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Irradiation of Skin with Visible Light Induces Reactive Oxygen Species and Matrix-Degrading Enzymes

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Irradiation of Skin with Visible Light Induces Reactive Oxygen Species and Matrix-Degrading Enzymes

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Daily skin exposure to solar radiation causes cells to produce reactive oxygen species (ROS), which are a primary factor in skin damage. Although the contribution of the UV component to skin damage has been established, few studies have examined the effects of non-UV solar radiation on skin physiology. Solar radiation comprises <10% of UV, and thus the purpose of this study was to examine the physiological response of skin to visible light (400-700 nm). Irradiation of human skin equivalents with visible light induced production of ROS, proinflammatory cytokines, and matrix metalloproteinase (MMP)-1 expression. Commercially available sunscreens were found to have minimal effects on reducing visible light-induced ROS, suggesting that UVA/UVB sunscreens do not protect the skin from visible light-induced responses. Using clinical models to assess the generation of free radicals from oxidative stress, higher levels of free radical activity were found after visible light exposure. Pretreatment with a photostable UVA/UVB sunscreen containing an antioxidant combination significantly reduced the production of ROS, cytokines, and MMP expression in vitro, and decreased oxidative stress in human subjects after visible light irradiation. Taken together, these findings suggest that other portions of the solar spectrum aside from UV, particularly visible light, may also contribute to signs of premature photoaging in skin.
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Irradiation of Skin with Visible Light Induces Reactive
Oxygen Species and Matrix-Degrading Enzymes
Frank Liebel
1
, Simarna Kaur
1
, Eduardo Ruvolo
2
, Nikiforos Kollias
2
and Michael D. Southall
1
Daily skin exposure to solar radiation causes cells to produce reactive oxygen species (ROS), which are a
primary factor in skin damage. Although the contribution of the UV component to skin damage has been
established, few studies have examined the effects of non-UV solar radiation on skin physiology. Solar radiation
comprises o10% of UV, and thus the purpose of this study was to examine the physiological response of skin to
visible light (400–700 nm). Irradiation of human skin equivalents with visible light induced production of ROS,
proinflammatory cytokines, and matrix metalloproteinase (MMP)-1 expression. Commercially available
sunscreens were found to have minimal effects on reducing visible light–induced ROS, suggesting that UVA/
UVB sunscreens do not protect the skin from visible light–induced responses. Using clinical models to assess
the generation of free radicals from oxidative stress, higher levels of free radical activity were found after visible
light exposure. Pretreatment with a photostable UVA/UVB sunscreen containing an antioxidant combination
significantly reduced the production of ROS, cytokines, and MMP expression
in vitro
, and decreased oxidative
stress in human subjects after visible light irradiation. Taken together, these findings suggest that other portions
of the solar spectrum aside from UV, particularly visible light, may also contribute to signs of premature
photoaging in skin.
Journal of Investigative Dermatology (2012) 132, 1901–1907; doi:10.1038/jid.2011.476; published online 9 February 2012
INTRODUCTION
The contribution of the UV (290–400 nm) component of solar
irradiation on the skin has been well studied; more recently,
studies have begun to explore the effects of non-UVR on skin
physiology. The spectral distribution of the solar energy at the
sea level comprises roughly 3–7% of UVR (290–400 nm),
44% of visible light (400–700 nm), and 53% of infrared (IR)
radiation (700–1440 nm; Frederick et al., 1989). The effects
of UVR are well documented. UVR from sunlight normally
consists of three wavelength regions: UVC (which is absorbed
by ozone in the atmosphere), UVB, and UVA. Energy from
the shorter-wavelength UVB is absorbed in greater amounts
by the epidermis and by keratinocyte DNA, compared
with the energy from UVA, which penetrates more deeply
into the dermal layers of the skin. Visible and IR light
wavelengths penetrate deep into the dermis and have been
thought to, following absorption, only produce heat. In
contrast to the extensive research on the damaging effect of
UV, few studies have looked at the effects of visible light
on skin.
It has been known for decades that reactive oxygen
species (ROS) produced in the skin following UV irradiation
are key mediators of oxidative damage to the skin. Cell
damage from UV also occurs through peroxidation of
membrane lipids via generation of lipid peroxides. UV
irradiation results in the rapid depletion of several endogen-
ous skin enzymes and antioxidants such as glutathione
reductase and catalase, as well as glutathione, tocopherol,
and ubiquinone. Exposure to UV has also been shown to
induce proinflammatory cytokines, such as IL-1a, and matrix
metalloproteinases (MMPs) in skin cells such as keratinocytes
and fibroblasts (Wlaschek et al., 1994; Wan et al., 2001).
Activation of MMPs leads to the breakdown of collagen,
the major structural component of skin, and also inhibits
new collagen synthesis. Induction of inflammatory cytokines
by UV, leading to overall skin inflammation, is another
significant contributor to the photoaging process in skin.
In addition to UV, IR has also been shown to cause oxi-
dative stress to skin; exposing human fibroblasts to IR
led to increased ROS formation, and enhanced expression
of MMPs (Schroeder et al., 2008). Additional studies exposed
photoprotected sites of healthy human skin to solar relevant
doses of IR radiation, and observed that the MMP-1
expression in the dermis, but not the epidermis, was
upregulated in 80% of the tested individuals (Cho et al.,
2008).
See related commentary on pg 1756
&2012 The Society for Investigative Dermatology www.jidonline.org 1901
ORIGINAL ARTICLE
Received 14 April 2011; revised 1 September 2011; accepted 4 October
2011; published online 9 February 2012
Parts of this work have been presented in a poster format at the Society for
Investigative Dermatology Annual Meeting, 2010.
