ABSTRACT Pomegranate seed oil was investigated for possible skin cancer chemopreventive efficacy in mice. In the main
experiment, two groups consisting each of 30, 4–5-week-old, female CD1 mice were used. Both groups had skin cancer initiated
with an initial topical exposure of 7,12-dimethylbenzanthracene and with biweekly promotion using 12-O-tetradecanoylphorbol
13-acetate (TPA). The experimental group was pretreated with 5% pomegranate seed oil prior to each TPA application.
Tumor incidence, the number of mice containing at least one tumor, was 100% and 93%, and multiplicity, the
average number of tumors per mouse, was 20.8 and 16.3 per mouse after 20 weeks of promotion in the control and pomegranate
seed oil-treated groups, respectively (P , .05). In a second experiment, two groups each consisting of three CD1 mice
were used to assess the effect of pomegranate seed oil on TPA-stimulated ornithine decarboxylase (ODC) activity, an important
event in skin cancer promotion. Each group received a single topical application of TPA, with the experimental group
receiving a topical treatment 1 h prior with 5% pomegranate seed oil. The mice were killed 5 h later, and ODC activity was
assessed by radiometric method. The experimental group showed a 17% reduction in ODC activity. Pomegrante seed oil (5%)
significantly decreased (P , .05) tumor incidence, multiplicity, and TPA-induced ODC activity.
INTRODUCTION
SKIN CANCER is the most common type of cancer in the
United States,1 with more than a million reported cases2
and 9,000 deaths per year.3 Increasing incidence of these
cancers due to constant exposure of skin to environmental
carcinogens, including both chemical agents and ultraviolet
radiation, provides a strong basis for chemoprevention with
both synthetic and natural, and internal and topical, remedies.
4 Further, skin cancer chemoprevention is a useful
model for cancer chemoprevention in general.5
Chemical and UVB radiation-induced skin carcinogenesis
in murine skin and possibly human skin is a stepwise
process of at least three distinct stages: initiation, promotion,
and progression. Experimental initiation in vivo is accomplished
by the topical application of a single dose of
a skin carcinogen such as 7,12-dimethylbenzanthracene
(DMBA), and is essentially irreversible. However, an initiation
dose of carcinogen may not produce visible tumors,
resulting only following prolonged and repeated application
of a tumor promoter such as 12-O-tetradecanoylphorbol 13-
acetate (TPA) to initiated skin.6,7 Promoters like TPA induce
ornithine decarboxylase (ODC), the rate-limiting enzyme
in the synthesis of polyamines8 and an important
molecular target for skin cancer chemoprevention.9 Other
targets may also involve promotion, or initiation or progression
events in the multistage process of neoplastic development.
Our previous work has highlighted the efficacy of topically
applied natural products derived from onion and garlic
oils,10 and more recently sandalwood oil11,12 and its constituent,
13 in preventing skin tumors in CD1 and SENCAR
mice. In the present work, we bring this experience to bear
on the study of pomegranate seed oil as a potential skin cancer
chemopreventive product.
Pomegranate fruit (Punica granatum) has been used
worldwide as an item of diet and medicine for millenia, and
has also been regarded as an important symbol in world religions
and mythologies and of medicine itself.14 We previously
demonstrated potent antioxidant and prostaglandininhibitory
activities for polyphenols extracted from pomegranate
seed oil and pomegranate fermented juice,15 as well
as a wide range of human breast cancer suppressive properties
in vitro, including promotion of apoptosis and inhibi-
tion of proliferation and invasion by the seed oil, and inhibition
of DMBA-initiated carcinogenesis in a mouse mammary
organ culture (MMOC) by the fermented juice
polyphenols.16 We recently showed chemopreventive activity
of the whole seed oil in the MMOC to be even stronger,
weight per weight, than that of the purified fermented juice
polyphenols.17
Pomegranate seed oil consists of .80% conjugated fatty
acids, the most important of which is the octadecatrienoic
acid, punicic acid. Punicic acid, like the ,1% polyphenols
in pomegranate seed oil, is an inhibitor of prostaglandin
biosynthesis.18 Punicic acid is also cytotoxic to mouse
leukemia cells, possibility related to inhibition of lipid peroxidation.
