Bioscience, Biotechnology, and Biochemistry

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Propionibacterium acnes related anti-inflammation

and skin hydration activities of madecassoside,
a pentacyclic triterpene saponin from Centella
asiatica

Xueqing Shen, Miaomiao Guo, Haiyuan Yu, Dan Liu, Zhi Lu & Yanhua Lu

To cite this article: Xueqing Shen, Miaomiao Guo, Haiyuan Yu, Dan Liu, Zhi Lu & Yanhua
Lu (2018): Propionibacterium�acnes related anti-inflammation and skin hydration activities of
madecassoside, a pentacyclic triterpene saponin from Centella�asiatica, Bioscience, Biotechnology,
and Biochemistry, DOI: 10.1080/09168451.2018.1547627

To link to this article: https://doi.org/10.1080/09168451.2018.1547627

Published online: 19 Nov 2018.

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BIOSCIENCE, BIOTECHNOLOGY, AND BIOCHEMISTRY
https://doi.org/10.1080/09168451.2018.1547627

Propionibacterium acnes related anti-inflammation and skin hydration activities

of madecassoside, a pentacyclic triterpene saponin from Centella asiatica
Xueqing Shena, Miaomiao Guoa, Haiyuan Yua, Dan Liub, Zhi Lub and Yanhua Lua
a
State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, People’s Republic of China;
b
Technology Center, Shanghai Inoherb Co. Ltd, Shanghai, People’s Republic of China

ABSTRACT ARTICLE HISTORY

Madecassoside is a major pentacyclic triterpene saponin from Centella asiatica with multiple Received 14 September 2018
pharmaceutical activities. In this study, we focused on its Propionibacterium acnes related anti- Accepted 29 October 2018
inflammation and skin hydration activities, both of which play important roles in skin home- KEYWORDS
ostasis and barrier function. Madecassoside significantly inhibited the pro-inflammatory Madecassoside; skin
cytokine IL-1β, TLR2 and nuclear translocation of NF-κB in P. acnes stimulated THP-1 human hydration; hyaluronan;
monocytic cells. In addition, madecasssoside exhibited significant effects on enhancement of Propionibacterium acnes;
skin hydration through increasing the key moisturizing contributors of aquaporin-3, loricrin anti-inflammation
and involucrin in HaCaT keratinocytes as well as hyaluronan (HA) secretion in human dermal
fibroblasts. The upregulation of HA synthases (HAS1, HAS2, HAS3) and inhibition to ROS
formation accounted for the increment of HA content. Together, the in vitro study implied the
potential medical and cosmetic application of madecassoside in skin protection.

