of a high-fat diet
(HFD) for 42 days to experimental rats. Piperine signifi-
cantly reduced the concentration of plasma and liver
lipids in obese rats to near normal levels, and HDL was
elevated [34]. Thus, piperine ingestion may influence fat
metabolism.
Moreover, our data indicate that the transcription of
GLUT4 and MCT1 was promoted by acute exercise after
short-time administration of piperine. Therefore, we
suggest that piperine, an active component of black pep-
per, may be used as a sports supplement for improving
exercise ability and adaption.
However, a limitation of this study is that we did not
use the mice only treated with piperine as a control
group. This is because we 1) intended to differentiate
our study from previous studies and 2) wanted to con-
firm the effects of supplementation of piperine in acute
exercise.
AMP-activated protein kinase (AMPK) is an enzyme
that controls key players of metabolic pathways, such as
glycolysis and fatty acid oxidation [35, 36]. Although we
did not evaluate AMPK signaling in this study, piperine
may regulate the AMPK pathway, as shown in previous
reports [37] and supported by our current data. Previ-
ously, several in vitro and in vivo studies attempted to de-
termine the mechanisms through which piperine affects
fat metabolism [3841]. Overall, our results show that
piperine treatment can improve the carbohydrate/fat me-
tabolism in skeletal muscle during acute exercise. Further
studies are needed to clarify the long-term effects of piper-
ine on exercise endurance capacity in athletes.
Conclusion
In conclusion, our findings suggest that piperine improve
beneficial energy metabolism during exercise by regulating
carbohydrate/fat metabolism without stimulating the innate
immune response or superoxide generation. Therefore, pip-
erine may have applications as a nutritional supplement for
improvement of exercise ability.
Additional files
Additional file 1: Figure S1. Effects of piperine on the expression of
glucose/fat metabolism-associated protein in the soleus muscle after
acute exercise. Total protein from soleus muscles was isolated, and the
expression of glucose metabolism-related genes was evaluated using
western blot. (JPEG 40 kb)
Additional file 2: Figure S2. Effects of piperine on exogenous hydrogen
peroxide (H2O2)-stimulated L6 skeletal muscle cells. L6 cells were treated
with piperine (10 μM) and presence or absence of H2O2 (100 μM) for 1
h. These morphological changes were observed by inverted microscope
(A). Reverse transcription polymerase chain reaction analysis was used to
determine CAT, NOX-1, APE/REF1 and Mn-SOD (B). These graphs are pre-
sented as mean ± standard error (P< 0.05). (JPEG 230 kb)
Abbreviations
AMPK: AMP-activated protein kinase; ANOVA: Analysis of variance;
CPT1: Carnitine palmitoyltransferase 1; CS: Citrate synthase; FITC: Fluorescein
isothiocynanate; GLUT4: Glucose transporter type 4; IFN: Interferon;
IL: Interleukin; MCT1: Monocarboxylate transporter 1; Mn-SOD: Manganese
superoxide dismutase; NADPH: Nicotinamide adenine dinucleotide
phosphate; NK: Natural killer; NOX: Nicotinamide adenine dinucleotide
phosphate; PBMC: Peripheral blood mononuclear cell; PCR: Polymerase chain
reaction; PMA: Phorbol 12-myristate 13-acetate; ROS: Reactive oxygen
species; RT: Reverse transcription; SEM: Standard error of the mean; TBS: Tris-
buffered saline; TBS/T: Tween 20; TCA: Tricarboxylic acid; Th: T helper; Zn-
SOD: Zinc superoxide dismutase
Acknowledgments
This paper was supported by the KU Research Professor Program of Konkuk
University.
Funding
This study was supported by the Basic Science Research Program through
the National Research Foundation of Korea (NRF) funded by the Ministry of
Education (2015R1D1A4A01019776).
Availability of data and materials
Data are all contained within the article.
Authorscontributions
JSK contributed to the study conception and experimental design, collected
data, and performed analyses. KPL interpreted the data and had primary
responsibility for the final content. DWL participated in the study conception.
KWL provided advice on the study design and management. All authors
Kim et al. Nutrition & Metabolism (2017) 14:43 Page 6 of 8
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were involved in editing the manuscript and read and approved the final
manuscript.
Competing interests
The authors declare that they have no competing interests.
Consent for publication
Not applicable.
Ethics approval
All experimental procedures were performed at the Animal Experiment
Research Center of Konkuk University. This study was conducted in
accordance with the ethical guidelines of the Konkuk University Institutional
Animal Care and Use Committee.
