Experimental Dermatology 2006: 15: 678–684 Copyright
The Authors 2006.

Journal compilation
2006 Blackwell Munksgaard

Blackwell Munksgaard . Printed in Singapore
doi: 10.1111/j.0906-6705.2006.00466.x
Experimental Dermatology
ISSN 0906-6705

Review Article

Botanical antioxidants in the prevention

of photocarcinogenesis and photoaging
Afaq F, Mukhtar H. Botanical antioxidants in the prevention of pho- Farrukh Afaq and Hasan
tocarcinogenesis and photoaging. Mukhtar
Exp Dermatol 2006: 15: 678–684.
The Authors 2006. Journal
Department of Dermatology, University of
compilation
2006 Blackwell Munksgaard.
Wisconsin, Madison, WI, USA
Abstract: Exposure of the skin to ultraviolet (UV) radiation, partic-
ularly its UV-B component (280–320 nm), from the sun results in ery-
thema, edema, hyperplasia, hyperpigmentation, sunburn cells,
immunosuppression, photoaging, and skin cancer. Amongst these var-
ious adverse effects of UV-B radiation, skin cancer and photoaging Key words: botanical antioxidant – photoaging
are of great concern. More recent changes in lifestyle have led to a sig- – photocarcinogenesis – photochemopreven-
nificant increase in the amount of UV-B radiation people receive lead- tion – ultraviolet radiation
ing to a surge in the incidence of skin cancer and photoaging. As Hasan Mukhtar PhD
these trends are likely to continue in the foreseeable future, the Helfaer Professor of Cancer Research
adverse effect of UV-B has become a major human health concern. Director and Vice Chair for Research
Therefore, development of novel strategies to reduce the occurrence of Department of Dermatology
skin cancer and delay the process of photoaging are highly desirable University of Wisconsin,
Medical Sciences Center
goals. One approach to reduce their occurrence is through photochem- Room #B25
oprevention, which we define as the use of agents capable of amelior- 1300 University Avenue
ating the adverse effects of UV-B on the skin. Photochemoprevention Madison
via use of botanical antioxidants, present in the common diet of WI 53706
human have gained considerable attention as photochemopreventive USA
agents for human use. Many such agents have also found a place in Tel.: +1-608-263-3927
skin care products. This review will focus on the effects of selected Fax: +1-608-263-5223
botanical antioxidants in the prevention of photocarcinogenesis and e-mail: hmukhtar@wisc.edu
photoaging. Accepted for publication 9 June 2006

Introduction
Solar ultraviolet (UV) radiation is the most
prominent and ubiquitous physical carcinogen
in our natural environment and is divided into
three regions depending on the wavelength,
short-wave UV-C (200–280 nm), mid-wave UV-
B (280–320 nm), and long-wave UV-A (320–
400 nm). UV-C in solar radiation is blocked by
the ozone layer and therefore its role in human
pathogenesis is minimal. UV-B and, to a much
lesser extent, UV-A radiation are responsible
for inducting various skin disorders including
skin cancer and photoaging (1,2). The role of
UV-B in skin cancer is well established. Because
greater than 90% of the solar radiation reach-
ing the earth’s surface is UV-A, in recent years
the role of UV-A in skin carcinogenesis has
begun to be appreciated (1). It has become clear
that UV-A accounts for at least 10% of the car-
cinogenic dose of the sunlight. UV irradiation
to skin results in erythema, edema, sunburn
cells, hyperplasia, immunosuppression, photo-
aging and photocarcinogenesis (3). UV-B irradi-
ation to skin has direct effects on biomolecules;
for example, the formation of cyclobutane pyr-
imidine dimers (CPDs) and pyrimidine (6-4)
pyrimidone photodimers (4), photoisomeriza-
tion of trans- to cis-urocanic acid (5) and gen-
eration of reactive oxygen species (ROS) (6).
These effects of UV-B may result in a variety of
skin disorders including skin cancer.