19 Pomegranate is one of only about a half dozen
plants known to contain conjugated fatty acids. A possible
relationship between the relative botanical isolation of
pomegranate and its singular chemistry and anticancer properties
has been noted.20
The purpose of the present investigation was to study
the chemopreventive effects of pomegranate seed oil on
DMBA-initiated and TPA-promoted skin tumor development
during the initiation and promotion phases in CD1
mice. Further, the effects of pomegranate seed oil on weight
gain and ODC activity in the experimental animals were also
evaluated.
MATERIALS AND METHODS
Pomegranate seed oil
Pomegranate seed oil was provided by Rimonest Ltd. (Rimonest
Ltd., Haifa, Israel; www.rimonest.com) from pomegranates
of the “Wonderful” cultivar, organically grown at
Kibbutz Sde Eliahu, Israel, in the year 2000. Seeds were
separated from their juice sacs, washed in water, and dried
in a convection current solar dryer. Oil extrusion was by
“cold press” at 80°C, using a Type 40A electric screw press
(Skeppsta Maskin, Orebro, Sweden). The oil was assayed
by an independent laboratory (Mylnfield Research Services,
Invergowrie, Dundee, Scotland) and shown to contain not
less than 80% conjugated fatty acids as triglycerols, diglycerols,
and monoglycerols.
Tumorogenesis protocol
The skin cancer protocol of Dwivedi et al.13 was used. In
brief, 4–6-week-old CD1 mice were divided into two groups,
each group containing 30 mice, as indicated in Table 1. The
mice were kept in an environmentally controlled room with
temperature, humidity, and light regulated. The backs of the
mice were shaved carefully with an electric clipper to avoid
cuts. The mice were allowed to rest for 2 days before carcinogenesis
was initiated.
Carcinogenesis was initiated with DMBA (200 nmol in
100 mL of acetone) applied topically. One week later, carcinogenesis
was promoted with TPA (5 nmol in 100 mL of
acetone), applied topically twice weekly. TPA treatment
continued throughout the duration of the experiment (20
weeks). Mice in group 1 served as the control and were pretreated
topically with 100 mL of acetone 1 h prior to each
TPA application. Mice in group 2 were pretreated topically
with 100 mL of 5% pomegranate seed oil in acetone 1 h
prior to each TPA application. Tumor counts and group
weights were taken on a weekly basis. Tumor incidence and
multiplicity were calculated and analyzed statistically.
ODC assay
Mice were divided into two groups, each containing three
mice. The backs of the mice were shaved carefully with an
electric clipper to avoid cuts. Mice in group 1 received 100
mL of acetone before TPA (5 nmol in 100 mL of acetone)
treatment topically. Mice in group 2 received 100 mL of 5%
pomegranate seed oil in acetone, before topical TPA (5 nmol
in 100 mL of acetone) treatment.
Mice were killed 5 h after the topical applications of TPA.
The dorsal epidermis was removed and homogenized in
phosphate buffer (pH 7.2) containing 0.1 mM pyridoxal
phosphate and 0.1 mM EDTA. The homogenate was centrifuged
at 105,000 g for 90 min and the supernatant collected
and used for the ODC assay. The assay mixture in
the main part of a Warburg flask was composed of 40 mL
of phosphate buffer (pH 7.2), 25 mL of pyridoxal phosphate,
25 mL of dithiothreitol, 25 mL of EDTA, 10 mL of L-ornithine
containing 0.5 mCi of DL[1-14C]ornithine, and 200
mL of epidermal supernatant.
The center well of the Warburg flask contained 400 mL
of ethanolamine and methoxyethanol used to absorb the
14CO2 produced in the main compartment. After incubation
at 37°C for 1 h, the reaction was stopped by the addition of
500 mL of citric acid. The mixture was stored in a dark place
overnight to ensure complete absorption of 14CO2 in the center
well. The contents of the center well were transferred to
a scintillation vial. The center well was washed with 0.5 mL
of ethanol four times, and the wash also added to the scintillation
vial, along with 10 mL of scintillation fluor. Radioactivity
was counted with a Beckman LS6000SE liquid
scintillation counter. The disintegrations per minute were
quantified. Assessment of ODC activity was accomplished
by measuring the production of 14CO2 from DL-[1-14C]ornithine.
Protein assay
Protein was assayed in the supernatant with a Bio-Rad
Protein Assay Kit. A standard curve was obtained using
bovine serum albumin. Absorbance values at 595 nm were
determined using the spectrophotometer. Protein concentra-
tions of the supernatant were extrapolated from the standard
curve data.