1
Preclinical Pharmacology, Johnson & Johnson Skin Research Center, CPPW,
a Unit of Johnson & Johnson Consumer Companies, Skillman,
New Jersey, USA and
2
Measurement Sciences, Johnson & Johnson Skin
Research Center, CPPW, a Unit of Johnson & Johnson Consumer Companies,
Skillman, New Jersey, USA
Correspondence: Michael D. Southall, Preclinical Pharmacology, Johnson &
Johnson Skin Research Center, CPPW, a Unit of Johnson & Johnson Consumer
Companies, 199 Grandview Road, Skillman, New Jersey 08558, USA.
E-mail: msoutha@its.jnj.com
Abbreviations: IR, infrared; MMP, matrix metalloproteinase; ROS, reactive
oxygen species
Although sunlight comprises up to 44% of visible light,
few studies have sought to determine the effects of visible
light on skin. Commercial sunscreens are designed to only
block wavelengths up to 380 nm, and thus skin with topical
sunscreens is not protected from the effects of visible light.
In the current study, we examined the effect of visible light
on the ROS, and MMP responses in skin in vitro. We report
that visible light can induce significant ROS production, and
this ROS mediates the release of proinflammatory cytokines
and MMP expression. Skin is exposed to visible light for
substantial durations of the day, and as skin contains several
chromophores for visible light the cumulative effects of
visible light could result in skin damage, which may
contribute to premature skin aging.
RESULTS
Exposure to visible light induces increases in epidermal ROS,
cytokine, and MMP production
To assess the role of visible light on skin, human epidermal
equivalents were exposed to a dose–response of visible
light, and the production of ROS, inflammatory cytokines,
and MMPs were determined. Visible light induced a dose-
dependent increase in intracellular hydrogen peroxide
formation, with visible light doses of 65, 130, and 180 Jcm
2
increasing ROS by 5-, 9-, and 18-fold, respectively
(Figure 1a). In comparison, a dose of 6.0 J cm
2
from a solar
simulator increased ROS by 19-fold. Visible light was also
found to increase the release of proinflammatory cytokines
from epidermal equivalents. IL-1arelease was increased
1.1-, 1.5-, and 2.5-fold after exposure to visible light doses
of 65, 130, and 180 J cm
2
, respectively, with 6.0 J cm
2
from a solar simulator increasing IL-1arelease by 2.8-fold
(Figure 1b). A similar effect was seen with release of IL-1
receptor antagonist, IL-6, GM-CSF, and IL-8 (data not shown).
In contrast, UV was found to increase the release of tumor
necrosis factor-a(TNFa), whereas visible light, even at doses
that induced other proinflammatory mediators, did not
increase TNFarelease. MMP release was also increased after
exposure to visible light. MMP-1 release was increased by
2-fold from visible light doses, compared with 2.8-fold
by solar simulator (Figure 1c), and MMP-9 release was
similarly increased by approximately 2-fold from visible light
doses.
Exposure to visible light induces the EGFR–p42/44 MAPK
pathway
The EGFR/extracellular signal–regulated kinase pathway has
been shown to be activated by UV irradiation in keratino-
cytes and other cell types.
2
To examine the effects of visible
light on this pathway, primary human keratinocytes were
exposed to UV solar light or dose–response of visible light
for specific periods of time. Treatment with the positive
control (TNFa) and exposure to UV light or visible light
(65–180 J cm
2
) resulted in increased phosphorylation of the
EGFR detected as phospho-tyrosine and the downstream
marker of proliferation p42/44 MAPK, suggesting activation
250
ab
c
300
250
200
150
100
50
0
200
150
100
50
0
Reactive oxygen species formation
(mean fluorescence units)
MMP-1 release (pg ml )
Inflammatory protein Il-1α
release (pg ml )
No
treatment
No
treatment
*
*
****
*
***
*
Visible light
800
700
600
500
400
300
200
100
0
No
treatment Visible li
g
ht
UVA/UVB Visible light UVA/UVB
UVA/UVB
180Jcm 6Jcm
180 J cm130 J cm65 J cm 6Jcm
130Jcm65Jcm
6Jcm180Jcm130Jcm65Jcm
Figure 1.Human epidermal skin equivalents were exposed to the indicated dose of visible light or UVA/UVB, and reactive oxygen species, proinflammatory
cytokine, or matrix metalloprotease (MMP) production was measured. Exposure to visible light significantly increased (a) reactive oxygen species, (b) IL-1a,
and (c) MMP-1 production in a dose-dependent manner. An asterisk indicates a significant difference (Po0.05) compared with unexposed skin using
analysis of variance with Newman–Keuls post hoc test.
1902 Journal of Investigative Dermatology (2012), Volume 132
F Liebel et al.
Visible Light Induces ROS, Inflammatory Cytokines, and MMPs in Skin
of the EGFR–ERK pathway (Figure 2a and b). Total EGFR and
ERK were also measured to show uniform protein loading.
Pretreatment of keratinocytes with 100 nMof the specific
EGFR inhibitor, Tyrphostin (AG 1478), for 20 minutes before
irradiation blocked the downstream effects of visible light on
the induction of ERK (Figure 2c), suggesting that the increased
phosphorylation of ERK resulted from the intermediate
activation of EGFR. These results demonstrate that visible
light can induce activation of the EGFR pathway in keratino-
cytes in a manner similar to UV.