In the USA alone 1.2 million new cases of
skin cancer are identified each year and this
accounts for 40% of all new cancer cases that

678

are diagnosed (1,7). Similar trend is also noted
in natives of many countries with predomin-
antly Caucasian population. The non-melanoma
skin cancers (NMSCs), comprising of basal cell
carcinomas (BCCs) and squamous cell carcino-
mas (SCCs), are the most frequently diagnosed
cutaneous malignancies and account for
approximately 80% and 16% of all skin can-
cers, respectively, whereas malignant melan-
omas account for only 4% of all skin cancers.
Both BCCs and SCCs are derived from the
basal layer of the epidermis of the skin. SCCs
are invasive and approximately 10% of these
cancers metastasize. On the other hand, BCCs
do not metastasize but can be locally invasive
and destructive. In the year 2006, 62 190 newly
diagnosed cases of melanoma, resulting in 7910
deaths are expected to occur while 2800 deaths
are expected to occur from non-epithelial skin
cancers (7).
Depending on the amount and form of the
UV-B radiation, as well as on the skin type of
the individual exposed, UV-B irradiation causes
premature skin aging, referred as photoaging (8).
Solar UV radiation has a profound effect on
exposed skin, producing accelerated aging chan-
ges consisting of fine and coarse wrinkling,
rough skin texture, dryness, telangiectasia, and
dyspigmentation abnormalities including lenti-
gines as well as guttate hypermelanosis and
hypomelanosis (9). Studies have suggested that
there is an increased generation of ROS in skin
upon UV-B and UV-A exposure. Increased ROS
generation can overwhelm antioxidant defense
mechanisms, resulting in oxidative stress and
oxidative photodamage of proteins and other
macromolecules in the skin. These ROS are
believed to be critical mediators of the photo-
aging and photocarcinogenesis processes
(1,3,10). ROS can modify proteins in tissue to
form carbonyl derivatives, which accumulate in
the papillary dermis of photodamaged skin (11).
Exposure of human or mouse skin to UV irradi-
ation results in the induction of a series of matrix
metalloproteinases (MMPs), which have been
implicated in photoaging (12). Together these
proteinases are capable of degrading the collagen
framework and other components of skin con-
nective tissue. Recent studies have suggested that
solar UV reduces collagen in photoaged human
skin by blocking transforming growth factor-
beta type II receptor/Smad signaling (13). UV
radiation results in the activation of activator
protein 1 (AP-1) that stimulates transcription of
MMP genes encoding MMP-1 (collagenase),
MMP-9 (gelantinase), and MMP-3 (stromelysin-
Prevention of photocarcinogenesis and photoaging
1) in skin cells (12). These changes apparently
occur through induction of AP-1 that is activa-
ted by a series of mitogen-activated protein kin-
ases (MAPKs). In addition, nuclear factor
kappa B (NF-jB) a transcription factor is acti-
vated by UV irradiation, which stimulates neu-
trophil attraction bringing neutrophil
collagenase (MMP-8) into the irradiation site to
further aggravate matrix degradation. Both AP-I
and NF-jB are activated by ROS that may pro-
vide the common denominator for driving this
complex biologic interaction (14,15). Oxidative
stress can also increase elastin messenger RNA
levels in dermal fibroblasts providing a mechan-
ism for the elastotic changes found in photoaged
dermis (15).
Botanical antioxidants have been shown to
be associated with reduced incidence of ROS-
mediated photocarcinogenesis and photoaging
(3,6). Consistent with this understanding in
recent years, botanical antioxidants have attrac-
ted considerable attention because of their skin
photoprotective effects (3,6). This has generated
a great interest in using dietary botanical sup-
plements rich in antioxidants for the prevention
of photocarcinogenesis and photoaging. This
review focuses on photochemopreventive effects
of selected botanical antioxidants. The biologi-
cal effects and postulated mechanism(s) of these
botanical antioxidants are listed in Table 1 and
Fig. 1.
