0.4 - L`Oréal Congress

P405
PROTECTIVE ACTIVITIES OF A BOTANICAL COMPLEX AGAINST
UV-INDUCED DAMAGE THROUGH THE MODULATION OF GENE EXPRESSION
P. Benech1; 4 , A. Patatian1, E. Lati2, P. Gasser2, L. Peno-Mazzarino2, E. Sandager3, C. Montastier3
1 LABORATOIRE GENEX, Longjumeau, France, 2 LABORATOIRE BIO-EC, Longjumeau, France,
3 HELENA RUBINSTEIN, Levallois-Perret, France, 4 AIX MARSEILLE UNIVERSITÉ, CNRS, NICN UMR 725, France
Introduction
UV rays are one of the most crucial environmental phenomenons in regards to their its capacity to induce premature skin ageing. This effect results from the disruption of homeostasis due to disregulated
ROS production induced by inflammation and mitochondrial alteration. The resulting accumulative damage affects cell growth controls and consequently cell renewal, leading to replicative senescence,
which contributes to skin ageing.
Since most of these effects rely on drastic changes at a gene expression level, whole human genome microarrays constitute a valuable tool to evaluate the capacity of ingredients to repair or reduce the
UV-induced alterations. Using this technology, the effects of pretreating skin explants with a botanical complex (BC) (composed of Litchi sinensis, Grifola frondosa, Scutellaria baicalensis, Nymphaea alba,
Senna Alata, Ophiopogon Japonicus & Oryza Sativa peptides) were investigated.
Material & methods
Skin explants were issued from a 60 year old female donor and incubated 24h before treatment. BC or its
excipient alone, as the control, was loaded on the skin surface every 24h for 4 days. Skin explants were either
left non-irradiated or irradiated with UVA and UVB rays [4 DEM (UVA =18J/cm2; UVB 0.6J/cm2)] for 3.5h. After 6
and 24h, RNAs were extracted and processed into fluorescent cRNAs for hybridisation to Agilent whole human
genome 4X44K microarrays. UV-modulated genes whose expression was reversed by the treatment as well as
genes regulated by BC alone were submitted to PredictSearch™, a text mining-based software, dedicated to
identifying functional networks.
Results
UV/ Excipient
6h
24h
1.10.0
0.70.0
0.80.0
1.0
1.8
0.7
1.1
1.90.1
1.30.1
2.20.1
3.00.3
0.80.0
1.00.1
1.60.1
0.90.1
1.20.0
1.30.1
1.20.0
A_23_P89431
3.20.4
2.00.1
1.30.1
1.10.0
0.90.1
2.00.1
CCL20
A_23_P17065
3.9
3.3
0.9
1.1
2.2
0.9
CCL24
A_23_P215491
3.1
2.2
1.0
0.9
1.1
1.1
PTGS2
A_24_P250922
4.30.3
2.60.3
0.70.0
1.30.1
0.60.1
0.90.0
6h
24h
6h
GeneName
ProbeName
IL6
A_23_P71037
2.30.2
2.00.1
1.90.1
IL8
A_32_P87013
10.0
16.1
IL1A
A_23_P72096
2.40.3
IL1B
A_23_P79518
CCL2
These results demonstrate that whereas BC pretreatment may enhance rather than repress the UV-induced expression of cytokines
such as IL-1A, IL-6 and IL-8, it elicits a significant attenuation of UV-induced genes as well as target genes of the AP1 complex. Indeed,
BC pretreatment leads to the decrease at 6h of the UV-induced expression of FOS and JUN, which are known to form the AP1 early
response transcription factor (Figure 1).
BC/ Excipient
BC+UV/UV
24h
On the one hand, UV rays induce a DNA damage response and apoptosis to avoid damaged cells from proliferating. On the other hand,
survival activities take place to allow cells to reinitiate a normal cell cycle. ATF3 and c-Jun activity, both induced in response to UV
irradiation, highlight these opposite processes (Figure 1). While ATF3 promotes apoptosis and cell cycle arrest through the inhibition
of the cyclin kinase D1 promoter [1], c-Jun is required for maintaining sufficient cyclin D1 activity to enable cell cycle progression [2].