Statistical analysis
The INSTAT software (GraphPad, San Diego, CA,
U.S.A.) was used for the data analysis. x2 was used for the
comparison of papilloma incidence and Student’s t test for
tumor multiplicity and ODC activity. Significance was considered
at P , .05.
RESULTS
The effects of pomegranate seed oil treatment on the incidence
of skin tumors in CD1 mice are shown in Fig. 1.
Skin tumors appeared in the sixth week of promotion after
the initial DMBA application in the control and treated
groups. Pomegranate seed oil treatment did not delay the appearance
of tumors, but significantly decreased (P , .05)
the rate at which the tumors developed. Skin tumor incidence
after 20 weeks of promotion was 100% and 93% for
the control and 5% pomegranate seed oil-treated groups, respectively.
The effects of pomegranate seed oil treatment on tumor
multiplicity in CD1 mice are shown in Fig. 2. Pomegranate
seed oil treatment significantly decreased (P , .05) the tumor
multiplicity throughout the 20 weeks of promotion. The
mean number of tumors per mouse was 20.8 and 16.3 for
the control and 5% pomegranate seed oil-treated groups, respectively.
Topical application of 5% pomegranate seed oil also significantly
inhibited (P , .05) TPA-induced epidermal ODC
activity. Fig. 3 illustrates the effects of pomegranate seed
oil treatment on TPA-induced epidermal ODC activity. The
ODC activity was 18.49 and 14.84 nmol of 14CO2/mg/h in
the control and 5% pomegranate seed oil-treated groups, respectively.
The pomegranate seed oil group has significantly
(P , .05) decreased ODC activity. Topical application of
5% pomegranate seed oil alone did not induce any epidermal
ODC activity. Topical application of 5% pomegranate
seed oil also did not have any effect on weight gain, as indicated
in Fig. 4.
CONCLUSIONS
Pomegranate seed oil (5%) topical applications significantly
decreased the incidence of skin tumor development,
skin tumor multiplicity, and ornithine decarboxylase activity
during 20 weeks of promotion. It is thus likely that the
inhibition of ornithine decarboxylase by the pomegranate
seed oil was at least partially responsible for the chemopreventive
effect.
As noted, pomegranate seed oil is very rich in punicic
acid, a known inhibitor of prostaglandin biosynthesis,
specifically by inhibiting cyclooxygenase (Cox 1 and Cox
2) and lipoxygenase.21 Pomegranate seed oil also inhibits the
upstream eicosanoid enzyme, phospholipase A2, expressed
by human prostate cancer cells.22 That prostaglandins at very
low concentrations promote ornithine decarboxylase23 suggests
that the inhibition of prostaglandin biosynthesis by
pomegranate seed oil might also contribute to its inhibition
of ornithine decarboxylase and, ultimately, to inhibition of
skin cancer promotion.
Overall, pomegranate seed oil appears to be a benign natural
product with potential as a topical chemopreventive
agent against skin cancer. More in-depth investigations, including
clinical studies, are warranted to evaluate this hypothesis
further.
ACKNOWLEDGMENTS
The authors wish to thank Mr. Eli Merom of Kibbutz Sde
Eliahu, Israel, for supplying the organically grown pomegranates
used in this study. Thanks also to Alexander
Botvinnik for technical assistance in the preparation of the
manuscript.
REFERENCES
1. Skin cancer (what you need to know about). National Institutes
of Heath, National Cancer Institute, NIH Publication no. 94-1563
(Revised April 1993).
2. Facts on Skin Cancer. American Cancer Society Publication 99-
200M. Rev. 8/95, no. 2049.
3. Cancer Facts & Figures—1996. American Cancer Society Publication
96-300M, no. 5008-96, p. 17.
4. Gupta S, Mukhtar H: Chemoprevention of skin cancer: current
status and future prospects. Cancer Metastasis Rev 2002;21:
363–380.
5. Richmond E, Viner JL: Chemoprevention of skin cancer. Semin
Oncol Nurs 2003;19:62–69.
6. Agarwal R, Mukhtar H: Cutaneous chemical carcinogens. In:
Pharmacology of the Skin (Mukhtar H, ed.). CRC Press, Boca Raton,
FL, 1991, pp. 371–387.
7. Boutwell RK: Some biological aspects of skin carcinogenesis.
Prog Exp Tumor Res 1984:4:207–250.