Visible light does not induce thymine dimer formation
UV exposure is widely known to cause DNA damage in skin,
including the formation of thymine (T–T) dimers (Sinha and
Hader, 2002; Marrot and Meunier, 2008). To investigate
whether visible light also leads to thymine dimer formation,
skin equivalents were treated with visible light or the positive
control (UV) and stained for T–T dimer formation. Whereas
UV led to strong induction of T–T dimers, visible light did not
result in T–T dimer formation even at higher doses of visible
light (Figure 3).
Antioxidants reduce the ROS, cytokine, and MMP
production induced by visible light
To determine whether antioxidants could reduce the
damage caused by visible light, an antioxidant combination
of Feverfew (Tanacetum parthenium) extract, Soy (Glycine
soja) extract, and Gamma Tocopherol were combined into
a UVA/UVB Sunscreen and the effect of visible light on
ROS, IL-1a, or MMP-1 release was determined. Exposure to
65 J cm
2
of visible light resulted in a significant increase in
Untreated
TNF-αUntreated EGF
Solar light
(J cm–2)
Visible light
(J cm–2)
p-ERK
Sham 160 160+
Tyrphostin
Visible li
g
ht (J cm–2)
Total ERK
4665
130 180
p-ERK
Total ERK
TNF-αSham 4 6 65 130 180
P-Tyrosine
Total EGFR
Solar light
(J cm–2)
Visible light
(J cm–2)
Figure 2.Primary human keratinocytes exposed to visible light were
evaluated for the levels of EGFR and ERK activation. (a,b) Primary human
keratinocytes were either left untreated, treated with the positive control
tumor necrosis factor-a(TNFa; 100 ng ml
1
), or exposed to UV solar light or
visible light for calculated periods of time. The sham group was incubated at
room temperature for an equivalent amount of time. Post exposure, the cells
were incubated at 371C for 20minutes, followed by cell lysis. Western blotting
with phospho-tyrosine (molecular weight corresponding to EGFR) and phospho-
extracellular signal–regulated kinase (ERK) antibodies demonstrated activation
of the EGFR–ERK pathway by multiple doses of visible light. (c) Pretreatment
with the selective EGFR inhibitor, Tyrphostin (100nM), for 20 minutes before
irradiation blocked the downstream effects of visible light on the induction of ERK.
Untreated
Visible light
(180 J cm–2)
Visible light
(240 J cm–2)
UV (6 J cm–2)
T–T dimers
Figure 3.Human epidermal skin equivalents were either left untreated or exposed to UV or visible light for calculated periods of time and stained for
T–T dimer formation. Exposure to UV led to induction of T–T dimers, whereas visible light did not have the same effect. Bar¼50 mm.
www.jidonline.org 1903
F Liebel et al.
Visible Light Induces ROS, Inflammatory Cytokines, and MMPs in Skin
ROS, IL-1a, and MMP-1 release. Pretreatment with a lotion
containing UVA/UVB Sunscreen alone had no effect on
reducing the damage from visible light. In contrast, the
addition of antioxidants into the same UVA/UVB Sunscreen
resulted in a significant reduction in the effects of visible
light, reducing ROS, IL-1a, and MMP-1 release by 78%, 82%,
and 87%, respectively (Figure 4). The direct effect of the anti-
oxidants alone, without sunscreen, was tested and similarly
mitigated the ROS, cytokine, and MMP release induced by
visible light (data not shown).
Measuring free-radical production in the skin from visible
light using chemiluminescence
We next sought to confirm the in vitro ROS results by
studying free-radical production on the skin of human
subjects. Areas of the skin high in porphyrin content such
as the forehead responded to low levels of visible light to
induce free-radical production, which could be measured
by photon emission or chemiluminescence. A 50 J cm
2
dose
at 150 mW cm
2
of visible light was able to significantly
increase the amount of free radicals by 85.8% over baseline
measurements (Figure 5a). The addition of an antioxidant
combination comprising Feverfew (T. parthenium) extract,
Soy (G. soja) extract, and Gamma Tocopherol to the sun-
screen was able to significantly reduce the free radicals by
54% (Figure 5b). These results are consistent with the in vitro
ROS results and clearly demonstrate that visible light expo-
sure induces free-radical production by the skin.
DISCUSSION
Although visible light comprises about 44% of solar light, few
studies have looked at just visible light alone. Fuchs et al.
(1989) have observed effects from UVA and visible light on
skin, whereas Cho et al. (2008) have looked at visible light
and IR; however, only Mahmoud et al. (2010) have discussed
the potential effects of visible light on skin. Visible light is the
only portion of the spectrum visible to the human eye and
responsible for general illumination. If we consider that the
solar irradiance in the visible range is about 50 mW cm
2
, the
doses used in this study (40–240 J cm
2
) would be equivalent
to outside midsummer sunlight exposure of approximately
15–90 minutes in Houston, TX. The data presented in
this paper suggest that visible light can produce some of the
same physiological effects as UV, including inflammation, ROS,
and matrix-degrading enzymes being generated in the skin.
