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R E S E A R C H A R T I C L E Open Access
The effect of Lactobacillus plantarum
hydrolysates promoting VEGF production
on vascular growth and hair growth of
C57BL/6 mice
Young Min Woo
2
, Ok Ju Kim
2
, Eun Sol Jo
2
, Min Young Jo
1
, Mee Young Ahn
1
, Young-Ho Lee
3
, Chun-ri Li
5
,
Sang-Hyeon Lee
1
, Jae-Suk Choi
4
, Jong Myung Ha
1
and Andre Kim
1*
Abstract
Purpose: Angiogenesis is critical in various biological processes, such as blood vessel growth, fetal differentiation,
wound healing, and organ regeneration. Various growth factors have been associated with vascular regeneration,
including insulin-like growth factor 1 (IGF-1), transforming growth factor-β(TGF-β), and basic fibroblast growth
factor (bFGF). One of the most important mediators of vascular regeneration is vascular endothelial growth factor
(VEGF). VEGF is known to increase vascular permeability, induce the proliferation of endothelial cells, and stimulate
capillary formation in vivo, which are core angiogenic functions.
Methods: The hydrolysates of lactic acid bacteria were produced by hydrolyzing Lactobacillus plantarum with
proteases, treated with MG-63 osteoblasts, and screened to obtain samples with an excellent VEGF production
effect. These samples were applied to human dermal papilla cells (hDPC) to examine the correlation between cell
growth and VEGF secretion. Furthermore, the hair growth rate was measured in hair growth experiments using
C57BL/6 male mice.
Results: The hydrolysates of the lactic acid bacteria produced in this study produced hair growth superior to the
growth obtained with 5% minoxidil in hair growth experiments using C57BL/6 male mice.
Conclusions: This study aims to develop a material for application to the scalp that promotes angiogenesis in the
scalp and facilitates the exchange of nutrients and wastes in the follicles to promote hair growth.
Keywords: Vascular endothelial growth factor (VEGF), Lactobacillus plantarum, Protease, Human dermal papilla cell
(hDPC), Hair growth
Background
Hair loss treatments applied directly to the scalp that are
currently researched include helping hair growth by nu-
trient supply and the release of harmful elements
through vasodilation or blood circulation promotion, re-
ducing stress on the hair follicle by inhibiting the gener-
ation of reactive oxygen species (ROS) or nitric oxide
(NO) that cause inflammation, reducing the risk factors
of follicles by inhibiting the formation of cytokines, and
strengthening the hair follicles by promoting angiogen-
esis (Hibino et al. 2004). Angiogenesis is a critical factor
in various biological processes such as blood vessel
growth, fetal differentiation, wound healing, and organ
regeneration. Various growth factors have been associ-
ated with vascular regeneration, including tumor necro-
sis factor (TNF), transforming growth factor-β(TGF-β),
and basic fibroblast growth factor (bFGF) (Suh et al.
2005). The formation of new blood vessels from existing
capillaries is accomplished by a combination of angio-
genic factors, extracellular matrix, and protease.
© The Author(s). 2019 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.
* Correspondence: adrk@silla.ac.kr
1
Major in Pharmaceutical Engineering, Division of Bioindustry, College of
Medical and Life Sciences, Silla University, 140 Baegyang-daero(Blvd),
700beon-gil(Rd.), Sasang-Gu, Busan 46958, South Korea
Full list of author information is available at the end of the article
Journal of Analytical Science
and Technolog
y
Woo et al. Journal of Analytical Science and Technology (2019) 10:18
https://doi.org/10.1186/s40543-019-0178-0
One of the most important mediators of vascular regen-
eration is vascular endothelial growth factor (VEGF).
VEGF is known to increase vascular permeability, induce
the proliferation of endothelial cells, and stimulate capil-
lary formation in vivo, which are core angiogenic func-
tions (Miele et al. 2000; Yeh et al. 1999). VEGF has been
reported to induce alkaline phosphatase activity and en-
hance the response to parathyroid hormone in osteoblasts
such as MG-63. Osteoblasts show the highest affinity for
VEGF receptors, suggesting that many functional VEGF
receptors are present in osteoblasts (Goad et al. 1996).
In particular, there are also reports that VEGF is se-
creted in dermal papilla cells. In light of the fact that
there are rich capillaries in the papilla during hair
growth, but that they disappear during the transition
into the resting period, it is thought that the growth of
blood vessels is associated with hair growth and thus
that VEGF is involved in hair growth. VEGF has been
also reported to reduce hair loss by promoting blood cir-
culation and the formation of new blood vessels around
follicles (Back et al. 1999).