8. O’Brien TG, Simsiman RC, Boutwell RK. Induction of the
polyamine-biosynthetic enzymes in mouse epidermis by tumorpromoting
agents. Cancer Res 1975;35:1662–1670.
9. Stratton SP, Dorr RT, Alberts DS. The state-of-the-art in chemoprevention
of skin cancer. Eur J Cancer 2000;36:1292–1297.
10. Dwivedi C, Rohlfs S, Jarvis D, Engineer FN. Chemoprevention
of chemically induced skin tumor development by diallyl sulfide
and diallyl disulfide. Pharm Res 1992;9:1668–1670.
11. Dwivedi C, Abu-Ghazaleh A. Chemopreventive effects of sandalwood
oil on skin papillomas in mice. Eur J Cancer Prev 1997;
6:399–401.
12. Dwivedi C, Zhang Y: Sandalwood oil prevents skin tumour development
in CD1 mice. Eur J Cancer Prev 1999;8:449–455.
13. Dwivedi C, Guan X, Harmsen WL, Voss AL, Goetz-Parten D, Koopman
EM, Johnson KM, Valluri HB, Matthees DD: Chemopreventive
effects of a-santalol on skin tumor development in CD1 and
Sencar mice. Cancer Epidemiol Biomarkers Prev 2003;12:151–156.
14. Langley P. Why a pomegranate? BMJ 2000;321:1153–1154.
15. Schubert SY, Lansky EP, Neeman I. Antioxidant and eicosanoid
enzyme inhibition properties of pomegranate seed oil and fermented
juice flavonoids. J Ethnopharmacol 1999;66:11–17.
16. Kim ND, Mehta R, Yu W, Neeman I, Livney T, Amichay A,
Poirier D, Nicholls P, Kirby A, Jiang W, Mansel R, Ramachandran
C, Rabi T, Kaplan B, Lansky E. Chemopreventive and adjuvant
therapeutic potential of pomegranate (Punica granatum)
for human breast cancer. Breast Cancer Res Treat 2002;71:
203–217.
17. Unpublished data presented at the 13th Annual Meeting of the
North American Menopause Society, Chicago 2002. Rajendra
Mehta, Department of Surgical Oncology, University of Illinois
at Chicago.
18. Nugteren DH, Christ-Hazelhof E. Naturally occurring conjugated
octadecatrienoic acids are strong inhibitors of prostaglandin
biosynthesis. Prostaglandins 1987;33:403–417.
19. Suzuki R, Noguchi R, Ota T, Abe M, Miyashita K, Kawada T: Cytotoxic
effect of conjugated trienoic fatty acids on mouse tumor and
human monocytic leukemia cells. Lipids 2001;36:477–482.
20. Longtin R: The pomegranate: nature’s power fruit? J Natl Cancer
Inst 2003;95:346–348.
21. Unpublished data, Robert Newman, Department of Pharmacology,
MD Anderson Cancer Center, Houston, TX.
22. Unpublished data, Wenguo Jiang, Department of Surgery, Cardiff
University, U.K.
23. Cameron CM, Rillema JA: Effects of prostaglandins on the prolactin
stimulation of lipid biosynthesis in mouse mammary gland
explants. Prostaglandins Leukot Med 1983;10:433–441.
POMEGRANATE SEED OIL AND SKIN CANCER IN MICE 161
experiment, two groups consisting each of 30, 4–5-week-old, female CD1 mice were used. Both groups had skin cancer initiated
with an initial topical exposure of 7,12-dimethylbenzanthracene and with biweekly promotion using 12-O-tetradecanoylphorbol
13-acetate (TPA). The experimental group was pretreated with 5% pomegranate seed oil prior to each TPA application.
Tumor incidence, the number of mice containing at least one tumor, was 100% and 93%, and multiplicity, the
average number of tumors per mouse, was 20.8 and 16.3 per mouse after 20 weeks of promotion in the control and pomegranate
seed oil-treated groups, respectively (P , .05). In a second experiment, two groups each consisting of three CD1 mice
were used to assess the effect of pomegranate seed oil on TPA-stimulated ornithine decarboxylase (ODC) activity, an important
event in skin cancer promotion. Each group received a single topical application of TPA, with the experimental group
receiving a topical treatment 1 h prior with 5% pomegranate seed oil. The mice were killed 5 h later, and ODC activity was
assessed by radiometric method. The experimental group showed a 17% reduction in ODC activity. Pomegrante seed oil (5%)
significantly decreased (P , .05) tumor incidence, multiplicity, and TPA-induced ODC activity.