Skin functions as the first defense line from deleterious
environmental invasion and excessive water loss [1]. To
keep skin homeostasis and aesthetically attractive, skin-
care products are widely used for anti-aging, hydration,
anti-melanogenesis, etc. Among these, anti-
inflammation and hydration are considered as the most
crucial skin care step for skin overall homeostasis and
barrier function, especially for the young people.
The integrity of skin is a precondition for skin
protection. However, acne inflammation, a chronic
dermatologic disorder prevalent in teenagers and
young adults, results in inflammatory lesions of vari-
able severity and thus is the biggest threat to skin
homeostasis and attractive appearance [2,3].
Propionibacterium acnes (P. acnes) proliferation in
the pilosebaceous unit plays a critical role in eliciting
inflammatory response, in addition to two other
actors of the pathophysiology of acne (hypersebor-
rhoea and abnormal follicular keratinization) [4].
And recent studies have revealed that only specific
P. acnes strains are associated with acne [5]. The
underlying inflammatory mechanism is largely due
to the activation of Toll-like receptor (TLR)-2 on
innate immune cells to secrete pro-inflammatory
cytokines like interleukin (IL)-1β, tumor necrosis
factor (TNF)-α and interleukin (IL)-8 via induction
of signaling pathways including nuclear factor (NF)-
κB [3,6].
Besides, as extrinsic and intrinsic factors like dry
environment, surfactant-induced damages and aging
would result in skin dryness, cell dysfunction and
poor barrier function, skin hydration is regarded as
the basic skin care step for skin homeostasis [1]. Skin
hydration depends on many proteins and humectants
in epidermis and dermis. In epidermis, aquaporin-3
(AQP3), filaggrin (FLG), loricrin (LOR) and involu-
crin (IVL) are the four most studied proteins
expressed by keratinocytes and play important roles
in hydration as their moisturizing and/or barrier
enhancing activities. All these four proteins have
been demonstrated to change sensitively in HaCaT,
an immortal cell line with similar differentiation
properties as human keratinocytes [7,8]. And in der-
mis, hyaluronan (HA), mainly synthesized by dermal
fibroblasts, is considered as a key humectant in skin
hydration for its outstanding water capacity [9,10].
HA synthesis is performed by HA synthases (HAS)
[9]. While HA degradation is carried out by hyalur-
onidases (HYAL) and non-enzymatic reactions
including reactive oxygen species (ROS) [11,12].
Recently, an increasing number of plant-derived
agents have been included in formulations of skincare
products [13]. Triterpenes, constituting the majority
of the components of many traditional medicinal
plants, are among a class of compounds with multiple
biological activities on skin cells including anti-
inflammation, hydration and anti-aging [14,15].
Madecassoside (Figure 1(a)) is one of the most
important pentacyclic triterpene saponins of
Centella asiatica, whose extract is widely used in