The growth factors secreted from dermal papilla cells
include basic fibroblast growth factor, insulin-like growth
factor-1 (IGF-1), hepatocyte growth factor (HGF), fibro-
blast growth factor 1 (FGF1), and keratinocyte growth fac-
tor (KGF) (Kwon et al. 2007;Philpottetal.1994; Mitsul et
al. 1997). Transforming growth factor beta 2 (TGFβ2) has
been found to be a key factor for inducing a regression
period, as it is expressed in follicles when the hair transi-
tions from the growth period into the regression period.
In addition, TGFβ2 has been found to inhibit the prolifer-
ation of epithelial cells and shorten the human hair cycle
(Park et al. 2016; Foitzik et al. 2005).
The representative drugs that stimulate hair growth
are minoxidil and 5α-reductase inhibitors such as finas-
teride and dutasteride. These drugs have been approved
by the U.S. Food and Drug Administration (FDA). Min-
oxidil was originally developed as a vasodilator for the
treatment of hypertension. Although its mechanism of
action on hair growth has not been clearly revealed, it is
thought that the increase of nutritional supply through
vasodilation and the K
+
channel opening effect induce
hair growth (Jamora et al. 2005; Buhl et al. 1990). Fur-
thermore, minoxidil is considered to induce VEGF ex-
pression, promote the growth of cells, including
follicular keratinocyte, that are used for hair growth, and
improve blood circulation by stimulating vascular endo-
thelial cells and smooth muscles, for minoxidil is also
used for hypertension treatment (Messenger et al. 2004).
Finasteride is a hair growth agent that prevents typical
male hair loss by inhibiting the activity of 5α-reductase,
a reductase that converts testosterone to dihydrotestos-
terone (DHT), which has the effect of preventing the
death of follicle cells by DHT. However, sustained drug
use is required to maintain the hair growth-promoting
effect of finasteride, and side effects, such as sexual dys-
function in men and birth defects in pregnant women,
have been reported (Do et al. 2011; Olsen et al. 2002).
To minimize these side effects, much research has
been conducted on therapeutic agents and treatments
that use natural materials for hair growth, and they sug-
gest the possibility of developing agents that produce
excellent hair growth from natural materials (Hwang et
al. 2017; Hyung et al. 2007; Shapiro et al. 1998). Cur-
rently, clinical trials are being conducted on the hair
growth-promoting effect of compounds using the root
extracts of Rosa multiflora as a main component. Fur-
thermore, the effects on hair growth of hair tonic, food
containing medicinal herbs, algae extracts, and black
bean extracts are being researched (Kim 2014; Choi et
al. 2011; Ha et al. 2013; Jeon et al. 2011).
The Lactobacillus plantarum used in these experi-
ments is widely used as a probiotic, and these lactic acid
bacteria can be easily found not only in nature, but also
in fermented foods and animal intestines (Chang et al.
2010; Choi et al. 2015). L. plantarum is used for anti-
microbial activity, ACE (angiotensin-converting enzyme)
inhibition, lipid metabolism, and hair growth (Axling et
al. 2012; Joo 2011; Lee et al. 2012). It is also used as an
individually acknowledged raw material for health foods
that are commercialized as lactic acid bacteria products
for skin health (Choi et al. 2013; Kang et al. 2009).
Therefore, in this study, the effects on hair growth of L.
plantarum proteins hydrolyzed using various proteases
were analyzed in vitro and in vivo.
Results and discussion
Results
Cytotoxicity test after treatment of Lactobacillus plantarum
hydrolysates
The L. plantarum enzyme hydrolysates were treated with
MG-63 cells and human dermal papilla cells (hDPC) at
concentrations of 0, 31.25, 62.5, 125, 250, and 500 μg/mL.
They were cultured for 24 h and analyzed by CCK assay.
The results are shown in Fig. 1. In the MG-63 cells
(Fig. 1a), there were no significant differences between the
control group and all groups except the Lactobacillus
plantarum by Protamex (LP-P) group for concentrations
of 0 up to 250 μg/mL. In the hDPC (Fig. 1b), the cells did
not show toxicity for concentrations of 0 up to 250 μg/ml.
Thus, cell growth experiments were carried out at a con-
centration of 200 μg/mL, which is the highest concentra-
tion that does not exhibit toxicity.