Interestingly, BC pretreatment prevented at 6 and 24h the UV-repressed expression of CLK2 and PPAN that promotes cell survival and
inhibits cell growth arrest [3, 4] as well as at 24h of TGFB1, an important negative regulator of keratinocyte proliferation (Figure 2).
Thus, in UV-irradiated cells pretreated with BC, the maintenance of TGFB1 expression at 24h may result from the reduced expression
of FOS and JUN at 6h. A maintained expression of TGFB1 may preserve keratinocyte motility capacity, which is known to be stimulated
by this factor.
Table 1: Expression of genes related to inflammation. Values are expressed as a fold change calculated from
ratios between the different conditions (UV versus excipient, BC pretreatment followed by UV exposure
versus UV alone, BC treatment versus excipient). When the same sequence probe for one given gene
was spotted at different locations in an array, the mean of the fold change was calculated and standard
deviation is indicated in exponent. Upregulated or downregulated gene expression were determined
according to a fold change (noted in bold) ≥1.5 or ≤0.6, respectively.
UV/ Excipient
6h
24h
BC/ Excipient
BC+UV/UV
6h
24h
6h
0.4
0.8
2.1
24h
GeneName
ProbeName
FOS
A_23_P106194
6.4
0.4
JUN
A_23_P201538
5.00.6
1.40.0
JUNB (a)
A_24_P241815
3,8
0,6
0.7
1.6
1.1
1.0
JUNB (b)
A_23_P4821
1.9
0.9
0.6
1.4
1.0
0.9
JUND
A_23_P365610
1.9
2.7
0.9
1.0
0.8
1.1
ATF3(a)
A_24_P33895
4.3
1.6
0.3
0.8
0.3
1.7
ATF3(b)
A_23_P34915
3.9
2.7
0.4
1.1
0.4
1.2
PMAIP1
A_23_P207999
5.20.9 5.50.3
0.40.0
1.30.0
0.60.1
0.90.1
CDKN1C
A_23_P428129
2.4
2.9
0.5
1.9
0.5
1.2
1.90.1
0.60.0
1.10.0
1.40.1
1.40.1
0.40.1 0.90.0
0.8
1.80.2 0.70.0
MMP9
A_23_P40174
1.60.1
MMP1
A_23_P1691
0.80.0
2.10.1
1.70.1
1.50.1
1.10.1
1.20.0
HUS1B(a)
A_23_P133730
2.0
3.1
0.5
2.2
0.3
0.3
HUS1B(b)
A_23_P423974
2.0
2.7
0.6
1.8
0.4
0.9
OXSR1(a)
A_24_P26114
3.2
2.9
0.4
0.9
1.8
0.6
OXSR1(b)
A_23_P418413
1.2
2.1
0.8
1.2
1.0
1.1
Activation
Activation
Inhibition
Activation
Inhibition
t
c
Ex
/
UV
6h
24h
/UV
V
+U
BC
6h
24h
/
6h
BC
ci
x
E
24h
ProbeName
TGM3
A_23_P57118
1.1
0.4
0.8
1.7
1.3
1.1
CLDN5(a)
A_23_P6321
1.0
0.7
1.7
1.0
1.5
1.1
CLDN5(b)
A_24_P79300
0.7
0.3
2.3
1.5
1.7
1.1
CLDN10
A_23_P48350
0.5
0.5
1.1
2.91
0.1
2.9
MVD
A_23_P152125
1.0
0.5
1.1
1.9
1.1
1.4
SC4MOL
A_23_P110184
0.4
0.4
0.7
1.8
0.9
1.0
SREBF2(a)
A_23_P419602
1.2
0.6
0.9
1.5
1.1
1.0
SREBF2(b)
A_23_P166502
1.0
0.3
1.0
2.5
1.1
1.2
ELOVL1
A_23_P23380
0.4
0.3
1.2
2.4
0.6
1.0
SCNN1A
A_24_P128323
0.8
0.3
1.0
2.3
0.7
1.1
AQP3(a)
A_23_P112481
0.4
0.4
1.2
1.5
0.7
0.9
AQP3(b)
A_23_P112482
0.5
0.1
1.1
2.6
0.8
0.9
PPAN
A_23_P27493
0.2
0.5
2.2
1.0
0.9
1.3
PPAN
A_23_P16096
0.3
0.3
2.0
1.9
0.71
1.3
CLK2(a)
A_23_P85888
0.4
0.5
2.3
1.5
1.3
1.1
CLK2(b)
A_32_P79610
0.3
0.4
2.3
1.5
1.0
0.9
CLK2(c)
A_24_P67027
0.4
0.7
2.0
1.3
1.2
1.0
TGFB1
A_24_P250922
1.10.1 0.50.0 0.90.1 2.10.1 1.10.0 1.30.1
TELO2
A_23_P206237
1.0
1.1
1.9
1.3
t
n
pie
GeneName
0.4
1.2
Gene expression