PublishersNote
Springer Nature remains neutral with regard to jurisdictional claims in published
maps and institutional affiliations.
Author details
1
Physical Activity & Performance Institute, Konkuk University, 120
Neungdong-ro, Gwangjin-gu, Seoul 05029, Republic of Korea.
2
Department
of Medical Science, School of Medicine Konkuk University, 120
Neungdong-ro, Gwangjin-gu, Seoul 05029, Republic of Korea.
3
Department
of Bio-Science, College of Natural Science, Dongguk University, Dongdae-ro
123, Gyeongju, Gyeongsangbuk-do 38066, Republic of Korea.
4
Department of
Physical Education, Laboratory of Exercise Nutrition, Korea University, 120
Neungdong-ro, Gwangjin-gu, Seoul 143-701, Republic of Korea.
Received: 22 December 2016 Accepted: 7 June 2017
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Supplementary resources (2)

... The mRNA levels were analyzed as previously reported [25]. The gastrocnemius muscles of untreated and starved rats were harvested, and total RNA was separated from L6 skeletal muscle cells using TRIzol, according to the manufacturer's instructions. ...
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Imaging techniques for diagnosing muscle atrophy and sarcopenia remain insufficient, although various advanced diagnostic methods have been established. We explored the feasibility of 18F-fluorocholine (18F-FCH) positron emission tomography/computed tomography (PET/CT) for evaluating skeletal muscle atrophy, as an imaging technique that tracks choline level changes in muscles. Cell uptake in L6 cells by 18F-FCH was performed in a complete medium containing serum (untreated group, UN) and a serum-free medium (starved group, ST). Small-animal-dedicated PET/CT imaging with 18F-FCH was examined in in-vivo models with rats that were starved for 2 days to cause muscle atrophy. After the hind limbs were dissected, starvation-induced in-vivo models were anatomically confirmed by reverse-transcription polymerase chain reaction to evaluate the expression levels of the atrophy markers muscle RING-finger protein-1 (MuRF-1) and atrogin-1. 18F-FCH uptake was lower in the starvation-induced cells than in the untreated group, and in-vivo PET uptake also revealed a similar tendency (the average standardized uptake value (SUVmean) = 0.26 ± 0.06 versus 0.37 ± 0.07, respectively). Furthermore, the expression levels of MuRF-1 and atrogin-1 mRNA were significantly increased in the starvation-induced muscle atrophy of rats compared to the untreated group. 18F-FCH PET/CT may be a promising tool for diagnosing skeletal muscle atrophy.
... Consistent with our results, Lee et al. discovered that the inhibition property of piperine in the migration of platelet-derived growth factor (PDGF)-BB-stimulated vascular smooth muscle cells through ROS scavenging [51]. Interestingly, piperine administration significantly suppressed NOX1 expression in the soleus muscles of mice after acute endurance exercise; similarly, piperine (10 µM) significantly blocked NOX1 expression in H 2 O 2 (100 µM)-stimulated L6 cells [52]. In this study, we observed the upstream molecules of ROS as LCA-stimulated Src/EGFR signals in human CRC cells. ...
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Piperine, a natural alkaloidal pungent product present in pepper plants, possesses the properties of anti-inflammatory and anti-metastasis. Lithocholic acid is a monohydroxy-5beta-cholanic acid with an alpha-hydroxy substituent at position 3; it is a secondary bile acid that plays a pivotal role in fat absorption, and has been discovered to mediate colorectal cancer (CRC) cell invasion and migration. However, the effect of piperine on angiogenesis has been poorly investigated. In the current study, we examined the role of piperine on LCA-stimulated angiogenesis by measuring interleukin-8 (IL-8) expression; moreover, we revealed the potential molecular mechanisms in CRC cells. Here, we showed that piperine inhibited LCA-stimulated endothelial EA. hy926 cell angiogenesis in a conditioned medium obtained from colorectal HCT-116 cells. Experiments with an IL-8 neutralizer showed that IL-8 present in the conditioned medium was the major angiogenic factor. Piperine inhibited LCA-stimulated ERK1/2 and AKT via the Src/EGFR-driven ROS signaling pathway in the colorectal cell line (HCT-116). Through mutagenesis and inhibitory studies, we revealed that ERK1/2 acted as an upstream signaling molecule in AP-1 activation, and AKT acted as an upstream signaling molecule in NF-κB activation, which in turn attenuated IL-8 expression. Taken together, we demonstrated that piperine blocked LCA-stimulated IL-8 expression by suppressing Src and EGFR in human CRC HCT-116 cells, thus remarkably attenuating endothelial EA. hy926 cell tube formation.
