InhibitoryEffectofCurcumin,ChlorogenicAcid,CaffeicAcid...
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[CANCER RESEARCH 48, 5941-5946, November 1, 1988] Inhibitory Effect of Curcumin, Chlorogenic Acid, Caffeic Acid, and Ferulic Acid on Tumor Promotion in Mouse Skin by 12-O-Tetradecanoylphorbol-13-acetate Mou-Tuan Huang, Robert C. Smart, Ching-Quo Wong, and Allan H. Conney Department of Chemical Biology and Pharmacognosy, College of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, New Jersey 08855-0789 [M-T. H., A. H. C.]; Roche Research Center, Hoffmann-La Roche Inc., Nutley, New Jersey 07110 [M-T. H., R. C. S., C-Q. W., A. H. C.]; and Toxicology Program, North Carolina State University, Raleigh, North Carolina 27695-7633 [R. C. SJ ABSTRACT The effects of topically applied curcumin, chlorogenic acid, caffeic acid, and ferulic acid on 12-O-tetradecanoylphorbol-13-acetate (TPA)- induced epidermal ornithine decarboxylase activity, epidermal DNA syn thesis, and the promotion of skin tumors were evaluated in female CD-I mice. Topical application of 0.5, 1, 3, or 10 iano\ of curcumin inhibited by 31, 46, 84, or 98%, respectively, the induction of epidermal ornithine decarboxylase activity by 5 nmol of TPA. In an additional study, the topical application of 10 ¿tinol of curcumin, chlorogenic acid, caffeic acid, or ferulic acid inhibited by 91, 25,42, or 46%, respectively, the induction of ornithine decarboxylase activity by 5 nmol of TPA. The topical application of 10 iimol of curcumin together with 2 or 5 nmol of TPA inhibited the TPA-dependent stimulation of the incorporation of |'I l|- thymidine into epidermal DNA by 49 or 29%, respectively, whereas lower doses of curcumin had little or no effect. Chlorogenic acid, caffeic acid, and ferulic acid were less effective than curcumin as inhibitors of the TPA-dependent stimulation of DNA synthesis. Topical application of 1, 3, or 10 fimol of curcumin together with 5 nmol of TPA twice weekly for 20 weeks to mice previously initiated with 7,12-dimethylbenz[a]anthra- cene inhibited the number of TPA-induced tumors per mouse by 39, 77, or 98%, respectively. Similar treatment of mice with 10 nmol of chloro genic acid, caffeic acid, or ferulic acid together with 5 nmol of TPA inhibited the number of TPA-induced tumors per mouse by 60, 28, or 35%, respectively, and higher doses of the phenolic acids caused a more pronounced inhibition of tumor promotion. The possibility that curcumin could inhibit the action of arachidonic acid was evaluated by studying the effect of curcumin on arachidonic acid-induced edema of mouse ears. The topical application of 3 or 10 ¿tmol of curcumin 30 min before the application of 1 /imol of arachidonic acid inhibited arachidonic acid- induced edema by 33 or 80%, respectively. INTRODUCTION The ground dried rhizome of the plant Curcuma longa Linn has been used for centuries as a naturally occurring medicine for the treatment of inflammatory and other diseases (1,2), and it has also been used as a coloring agent and spice in many foods. The powdered rhizome is commonly called turmeric. Curcumin (diferuloylmethane; Fig. 1) has been identified as the major pigment in turmeric, and this substance has also been used as a spice, food preservative, and yellow coloring agent (3-5). Curcumin, which is widely used in curry, is responsible for the yellow color of curry. Turmeric contains 1-5% curcumin while the curcumin content of turmeric oleoresin is approxi mately 40% (6). Recent studies have indicated that several compounds that possess antioxidant or antiinflammatory activity inhibit TPA1- induced tumors on mouse skin (7-12). Since curcumin has been reported to possess both antioxidant and antiinflammatory activity (13-17), we have evaluated the effect of curcumin on TPA-induced tumor promotion on mouse skin, and we have Received 5/6/88; revised 7/22/88; accepted 7/29/88. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 'The abbreviations used are: TPA, 12-O-tetradecanoylphorbol-13-acetate; DMBA, 7,12-dimethylbenz[a]anthracene; DMSO, dimethyl sulfoxide; HETE, hydroxyeicosatetraenoic acid; HPETE, hydroperoxyeicosatetraenoic acid. also evaluated the effects of the related compounds chlorogenic acid, caffeic acid, and ferulic acid as potential inhibitors of tumor promotion. MATERIALS AND METHODS Materials. TPA was purchased from CRC Inc., Chanhassen, MN. DL-['"C]Ornithine (58 Ci/mmol) and [3H]thymidine (5 Ci/mmol) were purchased from Amersham Corp., Arlington Heights, IL. fra/w-Reti- noic acid was obtained from Hoffmann-La Roche Inc., Basle, Switzer land. Fluocinolone acetonide, nonradioactive DL-ornithine, curcumin, chlorogenic acid, ferulic acid, and caffeic acid were purchased from the Sigma Chemical Co., St. Louis, MO, and DMBA was purchased from Calbiochem-Behringer, San Diego, CA. Aquasol was purchased from New England Nuclear, Boston, MA. Acetone and dimethyl sulfoxide were purchased from Burdick & Jackson Laboratories, Muskegon, MI. Arachidonic acid was purchased from Nu-Chek Prep Inc., Elysian, MN. Animals. Seven-week-old female CD-I mice were purchased from Charles River Laboratories, Kingston, NY, and kept in our animal facility at least 1 week before use. Mice were fed a Purina Laboratory Chow 5001 diet ad libitum (Ralston-Purina Co., St. Louis, MO) and kept on a 12-h light, 12-h dark cycle. Mice were provided drinking water ad libitum. The dorsal region of each mouse was shaved with electric clippers at least 2 days before treatment with TPA or DMBA. Only mice that did not show signs of hair regrowth were used. All compounds were applied to the dorsal shaved area of 8- to 9-week-old mice in 200 ß\ acetone or 200 ^1 of acetone:DMSO (90:10). Ornithine Decarboxylase Assay and Preparation of Epidermal I lo- mogenates. The preparation of epidermal homogenate and the deter mination of ornithine decarboxylase activity were done as described previously (18). Mice were treated topically with 200 /il of acetone, with TPA in acetone, or with curcumin and TPA together in acetone. In experiments with chlorogenic acid, caffeic acid, and ferulic acid, mice were treated with 200 n\ of acetone:DMSO (90:10), with TPA in acetone:DMSO, or with the test compound and TPA together in acetone:DMSO. Five h after treatment, the mice were sacrificed by cervical dislocation, and the dorsal area of the skin was removed. In order to remove the epidermis from the dermis, the skins were plunged into a 58°Cwater bath for 30 s, and then the skins were immediately submerged in an ice water bath as described by Slaga et al. (19). The epidermis was removed from the dermis by gentle scraping and placed in 1 ml of 50 mM potassium phosphate, pH 7.7, buffer containing 2 HIMdithiothreitol and 0.1 mM EDTA. The epidermis was homogenized on ice for 30 s with a Tekmar polytron tissumizer at full setting. The epidermal homogenate was centrifuged at 11,000 x g for 30 min at 4°C and the supernatant fraction was removed and stored overnight at —20°C prior to the determination of ornithine decarboxylase activity as described earlier (18). Protein was determined by the method of Lowry et a/., using bovine serum albumin as the standard (20). Quantitation of Epidermal DNA Synthesis. Mice were treated topi cally with 200 ii\ acetone, TPA in acetone, or curcumin and TPA together in acetone. Acetone:DMSO (90:10) was the solvent for exper iments with chlorogenic acid, caffeic acid, and ferulic acid. At 18 h after treatment, 10 ^1 of [3H]thymidine solution (5 Ci/mmol, 1 mCi/ ml) were injected i.p. into each mouse, and 40 min later the mice were sacrificed by cervical dislocation. The isolation of epidermal DNA and determination of the incorporation of [3H]thymidine into DNA was done as previously described (18). Tumor Studies on Mouse Skin. The dorsal region of 7-week-old 5941 on May 28, 2018. © 1988 American Association for Cancer Research. cancerres.aacrjournals.org Downloaded from
Transcript of InhibitoryEffectofCurcumin,ChlorogenicAcid,CaffeicAcid...