CONTACT Yanhua Lu luyanhua@ecust.edu.cn; Zhi Lu luzhi@inoherb.com

Propionibacterium acnes related anti-inflammation and skin hydration activities of madecassoside.
© 2018 Japan Society for Bioscience, Biotechnology, and Agrochemistry
2 X. SHEN ET AL.
Figure 1. Effects of madecassoside on THP-1 cell viability and IL-1β in P. acnes stimulated THP-1 cells. (a) Chemical structure of
madecassoside. (b) Effect of madecassoside on THP-1 cell viability was monitored by MTT. (c) Production of IL-1β was assayed by
a Human IL-1β ELISA kit. (d) The mRNA expression of IL-1β was measured by qRT-PCR. Each value represents the mean ± SD of
triplicate experiments. (##) P < 0.01 compared with control group; (**) P < 0.01 compared with only P. acnes stimulated group.
medicines, foods and cosmetics. Centella asiatica
extract has been demonstrated moisturizing and anti-
inflammatory effects in vivo [16]. However, the active
components of the extract and the underlying
mechanism were poorly understood. Our study was
conducted under the hypothesis that madecassoside
was one of the contributing agents of the hydration
and anti-inflammation effect of Centella asiatica
extract. This work aimed to explore the P. acnes
related anti-inflammation and skin hydration effects
of madecassoside in vitro. Thus, the present study
might cast new light on the medical and cosmetic
application of madecassoside in skin care.
Materials and methods
Materials
Madecassoside (purity> 98%) was provided by
Shanghai Inoherb Cosmetics Co., Ltd. (Shanghai,
China). BCA Protein Assay Kit and MTT assay kit
was purchased from Biotech well (Shanghai, China).
HAS2, HAS3 and IVL antibodies were purchased from
ABclonal (Wuhan, China). HAS1 and β-actin antibo-
dies were purchased from Proteintech (Wuhan,
China). NF-κB p65 antibody was got from Cell
Signaling Technology (Boston, USA). GAPDH and
Lamin B antibodies were purchased from Biotech well
(Shanghai, China) and Boster (Wuhan, China) respec-
tively. HRP-conjugated secondary antibodies of Goat
anti-rabbit IgG and Goat anti-mouse IgG were pur-
chased from Signalway Antibody (Shanghai, China).
Cell culture and bacteria preparation
Human dermal fibroblasts (HDF) were purchased
from ScienCell Research Laboratories (Shanghai,
China). HaCaT and human monocytic cell line
THP-1 were purchased from China Center for
Type Culture Collection (Shanghai, China). HDF
and HaCaT were cultured in DMEM medium.
THP-1 cells were cultured in RPMI-1640 medium.
Medium was all added with 10% (v/v) Fetal bovine
serum (FBS) (ExCell Bio, Taicang, China) and
cells were all cultured in a carbon dioxide incu-
bator with constant temperature (5% CO2, 37°C).
P. acnes (ATCC6919, Xiangfu biotech, Shanghai,
China) was cultured under anaerobic conditions in
Cooked Meat Medium. Then it was harvested and
resuspended in FBS-free RPMI-1640 medium
before its immediate application for stimulating
THP-1 cells (P. acnes: THP-1 = 100:1) [17].
Cell viability assay
HDF or HaCaT cells were digested and adjusted to
about 6000 cells/well, and then the cell suspension
was added in 96-well plates. After 12 hours, the
medium were replaced by DMEM of various concen-
trations of madecassoside. THP-1 cells (6000 cells/
well) were similarly seeded and treated with made-
cassoside. The cell viability was quantified by MTT
assays. Briefly, cells were first treated with 0.5 mg/mL
MTT for 4 hours. Then the supernatant was carefully
removed and replaced by 150 μL DMSO to dissolve
the formazan. The absorbance at 570 nm and 630 nm
were respectively detected as detection and reference
wavelengths to determine relative cell viability.
ROS assay
HDF cells (6-well plates, 3 × 105 cells/well) were