It is well known that ROS are produced in skin following
UV irradiation (Pathak and Stratton, 1968) and are major
mediators of oxidative damage to the skin. Singlet oxygen can
be generated from the action of UVA and endogenous
photosensitizers, such as porphyrins and flavins (Ravanat
et al., 2000), which can produce oxidative damage (Pelle
et al., 2003). In the current study, exposing human skin
equivalents to increasing doses, 40–180 J cm
2
, of visible
120 Visible light alone
Visible light + UVA/UVB
sunscreen
Visible light + UVA/UVB
sunscreen with
antioxidant combo
100
80
60
40
Percent response
20
0
ROS
***
IL-1 MMP-1
Figure 4.UVA/UVB sunscreen with an antioxidant reduces reactive
oxygen species (ROS), proinflammatory cytokine (IL-1), and matrix
metalloprotease (MMP) production induced by visible light. Exposure
to visible light significantly increased H
2
O
2
,IL-1a, and MMP-1 production,
and this increase was not affected by the UVA/UVB sunscreen. In contrast,
the UVA/UVB sunscreen containing an antioxidant blend significantly
reduced H
2
O
2
, IL-1a, and MMP-1 production. An asterisk indicates
a significant difference (Po0.05) compared with unexposed skin using
analysis of variance with Newman–Keuls post hoc test.
140
*
120
100
180
60
40
0
Chemiluminescence signal
(counts per second)
% Change vs. baseline
50 J cm–2 visible light exposure
Untreated
*
100
90
80
70
60
50
40
30
20
10
0
No topical treatment Sunscreen
w/ antioxidants
Sunscreen
20
Figure 5.Human subjects were exposed to visible light, and chemiluminescence was measured as a marker of reactive oxygen species. A50Jcm
2
dose
of visible light at 150 mW cm
2
significantly increased free-radical production; an asterisk indicates a significant difference (Po0.05) compared with untreated
skin (N¼40, a). UV sunscreens do not block non-UV longer wavelengths from stimulating free radicals in the skin. The addition of a potent antioxidant
combination to the formulation significantly reduced the number of free radicals (b); *Po0.05 compared with sunscreen alone (N¼12 per group of sunscreens,
N¼24 for visible light alone).
1904 Journal of Investigative Dermatology (2012), Volume 132
F Liebel et al.
Visible Light Induces ROS, Inflammatory Cytokines, and MMPs in Skin
light resulted in a dose-dependent increase in ROS produc-
tion similar to UV (Figure 1a). Visible light has been used in
light therapy for various conditions such as atopic dermatitis
(Byun et al., 2011), eczema (Krutmann et al., 2005), and also
in antimicrobial photochemotherapy (Soukos et al., 1998).
These dermatological treatments using visible light tend to be
focal applications to the affected lesional skin only to treat the
underlying immune response. It is interesting to speculate
that the therapeutic effects of visible light for atopic dermatitis
and eczema may be mediated by the production of ROS in
the skin, similar to the mechanism of psoralen plus UVA
treatments for psoriasis. Irradiation of normal human epider-
mal keratinocytes with UVB can generate a dose-dependent
increase in proinflammatory cytokines and expression of
MMPs (Figure 1a–c), which was also seen with visible light
exposure. IR light has also been shown to be a source of
oxidative stress for skin. Exposing human skin fibroblasts
to near-IR radiation was shown to induce ROS formation
and lead to the subsequent increased expression of MMPs
(Schroeder et al., 2007). Additional studies exposed photo-
protected sites of healthy human skin to solar relevant doses
of IR radiation. MMP-1 expression in the dermis, but not in
the epidermis, was upregulated in 80% of the tested
individuals (Schroeder et al., 2008). Cho et al. (2008) were
able to show that, in addition to UV, IR plus the visible
light spectrum within natural sunlight increases MMP-1 and
MMP-9 expression in vivo. Taken together, these findings
suggest that although the UV fraction of the solar spectrum
is harmful and can cause skin damage, the visible light and
the IR light portions of the spectrum may also induce skin
damage through to the dermis, which could lead to premature
photoaging of skin through the generation of free radicals.
Exposure to UVR has been shown to activate the EGFR–ERK
pathway in keratinocytes (Huang et al., 1996) and in human
skin (Knebel et al., 1996; Katiyar, 2001). The rapid phosphor-
ylation of EGFR by UV is known to be reactive oxygen
intermediate–mediated (Huang et al., 1996; Knebel et al.,
1996).To investigate the effects of visible light on this
pathway, primary human keratinocytes were exposed to
UV solar light or dose–response of visible light. Exposure to
UV light and visible light (65–180 J cm
2
) resulted in increased
phosphorylation of the EGFR (detected as phospho-tyrosine)
and the downstream marker of cell proliferation p42/44
MAPK, suggesting activation of the EGFR–ERK pathway (Figure
2a and b). To determine whether EGFR is a prerequisite for
ERK activation by visible light, human keratinocytes were
pretreated with the specific EGFR kinase inhibitor (Osherov
and Levitzki, 1994; Ellis et al., 2006), Tyrphostin (AG1478),
before visible light treatment. The presence of Tyrphostin
abolished the phosphorylation of ERK, suggesting that EGFR is
required for the downstream induction of ERK by visible light
(Figure 2c). Activation of the EGFR–ERK pathway has been
implicated in UV-induced epidermal hyperplasia (El-Abaseri
et al., 2006), and also in the activation of MMPs that may
accelerate skin aging (Kang et al., 2008).Induction of ERK
signaling by visible light might be upstream of the MMP-1
activation shown in human skin equivalents (Figure 1c), and
potentially any visible light–induced hyperplasia.
DNA damage by UVB irradiation results from photo-
chemical reactions consequent to direct absorption of
photons by DNA bases. The UV-induced DNA lesions that
have been studied in most detail are the cyclobutane
pyrimidine dimer and the 6–4 pyrimidine–pyrimidone photo-
product at adjacent pyrimidines (Nakajima et al., 2004).