[CANCER RESEARCH 48, 5941-5946, November 1, 1988]
Inhibitory Effect of Curcumin, Chlorogenic Acid, Caffeic Acid, and Ferulic Acid onTumor Promotion in Mouse Skin by 12-O-Tetradecanoylphorbol-13-acetate
Mou-Tuan Huang, Robert C. Smart, Ching-Quo Wong, and Allan H. Conney
Department of Chemical Biology and Pharmacognosy, College of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, New Jersey 08855-0789[M-T. H., A. H. C.]; Roche Research Center, Hoffmann-La Roche Inc., Nutley, New Jersey 07110 [M-T. H., R. C. S., C-Q. W., A. H. C.]; and Toxicology Program,North Carolina State University, Raleigh, North Carolina 27695-7633 [R. C. SJ
ABSTRACT
The effects of topically applied curcumin, chlorogenic acid, caffeicacid, and ferulic acid on 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced epidermal ornithine decarboxylase activity, epidermal DNA synthesis, and the promotion of skin tumors were evaluated in female CD-Imice. Topical application of 0.5, 1, 3, or 10 iano\ of curcumin inhibitedby 31, 46, 84, or 98%, respectively, the induction of epidermal ornithinedecarboxylase activity by 5 nmol of TPA. In an additional study, thetopical application of 10 ¿tinolof curcumin, chlorogenic acid, caffeic acid,or ferulic acid inhibited by 91, 25,42, or 46%, respectively, the inductionof ornithine decarboxylase activity by 5 nmol of TPA. The topicalapplication of 10 iimol of curcumin together with 2 or 5 nmol of TPAinhibited the TPA-dependent stimulation of the incorporation of |'I l|-
thymidine into epidermal DNA by 49 or 29%, respectively, whereas lowerdoses of curcumin had little or no effect. Chlorogenic acid, caffeic acid,and ferulic acid were less effective than curcumin as inhibitors of theTPA-dependent stimulation of DNA synthesis. Topical application of 1,3, or 10 fimol of curcumin together with 5 nmol of TPA twice weekly for20 weeks to mice previously initiated with 7,12-dimethylbenz[a]anthra-cene inhibited the number of TPA-induced tumors per mouse by 39, 77,or 98%, respectively. Similar treatment of mice with 10 nmol of chlorogenic acid, caffeic acid, or ferulic acid together with 5 nmol of TPAinhibited the number of TPA-induced tumors per mouse by 60, 28, or35%, respectively, and higher doses of the phenolic acids caused a morepronounced inhibition of tumor promotion. The possibility that curcumincould inhibit the action of arachidonic acid was evaluated by studying theeffect of curcumin on arachidonic acid-induced edema of mouse ears. Thetopical application of 3 or 10 ¿tmolof curcumin 30 min before theapplication of 1 /imol of arachidonic acid inhibited arachidonic acid-induced edema by 33 or 80%, respectively.
INTRODUCTION
The ground dried rhizome of the plant Curcuma longa Linnhas been used for centuries as a naturally occurring medicinefor the treatment of inflammatory and other diseases (1,2), andit has also been used as a coloring agent and spice in manyfoods. The powdered rhizome is commonly called turmeric.Curcumin (diferuloylmethane; Fig. 1) has been identified as themajor pigment in turmeric, and this substance has also beenused as a spice, food preservative, and yellow coloring agent(3-5). Curcumin, which is widely used in curry, is responsiblefor the yellow color of curry. Turmeric contains 1-5% curcumin
while the curcumin content of turmeric oleoresin is approximately 40% (6).
Recent studies have indicated that several compounds thatpossess antioxidant or antiinflammatory activity inhibit TPA1-induced tumors on mouse skin (7-12). Since curcumin has beenreported to possess both antioxidant and antiinflammatoryactivity (13-17), we have evaluated the effect of curcumin onTPA-induced tumor promotion on mouse skin, and we have
Received 5/6/88; revised 7/22/88; accepted 7/29/88.The costs of publication of this article were defrayed in part by the payment
of page charges. This article must therefore be hereby marked advertisement inaccordance with 18 U.S.C. Section 1734 solely to indicate this fact.
'The abbreviations used are: TPA, 12-O-tetradecanoylphorbol-13-acetate;DMBA, 7,12-dimethylbenz[a]anthracene; DMSO, dimethyl sulfoxide; HETE,hydroxyeicosatetraenoic acid; HPETE, hydroperoxyeicosatetraenoic acid.
also evaluated the effects of the related compounds chlorogenicacid, caffeic acid, and ferulic acid as potential inhibitors oftumor promotion.