Measurement of growth rate after treatment with
Lactobacillus plantarum hydrolysates
To determine the growth rates of MG-63 cells and
hDPC, cell growth was measured for 10 days. The results
Woo et al. Journal of Analytical Science and Technology (2019) 10:18 Page 2 of 9
Fig. 1 Determination of the viability of aMG-63 cell and bhDPC (24 h). The values are the means± SD of three independent experiments, and
the cell proliferation effect on the hydrolysates was represented as a percentage. The letters a, b, c, d, and e above the bars stand for significantly
different (p< 0.05) groups by one-way ANOVA, followed by Duncan’s multiple test. Cell viability was determined by CCK assay. *Values are the
means ± SD of three independent experiments (n=3)
Fig. 2 Growth of aMG-63 cells and bhDPC over 10 days. The control (con) group was not a treated sample, hydrolysates of L. plantarum (LP),
hydrolysates of L. plantarum by Alcalase (LP-A), hydrolysates of L. plantarum by bromelain (LP-B), and hydrolysates of L. plantarum by Protamex (LP-P)
Woo et al. Journal of Analytical Science and Technology (2019) 10:18 Page 3 of 9
are shown in Fig. 2. The L. plantarum enzyme hydroly-
sates were treated at a concentration of 200 μg/mL in
the medium. MG-63 cells showed steady growth until
the eighth day of growth, but the cell growth decreased
after the eighth day. The growth rate of the LP-P group
(see Fig. 1a) was lower than that of the other groups
both up until, and after, 24 h of CCK assay. As shown in
Fig. 2a, the LP, Lactobacillus plantarum by Alcalase
(LP-A), and Lactobacillus plantarum by bromelain
(LP-B) groups showed similar growth cycles to those of
the control group, whereas LP-P showed rapid growth of
106.0 × 10
5
cells/mL at day 4 after culture, 91.0 × 10
5
cells at day 6, and 93.3 × 10
5
cells/mL at day 8.
The growth of hDPC was measured for 10 days and
the results are shown in Fig. 2b. The samples showed
steady growth for 10 days. The LP-P group in particular
showed rapid growth on the eighth day. This result is
the same as that of the MG-63 cells. The LP-P group
showed the rapid growth of 5.28 × 10
5
cells/mL at day 6
of culture and 7.51 × 10
5
cells/mL at day 8.
Measurement of VEGF concentration
The amounts of VEGF and IGF-1 secreted from the
MG-63 and hDPC growth medium were measured by
ELISA, and the results are shown in Fig. 3(a, b). The
concentration of the LP-B-treated group in the MG-63
cells was 3730.44 pg/mL, which was 262.73% higher than
that of the control group. The concentrations of LP-A
and LP-P treatments were 2827.11 pg/mL and 2766.56
pg/mL, respectively, which were 199.11% and 194.84%
higher than those of the control group, respectively. The
results of VEGF measurement in the hDPC growth
medium are shown in Fig. 3b. Similarly to the MG-63
cell concentrations, the VEGF concentrations in the
LP-B group and the LP-P group were 148.58 pg/mL and
112.96 pg/mL, respectively, which were 221.60% and
168.47% higher than those of the control group (67.05
pg/mL), respectively. Madaan et al. (2017) reported that
minoxidil sulfate demonstrated stimulation of VEGF in
DPCs. The VEGF concentrations in the minoxidil (1 μM)
were about 100 pg/mL.
Fig. 3 The effect of the concentration of vascular endothelial growth factor (VEGF) and insulin-like growth factor 1(IGF-1) on L. plantarum and on L.
plantarum hydrolysates of MG-63 osteoblastic cell (a,c) and human dermal papilla cell (hDPC) (b,d). Data are expressed as a percentage of the control
result. The control (con) group was not a treated sample, hydrolysates of L. plantarum (LP), hydrolysates of L. plantarum by Alcalase (LP-A), hydrolysates
of L. plantarum by bromelain (LP-B), and hydrolysates of L. plantarum by Protamex (LP-P). All values are expressed as means ± S.D. (n= 3). The letters a,
b, c, and d above the bars stand for significantly different (p< 0.05) groups by one-way ANOVA, followed by Duncan’s multiple test
Woo et al. Journal of Analytical Science and Technology (2019) 10:18 Page 4 of 9
Measurement of IGF-1 concentration
The amount of IGF-1 secreted from the MG-63 growth
medium was measured by ELISA, and the results are
shown in Fig. 3(c, d). The concentration of the
LP-B-treated group in the MG-63 cells was 504.12 pg/
mL, which was 128.04% higher than that of the control
group. The concentrations of LP-A and LP-P treatments
were 500.71 pg/mL and 467.82 pg/mL, respectively,
which were 127.18% and 118.82% higher than those of
the control group, respectively. The results of IGF-1
measurement in the hDPC growth medium are shown
in Fig. 3d. The IGF-1 concentrations in the LP-P group
and the LP-B group were 97.82 pg/mL and 94.12 pg/mL,
respectively, which were 223.79% and 215.33% higher than
those of the control group (43.71 pg/mL), respectively.