... while suppressing the cytokines such as interleukin 4, interleukin 5, and interleukin 13 (Kim, Lee, Lee, & Lim, 2017). Thus, piperine acts as a retarder for immunomodulatory Th-2 cytokines. ...
Chapter
Black pepper is known for its pungent constituent piperine. It has wide culinary uses and also possess certain preservative and medicinal properties. Piperine, beyond pungency, has been found to possess bio-transformative effects that can detoxify pathogen, prevent oxidation, and improve the bioavailability and absorption of various drugs. Piperine also shows immunomodulatory, antiasthmatic, anticarcinogenic, antiulcer, antiamoebic, antioxidant, antiinflammatory, and antiulcer properties. On the other hand, curcumin, the curcuminoid of turmeric, mainly found in rhizomes of Curcuma spp., helped to prevent skin cancers, duodenum, forestomach, and colon cancers in mice. In India and many other regions of the world, piperine and curcumin have been part of the folk formulations used for treating various ailments for centuries. In this chapter, the recent developments regarding chemistry, extraction processes, pharmacological, and biological activities of both piperine and curcumin have been discussed.
... Piperine has been found to be useful to treat rhinitis, an allergyinduced by ovalbumin (Aswar et al., 2015). Piperine helps in inhibiting the airway inflammation during bronchial asthma by improving the expressions of eosinophil CC chemokine receptor (CCR3) and transforming growth factor (TGF-β) while suppressing the cytokines such as interleukin 4, interleukin 5, and interleukin 13 (Kim, Lee, Lee, & Lim, 2017). Thus, piperine acts as a retarder for immunomodulatory Th-2 cytokines. ...
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A Centum of Valuable Plant Bioactives
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Skeletal muscle (SkM) is essential for body movement, energy metabolism, and material metabolism, and directly influences the quality of human life. Aging, chronic diseases, and strenuous exercise often lead to various health problems associated with SkM, including muscle atrophy, loss of muscle mass and strength, and metabolic disorders. Various natural products(NaPs), mainly resveratrol (RES), quercetin (QUE), ursolic acid (UA), ecdysone (ECD; mainly 20-OH ECD, 20-HE), and vitamin D, have been reported to protect or regulate SkM health. Some of the products are functionally equivalent to sex hormones, and some are even referred to as "plant exercise pills." However, controversy persists regarding the role of NaPs in SkM health. Therefore, this review objectively summarizes the in vivo and vitro biological activities, molecular mechanisms, and clinical research results of studies on NaPs applied in the regulation of SkM health over the past decade. The present review could advance further research on NaPs and SkM health, and facilitate the revelation of new evidence that could facilitate the application of NaPs in ensuring SkM health.
... It attenuated microcystin-induced oxidative damage by increasing hepatic levels of GSH, SOD, CAT, and GSH-Px contents in mice [201]. Piperine also improved glucose and lipid metabolism in skeletal muscles during exercise in mice [202]. Most importantly, piperine treatment could reverse high fat diet-induced hepatic steatosis and insulin resistance in part by increasing serum levels of adiponectin and the hepatic expression of AdipoR1 and AdipoR2 in a dose-dependent manner in mice [203]. ...
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... The respiratory gas analysis was conducted using an open circuit respiratory system, as in previous studies [20][21][22]. Rat O 2 uptake and CO 2 production were measured using a mass analyzer (model RL-600, Alco Systems, Chiba, Japan) and a switching system (model AN16-A-S, Alco Systems, Chiba, Japan), using eight acrylic metabolic chambers (23 × 10 × 13 cm 3 ; chamber volume of approximately 3 L). The flow rate within the chamber was 3 L/min. ...