MATERIALS AND METHODS
Materials. TPA was purchased from CRC Inc., Chanhassen, MN.DL-['"C]Ornithine (58 Ci/mmol) and [3H]thymidine (5 Ci/mmol) werepurchased from Amersham Corp., Arlington Heights, IL. fra/w-Reti-noic acid was obtained from Hoffmann-La Roche Inc., Basle, Switzerland. Fluocinolone acetonide, nonradioactive DL-ornithine, curcumin,chlorogenic acid, ferulic acid, and caffeic acid were purchased from theSigma Chemical Co., St. Louis, MO, and DMBA was purchased fromCalbiochem-Behringer, San Diego, CA. Aquasol was purchased fromNew England Nuclear, Boston, MA. Acetone and dimethyl sulfoxidewere purchased from Burdick & Jackson Laboratories, Muskegon, MI.Arachidonic acid was purchased from Nu-Chek Prep Inc., Elysian, MN.
Animals. Seven-week-old female CD-I mice were purchased fromCharles River Laboratories, Kingston, NY, and kept in our animalfacility at least 1 week before use. Mice were fed a Purina LaboratoryChow 5001 diet ad libitum (Ralston-Purina Co., St. Louis, MO) andkept on a 12-h light, 12-h dark cycle. Mice were provided drinkingwater ad libitum. The dorsal region of each mouse was shaved withelectric clippers at least 2 days before treatment with TPA or DMBA.Only mice that did not show signs of hair regrowth were used. Allcompounds were applied to the dorsal shaved area of 8- to 9-week-oldmice in 200 ß\acetone or 200 ^1 of acetone:DMSO (90:10).
Ornithine Decarboxylase Assay and Preparation of Epidermal I lo-mogenates. The preparation of epidermal homogenate and the determination of ornithine decarboxylase activity were done as describedpreviously (18). Mice were treated topically with 200 /il of acetone,with TPA in acetone, or with curcumin and TPA together in acetone.In experiments with chlorogenic acid, caffeic acid, and ferulic acid,mice were treated with 200 n\ of acetone:DMSO (90:10), with TPA inacetone:DMSO, or with the test compound and TPA together inacetone:DMSO. Five h after treatment, the mice were sacrificed bycervical dislocation, and the dorsal area of the skin was removed. Inorder to remove the epidermis from the dermis, the skins were plungedinto a 58°Cwater bath for 30 s, and then the skins were immediately
submerged in an ice water bath as described by Slaga et al. (19). Theepidermis was removed from the dermis by gentle scraping and placedin 1 ml of 50 mM potassium phosphate, pH 7.7, buffer containing 2HIMdithiothreitol and 0.1 mM EDTA. The epidermis was homogenizedon ice for 30 s with a Tekmar polytron tissumizer at full setting. Theepidermal homogenate was centrifuged at 11,000 x g for 30 min at 4°C
and the supernatant fraction was removed and stored overnight at—20°Cprior to the determination of ornithine decarboxylase activity
as described earlier (18). Protein was determined by the method ofLowry et a/., using bovine serum albumin as the standard (20).
Quantitation of Epidermal DNA Synthesis. Mice were treated topically with 200 ii\ acetone, TPA in acetone, or curcumin and TPAtogether in acetone. Acetone:DMSO (90:10) was the solvent for experiments with chlorogenic acid, caffeic acid, and ferulic acid. At 18 hafter treatment, 10 ^1 of [3H]thymidine solution (5 Ci/mmol, 1 mCi/
ml) were injected i.p. into each mouse, and 40 min later the mice weresacrificed by cervical dislocation. The isolation of epidermal DNA anddetermination of the incorporation of [3H]thymidine into DNA was
done as previously described (18).Tumor Studies on Mouse Skin. The dorsal region of 7-week-old
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INHIBITION OF TPA-INDUCED TUMOR PROMOTION
CH
HO
O OII II
CH= CH-C-CH2-C-CH = CH
CURCUMIN
HOOC-CH= CH
FERULIC ACID
HOOC-CH=CH
CAFFEIC ACID
H OOC-CH = C
HOOCJ 1 H
CHLOROGENICACID
Fig. 1. Structures of curcumin, ferulic acid, cafTeic acid, and chlorogenic acid.