Green tea
Tea from the plant Camellia sinensis, of the
Theaceae family, is consumed by more than
two-thirds of the world’s population and is the
most popular beverage, next only to water.
Green tea contains four major types of poly-
phenols: ())-epicatechin (EC), ())-epicatechin
gallate (ECG), ())-epigallocatechin (EGC), and
())-epigallocatechin-3-gallate (EGCG). All of
these polyphenols act as potent antioxidants
and can scavenge ROS, such as lipid-free radi-
cals, superoxide radicals, hydroxyl radicals,
hydrogen peroxide, and singlet oxygen. EGCG,
which is the most abundant polyphenol present
in green tea has been considered to be the main
compound responsible for these effects, consti-
tuting approximately 40% of the total polyphe-
nolic mixture. In several publications from this
laboratory, it has been demonstrated that top-
ical application or oral feeding of a polyphenol-
ic fraction prepared from green tea (GTP)
prevents photocarcinogenesis (3,6,16). Long-
term oral feeding of GTP to mice exposed
679
Afaq and Mukhtar
Table 1. Botanical antioxidants for prevention of photocarcinogenesis
and photogaing
Botanical Target/mechanism(s)
antioxidants
Green, black Inhibits UV-B-induced skin
and oolong teas carcinogenesis
Inhibits UV-B-induced erythema,
edema, depletion of the antioxidant
enzyme system, ODC and COX-2
activities
Enhances UV-B-induced increases
in the number of p53- and
p21-positive cells in the epidermis
Reduces the size of the parametrial
fat pad and the thickness of the
dermal fat layer
Inhibits UV-B-induced expression of
the MMPs
Reduces UV-B-induced collagen
cross-linking
Reduces IL-10 and IL-12 production
Inhibits UV-B-induced release of
intracellular H2O2 and
phosphorylation of MAPKs
and NF-jB pathways
Enhances the rate and extent of
disappearance of the mutant
p53-positive patches
Pomegranate fruit Inhibits UV-B-induced
extract phosphorylation of MAPKs
and activation of NF-jB pathways
Inhibits phosphorylation of STAT3,
AKT, and ERK1/2
Inhibits UV-B-induced edema,
hyperplasia, infiltration of
leukocytes, generation of
hydrogen peroxide and DNA
damage in the form of CPDs
and 8-hydroxy-2¢-deoxyguanosine
Enhances UV-B-induced increases
in p53- and p21 protein expression
Resveratrol Inhibits UV-B-induced skin
inflammation and edema
Inhibits UV-B-induced activation
of NF-jB
Modulates cki–cyclin–cdk network
and MAPKs pathway
Inhibits tumor incidence and delays
the onset of tumorigenesis
Inhibits cell proliferation and
phosphorylation of survivin
Genistein Inhibits UV-A-induced enhancement
of the DNA-binding activity of
STAT1 by acting as a TPK inhibitor
Reduces c-fos and c-jun expression
Reduces edema and contact
hypersensitivity
Inhibits skin carcinogenesis and
cutaneous aging
chronically to UV radiation resulted in lower
tumor burden in these animals compared with
their non-GTP-fed control (3,6,16). Oral feeding
of GTP to SKH-1 hairless mice, followed by
irradiation with UV-B, resulted in significant
protection against UV-B radiation-mediated
cutaneous edema; depletion of the antioxidant-
defense system in the epidermis; induction of
680
UVB
Reference Skin
Botanical Antioxidants Act As
st
(3,6,16–18) Sunscreen (1 line) Antioxidant (2nd line) Redox regulation of
signal transduction
Radical scavenger
pathway (3rd line)
(3,6,16,36) Prevention of
Reduces oxidative stress
damage
Suppresses inflammation
Reduces DNA damage
p53 Suppresses proliferation
(19) Gene Transformation
DNA excission repair specific apoptosis in
inactivation
transformed cells
(20) Correction
Prevention
(26) Figure 1. Three modes of action of botanical antioxidants.