INTRODUCTION
SKIN CANCER is the most common type of cancer in the
United States,1 with more than a million reported cases2
and 9,000 deaths per year.3 Increasing incidence of these
cancers due to constant exposure of skin to environmental
carcinogens, including both chemical agents and ultraviolet
radiation, provides a strong basis for chemoprevention with
both synthetic and natural, and internal and topical, remedies.
4 Further, skin cancer chemoprevention is a useful
model for cancer chemoprevention in general.5
Chemical and UVB radiation-induced skin carcinogenesis
in murine skin and possibly human skin is a stepwise
process of at least three distinct stages: initiation, promotion,
and progression. Experimental initiation in vivo is accomplished
by the topical application of a single dose of
a skin carcinogen such as 7,12-dimethylbenzanthracene
(DMBA), and is essentially irreversible. However, an initiation
dose of carcinogen may not produce visible tumors,
resulting only following prolonged and repeated application
of a tumor promoter such as 12-O-tetradecanoylphorbol 13-
acetate (TPA) to initiated skin.6,7 Promoters like TPA induce
ornithine decarboxylase (ODC), the rate-limiting enzyme
in the synthesis of polyamines8 and an important
molecular target for skin cancer chemoprevention.9 Other
targets may also involve promotion, or initiation or progression
events in the multistage process of neoplastic development.
Our previous work has highlighted the efficacy of topically
applied natural products derived from onion and garlic
oils,10 and more recently sandalwood oil11,12 and its constituent,
13 in preventing skin tumors in CD1 and SENCAR
mice. In the present work, we bring this experience to bear
on the study of pomegranate seed oil as a potential skin cancer
chemopreventive product.
Pomegranate fruit (Punica granatum) has been used
worldwide as an item of diet and medicine for millenia, and
has also been regarded as an important symbol in world religions
and mythologies and of medicine itself.14 We previously
demonstrated potent antioxidant and prostaglandininhibitory
activities for polyphenols extracted from pomegranate
seed oil and pomegranate fermented juice,15 as well
as a wide range of human breast cancer suppressive properties
in vitro, including promotion of apoptosis and inhibi-
tion of proliferation and invasion by the seed oil, and inhibition
of DMBA-initiated carcinogenesis in a mouse mammary
organ culture (MMOC) by the fermented juice
polyphenols.16 We recently showed chemopreventive activity
of the whole seed oil in the MMOC to be even stronger,
weight per weight, than that of the purified fermented juice
polyphenols.17
Pomegranate seed oil consists of .80% conjugated fatty
acids, the most important of which is the octadecatrienoic
acid, punicic acid. Punicic acid, like the ,1% polyphenols
in pomegranate seed oil, is an inhibitor of prostaglandin
biosynthesis.18 Punicic acid is also cytotoxic to mouse
leukemia cells, possibility related to inhibition of lipid peroxidation.
19 Pomegranate is one of only about a half dozen
plants known to contain conjugated fatty acids. A possible
relationship between the relative botanical isolation of
pomegranate and its singular chemistry and anticancer properties
has been noted.20
The purpose of the present investigation was to study
the chemopreventive effects of pomegranate seed oil on
DMBA-initiated and TPA-promoted skin tumor development
during the initiation and promotion phases in CD1
mice. Further, the effects of pomegranate seed oil on weight
gain and ODC activity in the experimental animals were also
evaluated.
MATERIALS AND METHODS
Pomegranate seed oil
Pomegranate seed oil was provided by Rimonest Ltd. (Rimonest
Ltd., Haifa, Israel; www.rimonest.com) from pomegranates
of the “Wonderful” cultivar, organically grown at
Kibbutz Sde Eliahu, Israel, in the year 2000. Seeds were
separated from their juice sacs, washed in water, and dried
in a convection current solar dryer. Oil extrusion was by
“cold press” at 80°C, using a Type 40A electric screw press
(Skeppsta Maskin, Orebro, Sweden). The oil was assayed
by an independent laboratory (Mylnfield Research Services,
Invergowrie, Dundee, Scotland) and shown to contain not
less than 80% conjugated fatty acids as triglycerols, diglycerols,
and monoglycerols.