Nuclear DNA strand breaks are also readily produced by
incubation of keratinocytes with hydrogen peroxide (Armeni
et al., 2001), and hydroxyl radicals can be generated from
hydrogen peroxide through Fe
2þ
-mediated Fenton-type
reactions (Stewart et al., 1996). Our studies showed that
visible light–irradiated tissues did not induce thymine dimer
formation (Figure 3) even at concentrations sufficient to
induce significant increases in ROS. In comparison, UV
irradiation induced a pronounced DNA damage (Figure 3).
These results suggest that, in contrast to UV, visible light
photons may not be directly absorbed by DNA bases and
therefore may not result in thymine dimer formation. Visible
light could contribute to other forms of DNA damage, such as
8-Oxoguanine, which were not measured in the current
study. Indeed, exposure of AS52 Chinese hamster cells to a
visible light source was reported to produce 8-Oxoguanine
DNA damage with a relative maximum between 400 and
450 nm (Kielbassa et al., 1997).
To confirm the in vitro results, we evaluated the effects of
visible light on oxidative stress in a clinical model. UVA
(320 nm–400 nnm) exposure has been shown to be a source
of oxidative stress in the skin clinically (Ou-Yang et al.,
2004), but no studies have examined ROS from visible light
alone. To study the effects of visible light (400–700 nm) on
the skin of subjects, we used a photon detector to measure
the rate of photon emission or chemiluminescence. A visible
light dose of 50 J cm
2
was able to significantly increase
the photon emission on the skin by 85.8% compared with
baseline measurements (Figure 5a). A broad UVA/UVB
sunscreen was tested for its ability to inhibit free radicals
generated from visible light, and this resulted in no change in
the amount of photon emission. These data show that the UV
sunscreens do not block the longer non-UV wavelengths from
stimulating free radicals in the skin. When an antioxidant
combination including a Parthenolide-Depleted Feverfew
(T. parthenium) extract (Martin et al., 2008) was added to the
UV sunscreen, a 54% decrease in the amount of generated
free radicals from visible light was observed (Figure 5b).
These results are consistent with in vitro study that demon-
strated that a UVA/UVB sunscreen did not affect the visible
light–induced release of ROS, MMP-1, or proinflammatory
cytokines from epidermal tissues; however, the addition of a
potent antioxidant combination to a UVA/UVB sunscreen
significantly reduced the release of all these mediators (Figure 4).
These data demonstrate that sunscreens containing UVA/UVB
sunfilters alone do not prevent the free-radical production
from non-UV longer wavelengths of the solar spectrum.
When UV contacts the skin, the energy of the photons is
absorbed by chromophores, which can mediate the cellular
effects of UV exposure. A variety of chromophores, including
DNA and some aromatic amino acids, are known to absorb
UVR; however, biomolecules that can absorb longer wave-
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F Liebel et al.
Visible Light Induces ROS, Inflammatory Cytokines, and MMPs in Skin
lengths such as visible light are not fully understood. A few
potential targets for the actions of visible light may include
riboflavin, hemoglobin, and bilirubin, all of which show
absorbance peaks in the visible range of the spectrum, and
are found in the skin. Melanin and b-carotene have been
shown to absorb UV light, but can also act as endogenous
chromophores for visible light (Anderson and Parrish, 1981;
Mahmoud et al., 2008). It is also known that the depth of
penetration of optical radiation in skin is wavelength
dependent, and affected by position and absorption spectrum
of the corresponding chromophore in skin (Mahmoud et al.,
2008). In the dermis, optical scattering of light is considered
to be inversely proportional to wavelength and affects the
depth of optical penetration (Anderson and Parrish, 1981).
The approximate depth for penetration of visible light
(400–700 nm) in a fair-skinned Caucasian individual was
estimated to be between 90 and 750 mm by Anderson and
Parrish (1981), compared with a depth of 1.5–90 mm for UVR
(Anderson and Parrish, 1981). Thus, even though visible
light photons are less energetic than UV photons, due to the
deeper dermal penetration visible light may still have a
substantial effect on skin. Taken together, these results
demonstrate that visible light exposure can induce ROS,
which can lead to the release of proinflammatory cytokines
and MMPs in the skin, similar to the effects of UV, and
therefore visible light may contribute to the signs of
premature aging in the skin.
MATERIALS AND METHODS
UV and visible light–induced cytokine release
The UV light source used as a positive control was an 1,000 W
Oriel solar UV simulator (Oriel, Stratford, CT), equipped with an
atmospheric attenuation filter (Schott WG 320, Schott Optical,
Elmsford, NY; 1 mm thick) and a visible–IR filter (Schott UG 11;
1 mm thick). This filtered xenon light source provided a simulated
solar UVR spectrum (290–400nm) that was nearly devoid of visible
and IR radiation as determined using an IL-1400 Research Radiometer
(International Light, Newburyport, MA). The fluence rate of the light
sources was 7.52 mW cm
2
, measured using a calibrated Oriel
Thermopile Model 71767.
The visible light source used was the Fiber-Lite Model 170-D
(Dolan-Jenner Industries, Boxborough, MA) with a 150 W quartz
halogen lamp; a straight 8-mm Dolan-Jenner glass optical fiber was
used for irradiation. The source was filtered with three KG5/3-mm
Schott glass filters and one GG400/3-mm (Schott Optical Company)
was used to filter IR and UVR from the light source, respectively. The
light source was characterized spectrally by a calibrated Optronics
OL750 spectroradiometer, Optronics Laboratories, Orlando, FL.