(30)
epidermal ornithine decarboxylase (ODC) and
(3,16)
(31,32,35) cycloxygenase (COX) enzymes activities (3,16).
Oral administration of green tea, black tea, and
decaffeinated green and black teas inhibited the
(22,23) formation and size of malignant and non-malig-
nant tumors (17). Studies have shown that oral
consumption of green tea to mice with estab-
(40)
lished tumors resulted in a significant regression
of these tumors (18). Oral feeding of GTP to
(41) SKH-1 hairless mice enhanced UV-B-induced
(42) increases in epidermal wild type p53, p21, and
apoptotic sunburn cells in the epidermis (19).
Lu et al. (20) have shown that orally adminis-
tered green tea, black tea, and caffeine
decreased the size of the parametrial fat pad,
(42) and the thickness of the dermal fat layer both
(43) distant and directly under tumors. UV-B-
induced apoptosis in mouse skin is largely but
(44) not entirely dependent on wild-type p53 and
(45) Bax, and topical applications of caffeine
enhanced UV-B-induced apoptosis by p53- and
(46) Bax-independent mechanisms (21). Oral treat-
(47) ment of mice with caffeine or green tea during
chronic UV-B-irradiation changed the mutation
(51) profile of the p53 gene in early mutant p53-pos-
itive epidermal patches (22). Oral administra-
(52) tion of green tea as the sole source of drinking
(53) fluid or topical applications of caffeine immedi-
(49) ately after discontinuation of UV-B treatment
enhanced the rate and extent of disappearance
of the mutant p53-positive patches (23). Kra-
mata et al. (24) have demonstrated that the p53
mutation profile of UV-B-induced skin patches
and SCCs was very similar suggesting that these
patches are precursor lesions for SCCs.
Treatment of cultured cells with EGCG
directly inhibits the expression of MMPs such
as MMP-2, MMP-9, and neutrophil elastase
even in the absence of UV exposure (25). Chro-
nic exposure of mouse skin to UV-B has been

shown to induce the expression of MMP-2, -3,
-7, and -9, which are involved in the degrada-
tion of types-I and -III collagen fragments gen-
erated by collagenases (26), and type IV
collagen of the basement membrane (27). Oral
feeding of GTP to mouse as the sole
source in drinking water remarkably inhibited
UV-induced expression of these MMPs in
mouse skin suggesting that GTP has a potential
anti-photoaging effect (26). Treatment of
EGCG diminishes UV-A-induced skin damage
(roughness and sagginess) and protects from the
loss of dermal collagen in hairless mouse skin,
and also blocks the UV-induced increase of col-
lagen secretion and collagenase mRNA level in
cultured human epidermal fibroblasts and the
promoter-binding activities of AP-l and NF-jB
(28). In another study, feeding of green tea
extract to rats remarkably inhibited the age-
associated increase of the fluorescence in the
aortic collagen (29). Oxidative damage induced
by UV-B may also cause modifications of pro-
teins, e.g. collagen cross-linking and formation
of carbonyl derivatives. In a study with mice, it
was found that collagen cross-linking could be
reduced by green tea extract (30). EGCG topic-
ally applied to mice, before exposure to a single
dose of UV-B irradiation, inhibits hydrogen
peroxide and nitric oxide production both in
the dermis and epidermis (3,16). Topical appli-
cation of EGCG to the mouse skin prior to UV
irradiation was shown to result in the inhibition
of contact hypersensitivity response to contact
sensitizer, reduction in the number of infiltra-
ting macrophages (CD11b+ cells) and neu-
trophils, downregulation in UV-induced
production of Interleukin-10 (IL-10) and
increased production of IL-12 in the skin and
draining lymph nodes (16). EGCG was also
found to balance the alterations in the IL-10/
IL-12 cytokines and this may be mediated by
the antigen presenting cells in the skin and
draining lymph nodes or by blocking the infil-
tration of IL-10 secreting CD11b+ macro-
phages into the irradiated site (16).