seeded and pretreated with madecassoside for 24 h
and 200 μM H2O2 for 2 more hours [12] to assay the
level of ROS by a ROS assay kit (Beyotime, Shanghai).
And the data were confirmed by protein contents.
Enzyme linked immunosorbent assay (ELISA)
THP-1 cells seeded in 96-well plates (2 × 104 cells/well)
and 6-well plates (4 × 105 cells/well) were treated with
madecassoside for 4 h, and then stimulated by P. acnes
for 24 h. HDF cells (96-well plates, 6000 cells/well) and
 SKIN HYDRATION & ANTI-INFLAMMATION EFFECTS OF MADECASSOSIDE
HaCaT cells (6-well plates, 3 × 105 cells/well) were
treated with medium containing various concentration
of madecassoside for 24 hours. A Human IL-1β ELISA
kit (Biotech well, Shanghai, China) and a Human HA
ELISA kit (ExCell Bio, Taicang, China) were used to
analyze IL-1β and HA contents of the cell-free super-
natants collected from the 96-well plates. A TLR2
ELISA kit (Biotech well, Shanghai, China), a Human
AQP3 ELISA kit and a Human LOR ELISA kit (ExCell
Bio, Taicang, China) were respectively used to analyze
TLR2, AQP3 and LOR protein levels of the total pro-
tein extracted from the 6-well plates. And the data were
confirmed by protein contents. N-acetylglucosamine
(GlcNAc), a substrate for HA biosynthesis and demon-
strated to promote HA production in fibroblast, was
used as a positive control [18]. Retinoic acid and CaCl2
were respectively used as positive controls in AQP3
and LOR analysis [19,20].
RNA isolation and quantitative real time
polymerase chain reaction (qRT-PCR)
Total RNA of HDF, HaCaT and THP-1 cells (6-well
plates, 3 × 105 cells/well) was isolated using RNA extrac-
tion kit (KeyGEN BioTECH, Nanjing, China). Next,
cDNA was synthesized from total RNA and qRT-PCR
was performed by PrimeScript RT reagent kit and
SYBR® Premix Ex Taq™ II with gDNA Eraser (Takara,
Beijing, China) respectively under the manufacture’s
instruction. The sequences of primers used for qRT-
PCR in HDF, HaCaT and THP-1 cells were as
follows: GAPDH, forward: 5ʹ-TCC ACTGGCGTCTTC
ACC-3ʹ, reverse: 5ʹ-GGCAGAGATGATGACCCTTTT
-3ʹ; HAS1, forward: 5ʹ-GCGATACTGGGTAGCCT
TCA-3ʹ, reverse: 5ʹ-GGTTGTACCAGGCCTCA
AGA-3ʹ; HAS2, forward: 5ʹ-CCTCATCATCCAAAGC
3
CTGT-3ʹ, reverse: 5ʹ-AAACAGTTGCCCTTTGCATC
-3ʹ; HAS3, forward: 5ʹ-GTCATGTACACGGCCT
TCAA-3ʹ, reverse: 5ʹ-CCTACTTGGGGATCC
TCCTC-3ʹ; HYAL1, forward: 5ʹ-GTGCTGCCCTAT
GTCCAGAT-3ʹ, reverse: 5ʹ-ATTTTCCCAGCTCACC
CAGA-3ʹ; HYAL2, forward: 5ʹ-TCTACCATTGGCGA
GAGTG-3ʹ, reverse: 5ʹ-AGCAGCCGTGTCAGGTAAT
-3ʹ; HYAL3, forward: 5ʹ-CCTCCAGTGCCCTCTTCC
-3ʹ, reverse: 5ʹ-CTGTCCCAGGATGACCT
TGT-3ʹ; AQP3, forward: 5ʹ-CATCTACACCCTGGCAC
AGA-3ʹ, reverse: 5ʹ-GGCTGTGCCTATGAACTGGT
-3ʹ; LOR, forward: 5ʹ-CATGATGCTACCCGAGGTTT
-3ʹ, reverse: 5ʹ-ACTGGGGTTGGGAGGTAGTT-3ʹ;
FLG, forward: 5ʹ-GTTACAATTCCAATCCTGTTG
TTTTC-3ʹ, reverse: 5ʹ-CGTTGCATAATACCTTG
GATGATC-3ʹ; IVL, forward: 5ʹ-GTGGGGGAGAGAG
GGAATTA-3ʹ, reverse: 5ʹ-CTCACCTGAGGTTGGG
ATTG-3ʹ; β-actin, forward: 5ʹ-CGTGGACATCCGCA
AAGA-3ʹ, reverse: 5ʹ-GAAGGTGGACAGCGAGGC
-3ʹ; IL-1β, forward: 5ʹ-AAGCTGAGGAAGATGCTG
-3ʹ, reverse: 5ʹ-ATCTACACTCTCCAGCTG-3ʹ; TLR2,
forward: 5ʹ-CAGGAGCTCTTAGTGACCAAGTGAA
-3ʹ, reverse: 5ʹ-CACAAAGTATGTGGCATTGTCCAG
-3ʹ. The parameter of qRT-PCR was conducted as
a previous research [21]. Expression levels were normal-
ized to those of GAPDH or β-actin mRNA levels.
Western blot analysis
After treated with madecassoside for 24 hours, the
cells (6-well plates, 4 × 105 cells/well) were lysed to
collect the total protein. THP-1 cells were treated
with madecassoside for 4 hours and stimulated by
P. acnes for 1 hour before using a nuclear protein
and cytoplasmic protein extraction kit (KeyGEN
BioTECH, Nanjing, China) to get nuclear and
4 X. SHEN ET AL.