Tumorogenesis protocol
The skin cancer protocol of Dwivedi et al.13 was used. In
brief, 4–6-week-old CD1 mice were divided into two groups,
each group containing 30 mice, as indicated in Table 1. The
mice were kept in an environmentally controlled room with
temperature, humidity, and light regulated. The backs of the
mice were shaved carefully with an electric clipper to avoid
cuts. The mice were allowed to rest for 2 days before carcinogenesis
was initiated.
Carcinogenesis was initiated with DMBA (200 nmol in
100 mL of acetone) applied topically. One week later, carcinogenesis
was promoted with TPA (5 nmol in 100 mL of
acetone), applied topically twice weekly. TPA treatment
continued throughout the duration of the experiment (20
weeks). Mice in group 1 served as the control and were pretreated
topically with 100 mL of acetone 1 h prior to each
TPA application. Mice in group 2 were pretreated topically
with 100 mL of 5% pomegranate seed oil in acetone 1 h
prior to each TPA application. Tumor counts and group
weights were taken on a weekly basis. Tumor incidence and
multiplicity were calculated and analyzed statistically.
ODC assay
Mice were divided into two groups, each containing three
mice. The backs of the mice were shaved carefully with an
electric clipper to avoid cuts. Mice in group 1 received 100
mL of acetone before TPA (5 nmol in 100 mL of acetone)
treatment topically. Mice in group 2 received 100 mL of 5%
pomegranate seed oil in acetone, before topical TPA (5 nmol
in 100 mL of acetone) treatment.
Mice were killed 5 h after the topical applications of TPA.
The dorsal epidermis was removed and homogenized in
phosphate buffer (pH 7.2) containing 0.1 mM pyridoxal
phosphate and 0.1 mM EDTA. The homogenate was centrifuged
at 105,000 g for 90 min and the supernatant collected
and used for the ODC assay. The assay mixture in
the main part of a Warburg flask was composed of 40 mL
of phosphate buffer (pH 7.2), 25 mL of pyridoxal phosphate,
25 mL of dithiothreitol, 25 mL of EDTA, 10 mL of L-ornithine
containing 0.5 mCi of DL[1-14C]ornithine, and 200
mL of epidermal supernatant.
The center well of the Warburg flask contained 400 mL
of ethanolamine and methoxyethanol used to absorb the
14CO2 produced in the main compartment. After incubation
at 37°C for 1 h, the reaction was stopped by the addition of
500 mL of citric acid. The mixture was stored in a dark place
overnight to ensure complete absorption of 14CO2 in the center
well. The contents of the center well were transferred to
a scintillation vial. The center well was washed with 0.5 mL
of ethanol four times, and the wash also added to the scintillation
vial, along with 10 mL of scintillation fluor. Radioactivity
was counted with a Beckman LS6000SE liquid
scintillation counter. The disintegrations per minute were
quantified. Assessment of ODC activity was accomplished
by measuring the production of 14CO2 from DL-[1-14C]ornithine.
Protein assay
Protein was assayed in the supernatant with a Bio-Rad
Protein Assay Kit. A standard curve was obtained using
bovine serum albumin. Absorbance values at 595 nm were
determined using the spectrophotometer. Protein concentra-
tions of the supernatant were extrapolated from the standard
curve data.
Statistical analysis
The INSTAT software (GraphPad, San Diego, CA,
U.S.A.) was used for the data analysis. x2 was used for the
comparison of papilloma incidence and Student’s t test for
tumor multiplicity and ODC activity. Significance was considered
at P , .05.
RESULTS
The effects of pomegranate seed oil treatment on the incidence
of skin tumors in CD1 mice are shown in Fig. 1.
Skin tumors appeared in the sixth week of promotion after
the initial DMBA application in the control and treated
groups. Pomegranate seed oil treatment did not delay the appearance
of tumors, but significantly decreased (P , .05)
the rate at which the tumors developed. Skin tumor incidence
after 20 weeks of promotion was 100% and 93% for
the control and 5% pomegranate seed oil-treated groups, respectively.
The effects of pomegranate seed oil treatment on tumor
multiplicity in CD1 mice are shown in Fig. 2. Pomegranate
seed oil treatment significantly decreased (P , .05) the tumor
multiplicity throughout the 20 weeks of promotion. The
mean number of tumors per mouse was 20.8 and 16.3 for
the control and 5% pomegranate seed oil-treated groups, respectively.