Pretreatment of normal human epidermal
keratinocytes (NHEK) with EGCG inhibits
UV-B-induced oxidative stress-mediated phos-
phorylation of MAPK; phosphorylation and
degradation of IjBa and activation of IKKa
and NF-jB (31,32). In human keratinocyte cell
line HaCaT, EGCG inhibits UV-B-mediated
activation of AP-1 activity (33). EGCG was
also found to inhibit UV-B-induced expression
of c-fos, a major component of AP-1 (34). Top-
ical application of GTP to SKH-1 hairless mice
Prevention of photocarcinogenesis and photoaging
prior to multiple UV-B exposure down-regu-
lates UV-B-induced phosphorylation of MAPK
and activation of NF-jB (35). It has also been
demonstrated that topical application of green
tea extract to human skin prior to UV-B expo-
sure inhibited UV-B-mediated erythema,
reduced the number of sunburn cells, protected
epidermal Langherhans cells and reduced DNA
damage (36). In a study including 118 patients
with atopic dermatitis, the consumption of
three cups of Oolong tea (a combination of
green and red tea) for 6 months decreased the
severity of the disease (37). In another study,
topical application of GTP before UV-B radi-
ation was found to inhibit UV-B-induced CPDs
formation in a dose-dependent fashion (38).
Pomegranate
Pomegranate (Punica granatum, Punicaceae) is
a rich source of two types of polyphenolic com-
pounds; anthocyanidins (such as delphinidin,
cyanidin, and pelargonidin) and hydrolyzable
tannins (such as punicalin, pedunculagin, puni-
calagin, gallagic, and ellagic acid esters of
glucose) and possesses strong antioxidant and
anti-inflammatory properties (39). Our recent
study demonstrated that treatment of NHEK
with pomegranate fruit extract (PFE) inhibited
UV-B-mediated phosphorylation of ERKl/2,
JNK1/2, and p38 proteins in a dose- and time-
dependent manner. We also found that PFE
treatment of NHEK resulted in a dose- and
time-dependent inhibition of UV-B-mediated
degradation and phosphorylation of IjBa and
activation of IKKa. Employing immunoblot
analysis, enzyme linked immunosorbent assay
and electrophoretic mobility shift assay; we
found that PFE treatment to NHEK resulted in
a dose- and time-dependent inhibition of
UV-B-mediated nuclear translocation and phos-
phorylation of NF-jB/p65 at Ser536 (40). Taken
together, our data show that PFE protects
against the adverse effects of UV-B radiation by
inhibiting UV-B-induced modulations of NF-jB
and MAPKs pathways and provides a molecu-
lar basis for the photochemopreventive effects
of PFE. We have shown that treatment of
NHEK with PFE before exposure to UV-A
results in a dose-dependent inhibition of UV-A-
mediated phosphorylation of signal transducers
and activators of transcription-3 (STAT3) at
Tyr705, AKT at Ser473, and ERK1/2. We found
that UV-A radiation of NHEK resulted in the
phosphorylation of mammalian target of rapa-
mycin (mTOR) at Thr2448 and p70S6K at
681
Afaq and Mukhtar
Thr421/Ser424. PFE pretreatment resulted in a
dose-dependent inhibition in the phosphoryla-
tion of mTOR at Thr2448 and p70S6K at
Thr421/Ser424. PFE pretreatment was also found
to induce UV-A-mediated activation in the pro-
tein expression of Bax and Bad (proapoptotic
protein) and downregulation of Bcl-XL (anti-
apoptotic protein) (41). Recently, we have
demonstrated that oral feeding of PFE to SKH-
1 hairless mice resulted in the inhibition of
UV-B-induced increase in skin edema, hyper-
plasia, infiltration of leukocytes, generation of
hydrogen peroxide and DNA damage in the
form of CPDs and 8-hydroxy-2¢-deoxyguano-
sine (42). Furthermore, we found that oral feed-
ing of PFE resulted in the inhibition of
UV-B-induced PCNA, ODC, and COX-2 pro-
tein expression, and the induction of UV-B-
mediated increase in p21 and p53 protein
expression (42). The relevance of these findings
in cell culture system to photocarcinogenesis
and photoaging is being examined in our labor-
atory.