From the spectroradiometric measurements (from 350 to 1800 nm),
the light source has 0.14% of UVA (350–400 nm), 98.3% of visible
light, 1.7% of IRA (700–1400 nm), and 0.3% of IRB (Supplementary
Figure S1 online). For the doses used in the studies presented in this
paper, the contribution from UVA and IR (A and B) are negligible.
For the highest visible light dose used in this study, 240 J cm
2
,we
would have less than 0.34 J cm
2
of UVA and about 4 J cm
2
of IRA
being delivered to the irradiated sample. The spectral irradiance of
the visible light source is described in Mahmoud et al (2010). Skin
surface temperature of skin equivalents and media temperature
exposed to either UV or visible light sources were measured during
exposure periods and did not increase by 41
1
C.
Cell culture and skin equivalents
Normal human epidermal neonatal keratinocytes were obtained
from Lifeline Cell Technologies (Frederick, MD) and maintained
in Dermalife keratinocyte medium (Lifeline Cell Technologies,
Frederick, MD) with supplements. Reconstituted human epidermis
(EPI-200-HCF) was purchased from MatTek (Ashland, MA). After
reception, epidermal equivalents were incubated with a phenol-free
and hydrocortisone-free maintenance medium (MatTek at 371C for 24 h.
Assessment of UV and visible light–induced ROS and
mediator release from reconstituted epidermis
MatTek epidermal equivalent tissues were topically treated for
45 minutes with 5 mMof the hydrogen peroxide–sensitive fluorescent
probe 5-(and-6)-chloromethyl-20,70-dichlorodihydro-fluorescein dia-
cetate, acetyl ester (Invitrogen, Carlsbad, CA). After incubation, the
plate was rinsed with warm phosphate-buffered saline to remove
excess probe. A volume of 6 ml of sunscreen formulation or Feverfew
extract (10 mg ml
1
, wt/volt) was topically applied and spread
uniformly across the surface of the epidermal equivalent. Tissues
were then incubated for 1 hour before UV or visible light exposure.
Fluorescence was then measured using a CytoFluor Fluorescence
Plate Reader (PerSeptive Biosystems, Framingham, MA) set with the
following filter combination: excitation at 485 nm and emission at
530 nm. The tissues were then exposed to UV or visible light, and imme-
diately after exposure the tissues were measured for fluorescence.
For measurement of cytokines, the equivalents were trans-
ferred back to the maintenance medium and incubated at 371C
for 24 hours. After 24 hours, the medium below each equivalent was
collected and analyzed for secreted IL-1a, IL-1 receptor antagonist,
IL-6, TNFa, GM-CSF, IL-8, MMP-1, and MMP-9 by ELISA, using
commercially available immunoassay multiplex kits (Millipore,
Bedford, MA) on a Luminex L100 (Luminex, Austin, TX).
Western blotting
Primary keratinocytes grown in 12-well plates were either left untreated,
treated with TNFa(100 ng ml
1
) or exposed to UV solar light or
multiple doses of visible light for specific periods of time corresponding
to the doses. The sham group was left at room temperature for an
equivalent amount of time. Post exposure, cells were incubated at 371C
for 20 minutes, then washed with phosphate-buffered saline and lysed
with radioimmunoprecipitation lysis buffer containing 65 mMTris (pH
7.4), 150 mMNaCl, 1 mMEDTA (pH 8), 1% nonidet P-40, 0.25%
sodium deoxycholate, 50 mMNaF, 1 mMNa
3
VO
4
,1mMphenylmethyl-
sulfonyl f luoride, and 1 protease inhibitor cocktail (Sigma, St Louis,
MO). Lysates were centrifuged and total protein was estimated in the
supernatants using a BCA protein assay kit (Pierce–Thermo Fisher
Scientific, Rockford, IL). Protein (30 mg)wasloadedonSDS-PAGE
followed by immunoblotting with the specific antibodies (incubation
with primary antibodies diluted 1:1,000 for overnight at 41C) and
detection using the ECL chemiluminescence detection system
(Amersham Life Sciences–GE Healthcare, Piscataway, NJ).
Assessing DNA damage (thymine dimer staining)
Thymine (T–T) dimer staining of human skin equivalents was performed
as described previously (Martin et al., 2008). In brief, skin equivalents
1906 Journal of Investigative Dermatology (2012), Volume 132
F Liebel et al.
Visible Light Induces ROS, Inflammatory Cytokines, and MMPs in Skin
were either left untreated or exposed to UV or visible light, after
which they were immediately fixed in formalin and sent to Paragon
BioServices (Baltimore, MD) for T–T dimer staining.
Chemiluminescence
Photons emitted from the skin were measured by a red-sensitive,
tri-alkali photomultiplier with a 2.5-cm diameter photocathode surface
(Electron Tubes, Rockaway, NJ, model 9828SA). The photomultiplier
was in a Peltier-cooled housing unit (Electron Tubes, model CDM30)
and was routinely cooled to 18 to –201C to reduce detector noise.
The housing unit can be held with one hand as a probe and placed on
the skin area to be measured. The surface of the photocathode was
protected by a manual shutter (Products for Research, Danvers, Ma,
model PR318). The shutter is opened for measurements and closed
for instrument background. The distance between the skin and the
photocathode surface was approximately 1.5 cm. The photomulti-
plier was supplied with a potential of 1.15 kV of high voltage
(Brandenburg, Brierley Hill, UK, model 477) with a variation less
than ±2 V. After going through an amplifier and discriminator
(Electronic Tubes, model AD6), the output signals from the
photomultiplier were measured in a photon-counting module
(Electronic Tubes, model CT1) and recorded by a computer. All
measurements were recorded in a positively dark room; other-
wise, the background would overwhelm the signal (Ou-Yang
et al., 2004). The forehead was used for all the measurements.