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Fatigue is a comprehensive response to a variety of physiological and biochemical changes in the body. Combining strategies addressing relevant key factors simultaneously may enhance anti-fatigue efficiency. In this study, we aimed to develop new anti-fatigue agents with combining strategy based on bioinformatics analysis. Result indicated PI3k-Akt, cAMP, and p53 signaling pathway-related energy metabolism and antioxidant capacity were involved in exercise-induced physiological changes. A new representative complex agent (Z-005, containing arctigenin, caffeine, and glucose) addressing these key factors as well as central excitation was developed and tested. Z-005 showed a synergistic effect, greatly enhancing anti-fatigue efficiency than single components, in a mouse running wheel test. Mouse exhaustion distances were 2.40-fold greater than the control in the Z-005 group. Z-005 excited the central nervous system, increased the plasma superoxide dismutase and catalase concentrations, reduced lactic acid accumulation, maintained the glycogen content. Besides, the expression of key genes in PI3K, cAMP, and p53 pathways, as well as the phosphorylation of adenosine monophosphate-activated protein kinase (AMPK) were proved with enhancement by Z-005 administration. This composite formulation thus represents a new and practicable combined strategy for fatigue prevention, and the study findings can contribute to the development of treatments for fatigue-related sub-health.
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Summary Athletes make great efforts to increase their endurance capacity in many ways. Using nutrition supplements for stimulating lipolysis is one such strategy to improve endurance performance. These supplements contain certain ingredients that affect fat metabolism; furthermore, in combination with endurance training, they tend to have additive effects. A large body of scientific evidence shows that nutrition supplements increase fat metabolism; however, the usefulness of lipolytic supplements as ergogenic functional foods remains controversial. The present review will describe the effectiveness of lipolytic supplements in fat metabolism and as an ergogenic aid for increasing endurance exercise capacity. There are a number of lipolytic supplements available on the market, but this review focuses on natural ingredients such as caffeine, green tea extract, L-carnitine, Garcinia cambogia (hydroxycitric acid), capsaicin, ginseng, taurine, silk peptides and octacosanol, all of which have shown scientific evidence of enhancing fat metabolism associated with improving endurance performance. We excluded some other supplements owing to lack of data on fat metabolism or endurance capacity. Based on the data in this review, we suggest that a caffeine and green tea extract improves endurance performance and enhances fat oxidation. Regarding other supplements, the data on their practical implications needs to be gathered, especially for athletes. © 2016, Center for Academic Publications Japan. All rights reserved.
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NADPH oxidases (NOXs), also known as respiratory burst oxidase homologs (RBOHs), are the major source of reactive oxygen species (ROS), and are involved in many important processes in plants such as regulation of acclimatory signaling and programmed cell death (PCD). Increasing evidence shows that NOXs play crucial roles in plant immunity and their functions in plant immune responses are not as separate individuals but with other signal molecules such as kinases, Rac/Rop small GTPases and hormones, mediating a series of signal transmissions. In a similar way, NOX-mediated signaling also participates in abiotic stress response of plants. We summarized here the complex role and regulation mechanism of NOXs in mediating plant immune response, and the viewpoint that abiotic stress response of plants may be a kind of special plant immunity is also proposed.
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Purpose: The objective of this study was to evaluate the hepatoprotective effects of Hoveniae Semen Cum Fructus extract in ethanol induced hepatic damages. Methods: Hepatic damages were induced by oral administration of ethanol and then Hoveniae Semen Cum Fructus extract was administered. Results: Following Hoveniae Semen Cum Fructus extract administration, body and liver weights were increased, while aspartate aminotransferase, alanine aminotransferase, albumin, γ-glutamyl transferase, and triglyceride levels in the serum, triglyceride contents, tumor necrosis factor -α level, cytochrome (CY) P450 2E1 activity in the liver and mRNA expression of hepatic lipogenic genes, and Nitrotyrosine and 4-HNE-immunolabelled hepatocytes were decreased. However, mRNA expression of genes involved in fatty acid oxidation was increased. Also, as a protective mechanism for hepatic antioxidant defense systems, decreased liver MDA contents, increased glutathione contents, increased dismutase and catalase activities were observed when compared to the ethanol control. Conclusion: Hoveniae Semen Cum Fructus extract favorably protected against liver damages, mediated by its potent anti-inflammatory and anti-steatosis properties through the augmentation of the hepatic antioxidant defense system by NF-E2-related factor-2 activation, and down-regulation of the mRNA expression of hepatic lipogenic genes or up-regulation of the mRNA expression of genes involved in fatty acid oxidation.