Resveratrol
Resveratrol (trans-3,4¢,5-trihydroxystilbene) is a
polyphenolic phytoalexin found largely in the
skin and seeds of grapes, nuts, fruits, and red
wine. Resveratrol is a potent antioxidant with
anti-inflammatory and antiproliferative proper-
ties (43,44). Topical application of resveratrol to
SKH-1 hairless mice prior to UV-B irradiation
significantly inhibited UV-B-induced skin edema
and caused a significant decrease in UV-B-medi-
ated generation of hydrogen peroxide and infil-
tration of leukocytes (43). In another study,
pretreatment of NHEK with resveratrol inhib-
ited UV-B-mediated activation of NF-jB path-
way (44). Studies have demonstrated that
resveratrol imparts its protective effect against
multiple UV-B exposure via modulations in the
cki–cyclin–cdk network and MAPK pathway
(45). In long-term studies, topical application of
skin with resveratrol (both pre- and post-treat-
ment) has been shown to result in a significant
inhibition in tumor incidence and delay in the
onset of tumorigenesis (46). Further, in short-
term experiments, topical application of resvera-
trol prior to UV-B irradiation resulted in a
significant inhibition of UV-B-induced cellular
proliferation, protein, and messenger RNA
levels of survivin, and phosphorylation of survi-
vin in SKH-1 hairless mouse skin (47). The
effect of resveratrol on photoaging remains to be
examined.
682
Genistein
Soybeans are a rich source of flavonoids called
isoflavones, the most potent being genistein and
daidzein. Genistein (4¢,5,7-trihydroxyiso-flav-
one) is primarily present in soy, in ginkgo
biloba extract, Greek oregano, and Greek sage.
Genistein has been shown to possess antioxid-
ant and anti-carcinogenic effects in skin (48,49).
Cai and Wei (50) have shown that dietary
administration of genistein significantly
enhanced the activities of antioxidant enzymes
in the skin of SENCAR mice. Treatment of
human keratinocyte cell line NCTC 2544 with
genistein prevented UV-induced enhancement
of the DNA-binding activity of signal trans-
ducer and activator of transcription-1 by acting
as a tyrosine kinase inhibitor, thus limiting lipid
peroxidation and increases in ROS formation
(51). Studies have shown that topical applica-
tion of genistein before UV-B radiation reduced
c-fos and c-jun expression in the SENCAR
mouse skin in a dose-dependent manner (52).
Genistein also had a powerful potential to
reduce the inflammatory edema reaction and
suppressed contact hypersensitivity induced by
moderate doses of solar-simulated UV radiation
(53). Genistein substantially inhibits skin carcin-
ogenesis and cutaneous aging induced by UV
light in mice and photodamage in humans (49).
A recent study has demonstrated that genistein
potently minimizes the detrimental effects of
UV-B irradiation in human reconstituted skin
by preserving cutaneous proliferation and repair
mechanics, inhibition of oxidative, and photo-
dynamic damage to DNA (54).
Conclusions
Accumulating laboratory data indicate that
many botanical agents, with antioxidant proper-
ties exerts anti-inflammatory, cancer-preventive
and anti-photoaging effects in the skin. This
suggests the possibility that specific botanicals
might be used for the prevention and treatment
of a variety of human skin disorders. The use
of skin care products supplemented with several
effective agents working through different path-
ways in conjunction with the use of sunscreens
may be an effective approach for reducing
UV-B-generated ROS-mediated photoaging and
skin cancer in humans. Because UV-B radiation
plays an important role in cutaneous damage,
agent(s) that can protect against UV-B-radi-
ation could be ideal photochemoprotective
agent(s) for photoaging and skin cancer.

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