Statistical analysis
All data are presented as mean±SD. For in vitro studies, a one-way
analysis of variance with Newman–Keuls post hoc test was used
to determine significance. A value of Po0.05 was considered
significant. For chemiluminescence results, data were analyzed
using a Student’s t-test with significance set at Po0.05.
CONFLICT OF INTEREST
All authors are employees of Johnson & Johnson.
ACKNOWLEDGMENTS
We thank Jean Krutmann for discussions and suggestions on the effect of
non-UV light on skin.
SUPPLEMENTARY MATERIAL
Supplementary material is linked to the online version of the paper at http://
www.nature.com/jid
REFERENCES
Anderson RR, Parrish JA (1981) The optics of human skin. J Invest Dermatol
77:13–9
Armeni T, Battino M, Stronati A et al. (2001) Total antioxidant capacity
and nuclear DNA damage in keratinocytes after exposure to H
2
O
2
.
Biol Chem 382:1697–705
Byun HJ, Lee HI, Kim B et al. (2011) Full-spectrum light phototherapy for
atopic dermatitis. Int J Dermatol 50:94–101
Cho S, Lee MJ, Kim MS et al. (2008) Infrared plus visible light and heat from
natural sunlight participate in the expression of MMPs and type I
procollagen as well as infiltration of inflammatory cell in human skin
in vivo.J Dermatol Sci 50:123–33
El-Abaseri TB, Putta S, Hansen LA et al. (2006) Ultraviolet irradiation induces
keratinocyte proliferation and epidermal hyperplasia through the activation
of the epidermal growth factor receptor. Carcinogenesis 27:225–31
Ellis AG, Doherty MM, Walker F et al. (2006) Preclinical analysis of the
analinoquinazoline AG1478, a specific small molecule inhibitor of EGF
receptor tyrosine kinase. Biochem Pharmacol 71:1422–34
Frederick JE, Snell HE, Haywood EK (1989) Solar ultraviolet radiation at the
earth’s surface. Photochem Photobiol 50:443–50
Fuchs J, Huflejt ME, Rothfuss LM et al. (1989) Acute effects of near ultraviolet
and visible light on the cutaneous antioxidant defense system.
Photochem Photobiol 50:739–44
Huang RP, Wu JX, Fan Y et al. (1996) UV activates growth factor receptors
via reactive oxygen intermediates. J Cell Biol 133:211–20
Kang KA, Zhang R, Piao MJ et al. (2008) Inhibitory effects of triphlorethol-A
on MMP-1 induced by oxidative stress in human keratinocytes via ERK
and AP-1 inhibition. J Toxicol Environ Health A 71:992–9
Katiyar SK (2001) A single physiologic dose of ultraviolet light exposure to
human skin in vivo induces phosphorylation of epidermal growth factor
receptor. Int J Oncol 19:459–64
Kielbassa C, Roza L, Epe B et al. (1997) Wavelength dependence of oxidative
DNA damage induced by UV and visible light. Carcinogenesis 18:811–6
Knebel A, Rahmsdorf HJ, Ullrich A et al. (1996) Dephosphorylation of
receptor tyrosine kinases as target of regulation by radiation, oxidants or
alkylating agents. EMBO J 15:5314–25
Krutmann J, Medve-Koenigs K, Ruzicka T et al. (2005) Ultraviolet-free
phototherapy. Photodermatol Photoimmunol Photomed 21:59–61
Mahmoud BH, Hexsel CL, Hamzavi IJ et al. (2008) Effects of visible light on
the skin. Photochem Photobiol 84:450–62
Mahmoud BH, Ruvolo E, Hexsel CL et al. (2010) Impact of long-wavelength
UVA and visible light on melanocompentant skin. J Invest Dermatol
130:2092–7
Marrot L, Meunier JR (2008) Skin DNA photodamage and its biological
consequences. J Am Acad Dermatol 58(5 Suppl 2):S139–48
Martin K, Sur R, Liebel F et al. (2008) Parthenolide-depleted Feverfew
(Tanacetum parthenium) protects skin from UV irradiation and external
aggression. Arch Dermatol Res 300:69–80
Nakajima S, Lan L, Kanno S et al. (2004) UV light-induced DNA damage and
tolerance for the survival of nucleotide excision repair-deficient human
cells. J Biol Chem 279:46674–7
Osherov N, Levitzki A (1994) Epidermal-growth-factor-dependent activation
of the src-family kinases. Eur J Biochem 225:1047–53
Ou-Yang H, Stamatas G, Saliou C et al. (2004) A chemiluminescence study of
UVA-induced oxidative stress in human skin in vivo.J Invest Dermatol
122:1020–9
Pathak MA, Stratton K (1968) Free radicals in human skin before and after
exposure to light. Arch Biochem Biophys 123:468–76
Pelle E, Huang X, Mammone T et al. (2003) Ultraviolet-B-induced oxidative
DNA base damage in primary normal human epidermal keratinocytes
and inhibition by a hydroxyl radical scavenger. J Invest Dermatol
121:177–83
Ravanat JL, Di Mascio P, Martinez GR et al. (2000) Singlet oxygen induces
oxidation of cellular DNA. J Biol Chem 275:40601–4
Schroeder P, Lademann J, Darvin ME et al. (2008) Infrared radiation-induced
matrix metalloproteinase in human skin: implications for protection.