Hair growth (appearance)
After hair removal, the appearance of hair was visually
observed and recorded (Figs. 4and 5). On the first day
of sample application, the skin color of the backs of all
groups was scarlet, and the hair began to grow little by
little from the seventh day of sample application. After
9 days, the entire skin that had been epilated was black,
as the skin hair began to grow. On the 12th day after
sample application, hair had grown in all groups, al-
though there were differences among the groups. The
LP-P group and the LP-B group had more hair than the
control group. In particular, the LP-P group and the
LP-B group showed the best hair growth.
Hair growth (dorsal tissue)
It has been shown that when hair starts growing, the skin
turns gray or black. Based on this report, it was observed
that the LP-P group and the minoxidil group, which are
the experimental groups, had completely black medial
dorsal skin color and dilated blood vessels (Fig. 6). Be-
cause the hair had almost all grown out according to the
visual observation of its appearance, it was confirmed that
the hair had partially entered the regression period. The
con, LP, and LP-A groups showed partially light gray skin,
but, on visual observation, the part where the hair did not
grow was found not to be entering the growth period, as it
was observed in the medial dorsal tissues.
Discussion
Cell growth and VEGF were measured to determine
the effect of L. plantarum hydrolysates on the prolif-
eration and activity of MG-63 osteoblast-like cells and
hDPC. After L. plantarum was hydrolyzed with Prota-
mex, MG-63 osteoblast-like cells showed a high
growth rate of 257% on the fourth day of culture,
compared to the control group. The group hydrolyzed
with bromelain showed a growth rate of 105% com-
pared to the control group. This is thought to be due
to the hydrolyzed peptide of lactic acid bacteria.
hDPC produced steady hair growth for 10 days. In
particular, the LP-P group showed rapid hair growth
onday8,andthisresultisthesameasthatof
MG-63 cells.
Fig. 4 Photograph of hair growth in C57BL/6 mice. The animals were shaved with an electric clipper and the sample applied with 0.1 mL per to
the dorsal skin of mice for 2 weeks. The photographs were taken at days 0, 4, 7, 10, and 14. Con: distilled water
Woo et al. Journal of Analytical Science and Technology (2019) 10:18 Page 5 of 9
VEGF is a growth hormone that is closely correlated
with cell growth because it is secreted during the growth
of osteoblasts such as MG-63 to induce cell migration,
survival, and proliferation. It has been also reported to
reduce hair loss by forming new blood vessels and pro-
moting blood circulation around the hair follicles
(Deckers et al. 2000). For stem cell cultures used in cos-
metics, the concentration of VEGF was found to be
about 3500–4000 pg/mL. The VEGF measured in the
MG-63 cell media was 3730 pg/mL in bromelain hydro-
lysates and 2827 pg/mL in Protamex hydrolysates, which
is higher by 262.73% and 199.11% than the figures for
the control group (1420 pg/mL), respectively.
Cell growth was higher in the LP-P group than in the
LP-B group, but the VEGF secretion rate was higher in
the LP-B group. The VEGF secreted from the hDPC
growth medium was measured as 149 pg/mL in the
LP-B group and as 113 pg/mL in the LP-P group, which
was 222% and 169% higher than the figures for the con-
trol group, respectively.
Hair growth was observed in the C57BL/6 mice. On
the seventh day, the LP-P treatment group showed a
skin color close to black, and hair growth was visually
observed on the eighth day. In the LP-P, P-con, and
LP-B groups, many blood vessels were observed to have
expanded in the median dorsal skin tissues, and their
skin color changed to black.
Materials and methods
Strain and medium used
Lactobacillus plantarum was purchased from KCCM
(KCCM 11322, Korea Federation of Culture Collection,
Seoul, Korea). MRS broth (Difco, Detroit, MI, USA) was
used as the lactic acid bacteria culture medium and was
anaerobically cultured at 37 °C for 48 h.