Topical application of 5% pomegranate seed oil also significantly
inhibited (P , .05) TPA-induced epidermal ODC
activity. Fig. 3 illustrates the effects of pomegranate seed
oil treatment on TPA-induced epidermal ODC activity. The
ODC activity was 18.49 and 14.84 nmol of 14CO2/mg/h in
the control and 5% pomegranate seed oil-treated groups, respectively.
The pomegranate seed oil group has significantly
(P , .05) decreased ODC activity. Topical application of
5% pomegranate seed oil alone did not induce any epidermal
ODC activity. Topical application of 5% pomegranate
seed oil also did not have any effect on weight gain, as indicated
in Fig. 4.
CONCLUSIONS
Pomegranate seed oil (5%) topical applications significantly
decreased the incidence of skin tumor development,
skin tumor multiplicity, and ornithine decarboxylase activity
during 20 weeks of promotion. It is thus likely that the
inhibition of ornithine decarboxylase by the pomegranate
seed oil was at least partially responsible for the chemopreventive
effect.
As noted, pomegranate seed oil is very rich in punicic
acid, a known inhibitor of prostaglandin biosynthesis,
specifically by inhibiting cyclooxygenase (Cox 1 and Cox
2) and lipoxygenase.21 Pomegranate seed oil also inhibits the
upstream eicosanoid enzyme, phospholipase A2, expressed
by human prostate cancer cells.22 That prostaglandins at very
low concentrations promote ornithine decarboxylase23 suggests
that the inhibition of prostaglandin biosynthesis by
pomegranate seed oil might also contribute to its inhibition
of ornithine decarboxylase and, ultimately, to inhibition of
skin cancer promotion.
Overall, pomegranate seed oil appears to be a benign natural
product with potential as a topical chemopreventive
agent against skin cancer. More in-depth investigations, including
clinical studies, are warranted to evaluate this hypothesis
further.
ACKNOWLEDGMENTS
The authors wish to thank Mr. Eli Merom of Kibbutz Sde
Eliahu, Israel, for supplying the organically grown pomegranates
used in this study. Thanks also to Alexander
Botvinnik for technical assistance in the preparation of the
manuscript.
REFERENCES
1. Skin cancer (what you need to know about). National Institutes
of Heath, National Cancer Institute, NIH Publication no. 94-1563
(Revised April 1993).
2. Facts on Skin Cancer. American Cancer Society Publication 99-
200M. Rev. 8/95, no. 2049.
3. Cancer Facts & Figures—1996. American Cancer Society Publication
96-300M, no. 5008-96, p. 17.
4. Gupta S, Mukhtar H: Chemoprevention of skin cancer: current
status and future prospects. Cancer Metastasis Rev 2002;21:
363–380.
5. Richmond E, Viner JL: Chemoprevention of skin cancer. Semin
Oncol Nurs 2003;19:62–69.
6. Agarwal R, Mukhtar H: Cutaneous chemical carcinogens. In:
Pharmacology of the Skin (Mukhtar H, ed.). CRC Press, Boca Raton,
FL, 1991, pp. 371–387.
7. Boutwell RK: Some biological aspects of skin carcinogenesis.
Prog Exp Tumor Res 1984:4:207–250.
8. O’Brien TG, Simsiman RC, Boutwell RK. Induction of the
polyamine-biosynthetic enzymes in mouse epidermis by tumorpromoting
agents. Cancer Res 1975;35:1662–1670.
9. Stratton SP, Dorr RT, Alberts DS. The state-of-the-art in chemoprevention
of skin cancer. Eur J Cancer 2000;36:1292–1297.
10. Dwivedi C, Rohlfs S, Jarvis D, Engineer FN. Chemoprevention
of chemically induced skin tumor development by diallyl sulfide
and diallyl disulfide. Pharm Res 1992;9:1668–1670.
11. Dwivedi C, Abu-Ghazaleh A. Chemopreventive effects of sandalwood
oil on skin papillomas in mice. Eur J Cancer Prev 1997;
6:399–401.
12. Dwivedi C, Zhang Y: Sandalwood oil prevents skin tumour development
in CD1 mice. Eur J Cancer Prev 1999;8:449–455.
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jagger - am Donnerstag, 10. Juni 2004, 17:17 - Rubrik: Wirksamkeit Granatapfel