UV6+/24+ or Inhibition
UV6-/24- Gene expressionor
induced or repressed
by UV induced or r
or
complex
Gene expression induced or repressed by Botanical
complex
UV6+/24+ or UV6-/24- Gene expression induced or repressed by UV rays
or
Gene expression induced or repressed by Botanical
complex
Figure 1: Differential expression of UV-induced genes in absence or in presence of BC pretreatment.
Table 2: Expression of genes related to AP1, apoptosis, cell cycle, ECM degradation, DNA damage and
oxidative stress response.
n
e
i
ip
UV6+/24+ or UV6-/24- Gene expression induced or
TELO2, identified as a factor required for telomere length maintenance, functions as an S-phase checkpoint protein in the cell
cycle and like TGFB1, can be considered as a regulator of the DNA damage response. Interestingly, conditional deletion of TELO2
in embryonic fibroblasts has been shown to compromise the response of embryonic fibroblasts to UV rays [5]. It has been
proposed that TELO2 links cell cycle progression, apoptosis, and telomere length regulation [6]. Furthermore, our results suggest
strongly that BC protects skin dryness triggered by UV through maintaining the expression of genes, which ultimately ensures
cutaneous hydration (Figure 2). Although BC treatment and UV exposure share the ability to increase IL1A, the downstream
signaling pathways, including NFkB and p38 (MAPK), which lead to regulate ATF3 and PMAIP1/NOXA, may differ.
Figure 2: Differential expression of UV-repressed genes in absence or in presence of BC pretreatment
Table 3: Genes for which BC pretreatment prevents the UV-repressed expression.
CONCLUSION
The cosmetic skincare product, which contains BC will prevent excessive UV-induced apoptosis, reducing cellular stress, and will protect the skin barrier against UV-induced damage. According to
these protective effects and to its ability to maintain TELO2 expression, our results suggest that a cosmetic product with this BC is a powerful anti-ageing skincare.
References
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2.Wisdom R1, Johnson RS, Moore C. c-Jun regulates cell cycle progression and apoptosis by distinct mechanisms. EMBO J. 1999 Jan 4;18(1):188-97.
3.Nam SY1, Seo HH, Park HS, An S, Kim JY, Yang KH, Kim CS, Jeong M, Jin YW. Phosphorylation of CLK2 at serine 34 and threonine 127 by AKT controls cell survival after ionizing radiation. J Biol Chem. 2010 Oct 8;285(41):31157-63
4.Wisdom R1, Johnson RS, Moore C. c-Jun regulates cell cycle progression and apoptosis by distinct mechanisms. EMBO J. 1999 Jan 4;18(1):188-97.
5.Takai H1, Wang RC, Takai KK, Yang H, de Lange T. Tel2 regulates the stability of PI3K-related protein kinases. Cell. 2007 Dec 28;131(7):1248-59.
6.Jiang N1, Bénard CY, Kébir H, Shoubridge EA, Hekimi S. Human CLK2 links cell cycle progression, apoptosis, and telomere length regulation. J Biol Chem. 2003 Jun 13;278(24):21678-84
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