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The phrases “ free radicals” and “reactive oxygen species” (ROS) are frequently used interchangeably although this is not always correct. This article gives a brief description of two mentioned oxygen forms. During the first two-three decades after ROS discovery in biological systems (1950-1970 years) they were considered only as damaging agents, but later their involvement in organism protection and regulation of the expression of certain genes was found. The physiological state of increased steady-state ROS level along with certain physiological effects has been called oxidative stress. This paper describes ROS homeostasis and provides several classifications of oxidative stresses. The latter are based on time-course and intensity principles. Therefore distinguishing between acute and chronic stresses on the basis of the dynamics, and the basal oxidative stress, low intensity oxidative stress, strong oxidative stress, and finally a very strong oxidative stress based on the intensity of the action of the inductor of the stress are described. Potential areas of research include the development of this field with complex classification of oxidative stresses, an accurate identification of cellular targets of ROS action, determination of intracellular spatial and temporal distribution of ROS and their effects, deciphering the molecular mechanisms responsible for cell response to ROS attacks, and their participation in the normal cellular functions, i.e. cellular homeostasis and its regulation. © 2015, Natsional’na Akademiya Nauk Ukrainy. All rights reserved.
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Metabolic alterations and cardiovascular diseases, such as atherosclerosis, are associated with lifestyle modifications, particularly the increase of physical inactivity and poor eating habits, which contribute to one of the main causes of death in modern times. Cardiovascular diseases are positively correlated with several illnesses, such as obesity, hypertension and dyslipidemia, and these disorders are known to contribute to changes in immune cells, cytokines and metabolism. Atherosclerosis is a chronic inflammatory disease characterized by the formation of lipid plaques and fibrous tissue (atheroma) in the artery walls and this process is related to the oxidation of LDL-c (low density lipoprotein) and the formation of a particle, termed LDLox, which can generate toxic injury to the vessel wall. In this atherogenic process there is an inflammatory response generated by the injury in the vascular endothelium, which in itself is able to express and secrete a variety of molecules, such as myeloid colony-stimulating factors (M-CSF), monocyte chemotactic protein-1 (MCP-1) and tumor necrosis factor alpha (TNF-α), that act as activators of the immune system. Therefore, the main purpose of this review is to highlight the immuno-metabolic alterations involving the thickening and stiffness of arteries observed in atherosclerosis, and how chronic exercise can act as an anti-inflammatory and anti-atherogenic approach.
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Purpose: Neurofibrillary tangles, one of pathological features of Alzheimer's disease, are produced by the hyperphosphorylation and aggregation of tau protein. This study aimed to investigate the effects of treadmill exercise on PI3K/AKT/mTOR signal transmission, autophagy, and cognitive ability that are involved in the hyperphosphorylation and aggregation of tau protein. Methods: Experimental animals (NSE/htau23 mice) were divided into non-transgenic control group (Non-Tg-Control; CON; n = 7), transgenic control group (Tg-CON; n = 7), and transgenic exercise group (Tg-Treadmill Exercise; TE; n = 7). The Tg-TE group was subjected to treadmill exercise for 12 weeks. After the treadmill exercise was completed, the cognitive ability was determined by conducting underwater maze tests. Western blot was conducted to determine the phosphorylation status of PI3K/AKT/mTOR proteins and autophagy-related proteins (Beclin-1, p62, LC3-B); hyperphosphorylation and aggregation of tau protein (Ser199/202, Ser404, Thr231, PHF-1); and phosphorylation of GSK-3β, which is involved in the phosphorylation of tau protein in the cerebral cortex of experimental animals. Results: In the Tg-TE group that was subjected to treadmill exercise for 12 weeks, abnormal mTOR phosphorylation of PI3K/AKT proteins was improved via increased phosphorylation and its activity was inhibited by increased GSK-3β phosphorylation compared with those in the Tg-CON group, which was used as the control group. In addition, the expression of Beclin-1 protein involved in autophagosome formation was increased in the Tg-TE group compared with that in the Tg-CON group, whereas that of p62 protein was reduced in the Tg-TE group compared with that in the Tg-CON group. Autophagy was activated owing to the increased expression of LC3-B that controls the completion of autophagosome formation. The hyperphosphorylation and aggregation (Ser199/202, Ser404, Thr231, PHF-1) of tau protein was found to be reduced in the Tg-TE group compared with that in the Tg-CON group. Furthermore, in the underwater maze test, the Tg-TE group showed a reduced escape time and distance compared with those of the Tg-CON group, suggesting that learning and cognitive ability were improved. Conclusion: These findings suggest that aerobic exercise such as treadmill exercise might be an effective approach to ameliorate the pathological features (or neurofibrillary tangles) of Alzheimer's disease.
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