Production of Lactobacillus plantarum protein enzyme
hydrolysates
The protease activity of L. plantarum, and Alcalase
(Novozyme Co., Bagsvaerd, Denmark), bromelain
(Novozyme), and Protamex (Novozyme), which are dif-
ferent types of protease, were reacted with the cultured
L. plantarum by adjusting the active optimal
temperature and pH (Alcalase, 50℃, pH 7; bromelain,
45℃, pH 7; Protamax, 60℃, pH 7). They were added to
make a 1% protein concentration and hydrolyzed for 2 h
Fig. 5 Hair growth score. Hair regrowth was evaluated inspecting by the gray- or black-colored area (%) on the reverse side of the skin
Fig. 6 Hair growth on the reverse side of the skin. After the gray skin color was detected, the skin was isolated and the reverse side was
photographed to evaluate dilated blood vessels
Woo et al. Journal of Analytical Science and Technology (2019) 10:18 Page 6 of 9
(LP-A: L. plantarum hydrolysates by Alcalase; LP-B: L.
plantarum hydrolysates by bromelain; LP-P: L. plantarum
hydrolysates by Protamex). The protein concentration was
determined by drawing a standard curve using the Bradford
method with bovine serum albumin (BSA) at R
2
=0.9951 as
a standard sample. These L. plantarum protein enzyme hy-
drolysates were freeze-dried and used as samples.
Cell culture
In this study, MG-63 osteoblast-like cells were used as con-
trols for hDPC. The human osteoblast-like MG-63 cells
used in this study were purchased from Korea Cell Line
Bank (KCLB, Seoul, Korea), and the human dermal papilla
cells (hDPC) were purchased from Cell Engineering for
Origin (CEFO Co., Seoul, Korea). The cultured cells were
subcultured in our laboratory. Ten percent FBS (Gibco,
Grand Island, NY, USA) and 1% antibacterial-antifungal so-
lution were added to the DMEM (Gibco, Grand Island, NY,
USA). During the culture, 5% CO
2
was continuously sup-
plied while maintaining the temperature at 37 °C.
Cell toxicity and cell growth rate after treatment of
Lactobacillus plantarum protein enzyme hydrolysates
A CCK assay was used to verify cytotoxicity. MG-63
cells and hDPC were dispensed at a concentration of
6×10
3
cells/well on a 96-well plate. After 24 h, the L.
plantarum protein enzyme hydrolysates were added at
concentrations of 0, 31.25, 62.5, 125, 250, and 500 μg/
mL and cultured for another 24 h. After the culture, a
CCK reagent (Dongin LS, Seoul, Korea) was added, and
the hydrolysates were cultured for 2 h at 37 °C in a 5%
CO
2
incubator and measured at 450 nm using an ELISA
reader. Each treatment group underwent the experimen-
tal treatment three times, and the cell proliferation effect
on the hydrolysates was represented as a percentage.
To determine the cell growth rate, MG-63 cells were
plated on a 100-mm dish at a cell number of 1 × 10
6
cells/mL, and hDPC was plated on a 100-mm dish at a
cell number of 1 × 10
5
cells/mL. The L. plantarum pro-
tein hydrolysates were reacted with tryphan blue solu-
tion at 1:1, and the unstained live cells were counted
every other day for 10 days and observed at × 20 magni-
fication using an inverted microscope.
Measurement of growth factor
The vascular endothelial growth factor (VEGF) and
insulin-like growth factor 1(IGF-1) concentrations were
measured using the ELISA Kit (R & D Systems, Minne-
apolis, MN, USA). Fifty microliters of assay diluent was
added to 200 μL of standard and culture samples,
reacted at 37 °C for 2 h, and then washed three times.
After that, 200 μL of conjugate was added and reacted at
37 °C for 2 h again. After washing three times, 200 μLof
substrate solution was added and reacted at 37 °C for 20
min. Then, 50 μL of stop solution was added to stop the
reaction. The absorbance was measured three times, and
the absorbance value was calculated by subtracting the.
Experimental animals
Five-week-old C57BL/6 male mice were purchased from
Samtako Bio Korea (O-San, Korea). The animals were
allowed to eat food and drink water freely at a
temperature of 24 °C ± 0.5 °C, a humidity of 55–65%, and
12 h of light cycle. They were used for the experiment
after a 7-day period of adaptation.