cytoplasmic proteins. Protein samples (20 μg) were Statistical analysis

separated by SDS-PAGE before transferred to
PVDF membranes (Millipore, Bedford, MA, USA).
After blocking with 5% BSA for 2 hours, HAS1,
HAS2, HAS3, IVL, NF-κB p65 (1:1000), Lamin
B (1:200), GAPDH (1:2000) and β-actin (1:6000)
were incubated with primary antibodies overnight
and HRP-conjugated antibodies for 2 hours. The
immune complexes of antibody were visualized
with an ECL system (Biotech well, Shanghai,
China). Band intensity was quantified by Smart
View Bio-electrophoresis Image Analysis System
(Furi, Shanghai, China).
Immunofluorescence assay
THP-1 cells (6-well plates, 4 × 105 cells/well), either
treated or untreated with madecassoside, stimulated
or unstimulated with P. acnes, were harvested and
rinsed. Next, cells were treated as previous
described [21] and then slides were sealed by anti-
fluorescence mounting medium and observed with
a fluorescent microscope.
All data were presented as the mean ± standard
errors. One-way analysis of variance (ANOVA)
and LSD were employed to analyze the significant
differences between groups with IBM SPSS V22.0.
P < 0.05 was considered statistically significant.
Results
Effects of madecassoside on IL-1β in P. acnes-
stimulated THP-1 cells
The level of IL-1β, a representative pro-
inflammatory cytokines and possible therapeutic
targets in acne [3], was analyzed by ELISA in
P. acnes induced THP-1 cells. Madecassoside (up
to 500 μM) exhibited slight effect on the viability of
THP-1 cells (> 90%; Figure 1(b)). Similar to our
previous research [6], secretion of IL-1β was low in
untreated groups while drastically increased in
groups incubated with live P. acnes (P. acnes:
THP-1 = 100:1). When madecassoside (250 μM
and 500 μM) was added in the system of

Figure 2. Effects of madecassoside on TLR2 and the nuclear translocation of NF-κB in P. acnes-stimulated THP-1 cells. (a) The
mRNA expression of TLR2 was measured by qRT-PCR. (b) The protein expression of TLR2 was measured by ELISA. The
translocation of NF-κB was detected by immunofluorescence (c) and western blot (d, e). The control group was incubated
without madecassoside and bacteria. Each value represents the mean ± SD of triplicate experiments. (##) P < 0.01 compared
with control group; (*) P < 0.05 and (**) P < 0.01compared with only P. acnes stimulated group.
 SKIN HYDRATION & ANTI-INFLAMMATION EFFECTS OF MADECASSOSIDE 5

Figure 3. Effects of madecassoside on the HaCaT cell viability and AQP3, FLG, LOR, IVL expression in HaCaT. (a) Effect of
madecassoside on HaCaT cell viability was monitored by MTT. (b) Effects of madecassoside on mRNA expression of AQP3, FLG,
LOR and IVL were analyzed by qRT-PCR. (c, d) Effects of madecassoside on protein expression of AQP3 and LOR of HaCaT cells
were measured by ELISA. (e) Effect of madecassoside on protein expression of IVL of HaCaT cells was measured by western blot.
Each value represents the mean ± SD of triplicate experiments. (*) P < 0.05 and (**) P < 0.01 as compared with control group.

cocultivation, the release of IL-1β was significantly
inhibited (Figure 1(c)). We further analyzed the
mRNA levels of IL-1β by qRT-PCR. As shown in
Figure 1(d), madecassoside (500 μM) significantly
inhibited the transcription of IL-1β in P.acnes-
stimulated THP-1 cells.
Effects of madecassoside on TLR2 in P.
acnes-stimulated THP-1 cells
TLR2 signaling is a target for suppressing inflam-
matory cytokine response inflammatory conditions
and has been regarded to lead to a partial induc-
tion of IL-1β [3]. Here we studied the effect of
madecassoside on TLR2 signaling by qRT-PCR
and ELISA. The data (Figure 2(a,b)) revealed that
TLR2 transcription and protein levels were both
significantly increased after P. acnes stimulation,
while markedly decreased after the treatment of
madecassoside at concentration of 500 μM.
Effects of madecassoside on NF-κB signaling
pathway in P. acnes-stimulated THP-1 cells
An immunofluorescence assay and western blot ana-
lysis were used to figure out the effects of madecasso-
side on the NF-κB translocation. After incubation
with P. acnes, NF-κB translocated from the cytoplasm
to the nucleus. And the fluorescence intensity
increased, indicating that the level of NF-κB also
increased (Figure 2(c)). Compared to that in the
stimulated group, the P. acnes-induced translocation
of NF-κB was markedly suppressed under the pre-
treatment of 250 and 500 μM madecassoside for
4 hours (Figure 2(d,e)).
Effects of madecassoside on LOR, IVL, FLG and
AQP3 in HaCaT
Madecassoside at a dosage of 0–100 μM exhibited
slight effect on the viability of HaCaT (> 90%;
Figure 3(a)). Quantified RT-PCR and ELISA were
applied to elucidate whether madecassoside affected
moisturizing contributors (LOR, IVL, AQP3 and
FLG) of epidermis. Compared with the untreated
groups, the mRNA expression of LOR, IVL and
AQP3 were significantly upregulated in a dose-
dependent manner (Figure 3(b)) while FLG insignif-
icantly changed. To confirm the results, following
detections of protein expression of AQP3, LOR and
IVL were conducted and significant promotion was
observed in the group treated with 100 μM madecas-
soside (Figure 3(c-e)).
6 X. SHEN ET AL.