J Invest Dermatol 128:2491–7
Schroeder P, Pohl C, Calles C et al. (2007) Cellular response to infrared
radiation involves retrograde mitochondrial signaling. Free Radic Biol
Med 43:128–35
Sinha RP, Hader DP (2002) UV-induced DNA damage and repair: a review.
Photochem Photobiol Sci 1:225–36
Soukos NS, Ximenz-Fyvie L, Hamblin MR et al. (1998) Targeted antimicribal
photochemotherapy. Antimicrob Agents Chemother 42:2595–601
Stewart MS, Cameron GS, Pence BC et al. (1996) Antioxidant nutrients protect
against UVB-induced oxidative damage to DNA of mouse keratinocytes
in culture. J Invest Dermatol 106:1086–9
Wan Y, Belt A, Wang Z et al. (2001) Transmodulation of epidermal growth
factor receptor mediates IL-1 beta-induced MMP-1 expression in
cultured human keratinocytes. Int J Mol Med 7:329–34
Wlaschek M, Heinen G, Poswig A et al. (1994) UVA-induced autocrine
stimulation of fibroblast-derived collagenase/MMP-1 by interrelated loops
of interleukin-1 and interleukin-6. Photochem Photobiol 59:5506
www.jidonline.org 1907
F Liebel et al.
Visible Light Induces ROS, Inflammatory Cytokines, and MMPs in Skin
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Reactive oxygen species are considered to play an important role in cutaneous pathology. Enzymic and non-enzymic antioxidants can prevent oxidative damage but may be overcome by strong pro-oxidative stimuli. The acute effect of a single exposure to near ultraviolet (UVA)/visible radiation (greater than 320 nm) on various skin antioxidants was examined in hairless mice immediately after irradiation. Impairment of cutaneous catalase and glutathione reductase activity was observed. Superoxide dismutase and glutathione peroxidase were not significantly influenced. Inhibition of catalase may render skin more susceptible to the damaging effects of hydrogen peroxide and its reaction products such as the hydroxyl radical. Partially diminished glutathione reductase activity is not accompanied by a change in reduced/oxidized glutathione level immediately after irradiation. There was a tendential (not statistically significant) decrease in cutaneous tocopherol, ubiquinol + ubiquinone 9 and ascorbic acid levels, either indicating direct photodestruction or consumption by reaction products of photooxidative stress. This partial impairment of the cutaneous antioxidant defense system by near ultraviolet/visible light, showing that the most susceptible component in skin is catalase, suggests possible pharmacological interventions.
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Human skin exhibited a stable electron spin resonance (ESR) signal before irradiation, only if the samples were pigmented. The single almost symmetrical absorption peak with g = 2.003 and a line width of 5–6 gauss at 77 °K is associated with melanin. It was also observed in pigmented human and guinea pig hair, melanosomes, and melanin granules obtained from B-16 mouse melanoma and DOPA-melanin. Irradiation with wavelengths greater than 320 mμ, including ruby laser (λ = 694 mμ) and incoherent 600–700 mμ light, at 77 °K enhanced the melanin signals in pigmented samples only. “White” skin gave no intrinsic melanin free-radical signal, and irradiation with similar wavelengths produced no detectable free radicals. Ultraviolet radiation (λ < 320 mμ) enhanced the melanin signal in pigmented samples and also produced other radicals with a line width of approximately 24 gauss in both “white” and pigmented skin. Radiation-induced free radicals were only detected at 77 °K and were unstable at 300°K. The photo-enhanced melanin free-radical signal was also unstable, but the intrinsic melanin signal was not destroyed by warming. Radical yields measured in “white” and pigmented skin as a function of ultraviolet exposure dose showed that fewer radicals were generated in pigmented skin. The significance of this observation in relation to the photoprotective function of melanin is discussed.
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The precise role of src-type kinases as signal transducers has been under intensive investigation but only in a few instances has their role been revealed in any detail. Thus, src, fyn and yes are activated upon stimulation by platelet-derived growth factor or colony-stimulating factor in cells expressing high levels of these receptors. Activation of src-family kinases by other receptor tyrosine kinases such as the epidermal-growth-factor (EGF) receptor has not been directly demonstrated. In this report, we demonstrate EGF-dependent activation of src-family tyrosine kinases in NIH3T3 cells overexpressing the human EGF receptor. Activation is rapid (< 1 min) and persistent (up to 16 h). Furthermore, we show a correlation between the level of EGF receptor expressed and the degree of src-family kinase activation. We show that src-family kinase activity is also activated by addition of EGF to PC12 cells, which endogenously express relatively high levels of EGF receptor. Most strikingly, we show that A431 cells, which endogenously express very high levels of EGF receptor, show 10-fold elevated src-family kinase activity as compared to DHER14 cells, and that this activity is constitutive. This activity is completely blocked by AG1478, a specific inhibitor of the EGF-receptor tyrosine kinase activity, pointing to a direct link between overexpression of the EGF receptor and enhanced src-family kinase activity. Our findings suggest that EGF-dependent src-family kinase activity is detectable only when the levels of EGF receptor reach a specific level. Additionally, high levels of EGF receptor, as in A431 cells, may contribute to the elevated activation of src-family kinases. Sustained src-family kinase activation, similar to that seen in v-src-transformed cells, may play a role in tumorogenesis and tumor maintenance.