Application of the sample
To apply the resting period during the circulation period
of the hair, the back hair was first removed using an
electric razor, and then the remaining hair was removed
using a depilatory (Niclean, Ildong Pharmaceutical).
There was no damage to the skin during hair removal,
and continuous observation showed no inflammation.
For each group, 100 μL of each extract and 5% minoxidil
(Hyundai Pharm. Co., Korea) were applied to the backs
whose hair had been removed twice a day (at 10:00 a.m.
and 6:00 p.m.) for 14 days. Distilled water was used as a
control.
Visual observation of hair growth
To verify the hair growth with the naked eye at 0, 4, 7,
10, and 14 days after the start of the experiment, Zoletil
(Virbac, Paris, France) and Rumpun (Bayel Korea Co.
Seoul, Korea) were mixed at 9:1, and the mice were
anesthetized (10/10 g) by intraperitoneal injection and
then photographed. The hair growth of each group was
rated by visual observation as 0–9% (0 points), 10–19%
(1 point), 20–29% (2 points), 30–39 (3 points), 40–49%
(4 points), 50–59% (5 points), 60–69% (6 points), 70–
79% (7 points), 80–89% (8 points), and 90–100% (9
points). On the 14th day of the experiment, the mice
were killed by cervical dislocation, the dorsal tissues
were removed, and the inside of the extracted skin was
visually observed.
Statistical processing
All experimental results are expressed as means and
standard deviations. A one-way ANOVA was conducted
for the resulting data using SPSS 20 (SPSS Inc., Chicago,
IL, USA), and the data were tested with Duncan’s mul-
tiple test at p< 0.05.
Conclusions
The L. plantarum protein hydrolyzed with Protamex
promoted hair growth more than did the 5% minoxidil.
Minoxidil, which was developed as a hypertension treat-
ment, is currently the best hair growth drug applied to
the scalp. However, if used for more than 1 month, it
Woo et al. Journal of Analytical Science and Technology (2019) 10:18 Page 7 of 9
causes side effects such as skin itching and erythema, as
well as systemic side effects, such as a decrease in blood
pressure. Thus, minoxidil is difficult to use for a long
period. However, the hydrolysates of lactic acid bacteria
proposed in this study demonstrated superior effects to
5% minoxidil without these side effects through hair
growth experiments using C57BL/6 male mice. These
results suggest that L. plantarum and its hydrolysates
can be used as basic materials for the production of cos-
metic ingredients, health foods, and medicines for
growth, osteoporosis, etc.
Abbreviations
bFGF: Basic fibroblast growth factor; IGF-1: Insulin-like growth factor 1;
LP: Lactobacillus plantarum; LP-A: Lactobacillus plantarum by Alcalase; LP-
B: Lactobacillus plantarum by bromelain; LP-P: Lactobacillus plantarum by
Protamex; TGF-β: Transforming growth factor-β; VEGF: Vascular endothelial
growth factor
Acknowledgements
This work was supported by the Technology Innovation Program (10,077,377,
Development of hair and anti-aging cosmetic materials using lactic acid bac-
teria enzyme hydrolysates) funded By the Ministry of Trade, Industry & En-
ergy (MOTIE, Korea) and supported by the Brain Busan 21+ project (BB21+).
Funding
Not applicable.
Authors’contributions
YW, OK, EJ, and MJ carried out the experiments. YW, MA, YL, and CL drafted
the manuscript. SL, JC, JH, and AK guided the research and modified the
manuscript. All authors read and approved the final manuscript.
Competing interests
The authors declare that they have no competing interests.
Publisher’sNote
Springer Nature remains neutral with regard to jurisdictional claims in
published maps and institutional affiliations.
Author details
1
Major in Pharmaceutical Engineering, Division of Bioindustry, College of
Medical and Life Sciences, Silla University, 140 Baegyang-daero(Blvd),
700beon-gil(Rd.), Sasang-Gu, Busan 46958, South Korea.
2
Department of
Natural Science Institute, Silla University, Busan 46958, South Korea.
3
Protein
Structure Group, Korea Basic Science Institute, Chungcheongbuk-do 28119,
South Korea.
4
Department of Food Engineering, Silla University, Busan 46958,
South Korea.
5
School of Acu-moxibustion and Tuina, Liaoning University of
Traditional Chinese Medicine, Shenyang 110847, China.
Received: 14 December 2018 Accepted: 5 April 2019
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