Figure 4. Effect of madecassoside on the HDF cell viability and HA content. (a) Effect of madecassoside on HDF cell viability was
monitored by MTT. (b) HA content was measured by ELISA. (c) Effects of madecassoside on mRNA expression of HA related
enzymes were analyzed by qRT-PCR. (d) HAS1, HAS2 and HAS3 protein levels were analyzed by western blot respectively. (e) The
ROS changes were monitored by using ROS assay kit and the result was confirmed by protein contents. Each value represents
the mean ± SD of triplicate experiments. (*) P < 0.05 and (**) P < 0.01 as compared with control group; (##) P < 0.01 compared
with only H2O2 stimulated group.

Effects of madecassoside on HA content in HDF on H2O2-stimulated ROS expression (Figure 4(e))

HDF cells treated with madecassoside at a dosage of
0–50 μM exhibited slight effect on cell viability
(> 90%; Figure 4(a)). ELISA was adopted to detect
the effect of madecassoside on the content of HA.
Compared with the untreated group, hyaluronan pro-
duced by madecassoside treated HDF was upregu-
lated in a dose-dependent manner (Figure 4(b)).
HDF treated with 50 μM madecassoside produced
33.37 ± 2.20 ng/mL HA, a bit higher than the group
treated with GlcNAc (32.66 ± 1.91 ng/mL).
Effects of madecassoside on HA synthesis and
degradation in HDF
Then the mRNA expression of some HA related
genes (HA synthases and hyaluronidase) and ROS,
a nonenzymatic factors, were closely analyzed to elu-
cidate the mechanism of HA upregulation. The
results (Figure 4(c)) showed that three HA synthases
(HAS1, HAS2, HAS3) were significantly upregulated
with a dose-dependent pattern while hyaluronidases
(HYAL1, HYAL2, HYAL3) exhibited no significant
changes in mRNA expression. Further study on HAS
protein expression (Figure 4(d)) was basically in
accordance with the mRNA expression. But
a marked suppression with a dose-dependent pattern
was observed.
Discussion
Madecassoside is one of the most studied terpenoids
saponin in Centella asiatica with multiple biological
activities in different types of cells [22–25]. And
recent literature has highlighted its skin beneficial
activities including wound healing, UV induced
inflammation, antioxidation, etc [23,26,27]. Our
in vitro study exhibited the skin protective effect
(anti-inflammation and hydration) of madecassoside
and thereby madecassoside might be a contribution
component of moisturizing and anti-inflammatory
properties of Centella asiatica demonstrated by
a previous in vivo study [16].
Acne inflammation is the biggest threat to skin
integrity for the young people and adverse to skin
homeostasis [2]. Some other bacterial strains (e.g.,
Staphylococcus aureus, Staphylococcus epidermidis)
could also partly exacerbate acne inflammatory
event, but are far less significant than P. acnes [28].
Here we have certified that madecassoside could sig-
nificantly inhibit the production of IL-1β released by
P. acnes stimulated THP-1 cells, which suggested
a suppressed P. acnes-induced inflammatory
response. As a wide variety of studies have unravelled
 SKIN HYDRATION & ANTI-INFLAMMATION EFFECTS OF MADECASSOSIDE
the roles of TLR2 and NF-κB in P. acnes stimulated
inflammation [3,6], we assessed whether madecasso-
side could affect TLR2 and NF-κB in P. acnes-
stimulated THP-1 cells. The decreased TLR2
expression and the inhibited activation of the NF-κB
signaling pathway were observed. NF-κB directly
bound to the IL-1β gene promoter and activated its
transcription [29], so the inhibited translocation of
NF-κB might also account for the downregulation of
IL-1β mRNA expression. And these indicated that
madecassoside could suppress inflammation activities
of P. acnes via a mechanism of diminished NF-κB
activation and TLR2-mediated signaling pathways,
similar to a previous study on Rosmarinus officinalis
extract [30].
Hydration is also vital in skin care as water endows
skin with continuous deformation, suppleness and
overall homeostasis. Skin moisturization relies on
many hydration-related components of epidermis and
dermis. In this study, the marked upregulation expres-
sion of AQP3 was observed in madecassoside treated
HaCaT, which was quite similar to the results of
a study on A. houstonianum ethanolnolic extract [7].
Induction of AQP3 expression is concluded as
a potential therapy for dry skin conditions [31] and
our results thus suggested increased skin hydration
effect as AQP3 facilitates water and glycerol transport.
In addition, a marked increased expression of two
major cornified envelope components (LOR and IVL)
suggested an enhanced barrier function and thus less
skin moisture loss [8]. The mRNA expression of filag-
grin was not increased and it might disclose an inactive
effect of madecassoside on NMF increment. Thereby,
the increased expression of AQP3 and two envelope
proteins might be concluded as three epidermal hydra-
tion targets of madecassoside.
Another well-known water binding substance in
skin is HA. HA regulates water balance and osmotic
pressure, promotes cell motility and proliferation,
participates in wound healing, etc [9]. Our data
suggested that madecassoside could increase the
production of HA in HDF, thereby contributing to
the hydration of dermis layer due to the increment
of HA-bound water. As skin hydration is considered
to be the key step of anti-aging care, our findings
might also partially account for in vivo results of the
improvement of madecassoside in skin aging state
[32]. Many researches on hydration or aging
included the analysis of HAS, HYAL and ROS to
elucidate changes in HA level [8,12,33]. And the
upregulation of three HA synthases and inhibition
to ROS formation observed could partially account
for the increment of HA content. The findings were
also in accord with a previous study, which showed
it was HAS instead of HYAL that play a major role
in hyaluronan production by human fibroblasts
[34]. As it was reported that transforming growth
7
factor-β1 (TGF-β1), basic fibroblast growth factor
(bFGF), epidermal growth factor (EGF), and plate-
let-derived growth factor-BB (PDGF-BB) could
upregulate HAS genes expression, further studies
are needed to explain the mechanism underlying
madecassoside-induced changes in HAS1, HAS2
and HAS3 mRNA expression [35]. Owing to the
unique wound healing properties of madecassoside
[23] and the important role that HA played in
wound healing [9], there might be some connec-
tions between our findings about HA and wound
healing activities of madecassoside.
In conclusion, this study exhibited that madecas-
soside could enhance skin hydration through upre-
gulating key moisturizing contributors (AQP3, LOR,
IVL and HA) in skin and inhibiting P. acne-related
inflammation to keep skin healthy. The in vitro find-
ings of this study supported further exploration of
madecassoside as ingredient of moisturizers with pro-
tective effect against acne inflammation.
Author contributions
Xueqing Shen, Yanhua Lu, Zhi Lu and Dan Liu conceived
and designed the study. Xueqing Shen, Miaomiao Guo and
Haiyuan Yu performed the experiments. Xueqing Shen
wrote and edited the manuscript.
Disclosure statement
The authors have no conflicts of interest to declare.
Funding
This work was supported by the Industry-University-
Institute collaboration of Baoshan District Committee of
Science and Technology (No. 17-C-1) and the
Fundamental Research Funds for the Central Universities
(No. 22221818014).
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