female CD-I mice was shaved with an electric clipper. Two days latergroups of 30 mice were treated topically with 200 nmol DMBA in 200til acetone, and control mice received 200 ill of acetone alone. After 1week, mice were treated topically with 200 til of acetone, 5 nmol TPA,or 5 nmol TPA applied simultaneously with curcumin in 200 ^1 ofacetone twice weekly for 20 weeks. In another tumor study designed toevaluate the effect of chlorogenic acid, cafTeic acid, and ferulic acid onTPA-induced tumor formation, acetone:DMSO (90:10) was used asthe solvent because chlorogenic acid at the doses examined was notcompletely soluble in acetone. Tumors of at least 1 mm in diameterwere counted and recorded once every 2 weeks, and the results areexpressed as the average number of tumors per mouse and percentageof tumor-bearing mice. Control mice that were first initiated with 200nmol DMBA and then treated with 3 //mol curcumin in acetone oracetone alone twice weekly for 20 weeks also were examined. Likewise,control mice treated with acetone alone in place of DMBA and thentreated twice weekly with 5 nmol TPA in 200 M!acetone for 20 weekswere also included in the study.
RESULTS
Effect of Curcumin, Chlorogenic Acid, Caffeic Acid, and Ferulic Acid on TPA-induced Ornithine Decarboxylase Activity inMouse Epidermis. The effect of topically applied curcumin,chlorogenic acid, caffeic acid, or ferulic acid on TPA-induced
ornithine decarboxylase activity in mouse epidermis was examined and the results are shown in Table 1. Curcumin stronglyinhibited the TPA-induced increase in epidermal ornithine decarboxylase activity in a dose-dependent manner. Topical application of 0.5, 1, 3, or 10 ¿¿molof curcumin with 5 nmol ofTPA inhibited the TPA-induced increases in epidermal ornithine decarboxylase activity by 31,46, 84, or 98%, respectively.Application of ira/w-retinoic acid (1.3 nmol), a known inhibitorof TPA-induced ornithine decarboxylase activity (21), inhibitedthe TPA-induced increase in epidermal ornithine decarboxylaseactivity by 88%. The effects of several naturally occurringphenolic acids that are structurally related to curcumin werestudied (Table 1, experiment 2). Topical application of 10 ¿tmolof chlorogenic acid, caffeic acid, or ferulic acid inhibited TPA-induced increases in ornithine decarboxylase activity by 25,42,or 46%, respectively, and higher doses of the phenolic acidsresulted in greater inhibition. The results of these studies indi
cate that curcumin is about 10-fold more active than the phenolic acids. The results of an additional study indicated that ani.p. injection of curcumin could inhibit the induction of ornithine decarboxylase activity in mouse epidermis that occurredafter the topical application of TPA. The i.p. injection of 40 or120 /¿molof curcumin (dissolved in dimethyl sulfoxide) intomice l h before the topical application of 5 nmol TPA inhibitedthe induction of epidermal ornithine decarboxylase activity by46 or 86%, respectively (data not shown). During the course ofthis study, it was found that curcumin in the vehicle precipitatedout of solution in the abdominal cavity and thus may not havebeen completely bioavailable. Curcumin could be seen as yellowflecks in the abdominal cavity throughout the experimentalperiod of 6 h.
Effect of Curcumin, Chlorogenic Acid, Caffeic Acid, and Ferulic Acid on TPA-induced DNA Synthesis in Mouse Epidermis.Topical application of 1, 3, or 10 nmo\ of curcumin togetherwith 2 nmol of TPA inhibited the TPA-induced increase inDNA synthesis by 4, 14, or 49%, respectively (Table 2, experiment 1). The topical application of fluocinolone acetonide (2nmol), a known inhibitor of TPA-induced DNA synthesis,inhibited the TPA-induced increase by 86%. Topical application of 1, 3, or 10 ¿¿molof curcumin simultaneously with 5nmol of TPA inhibited the TPA-induced increase in the incorporation of [3H]thymidine into epidermal DNA by 3,4, or 29%,
respectively (Table 2, experiment 2). Topical application of 20/¿molof chlorogenic acid, caffeic acid, or ferulic acid togetherwith 5 nmol of TPA inhibited the TPA-induced increase in theincorporation of [3H]thymidine into epidermal DNA by 7, 33,
or 15%, respectively.Effect of Curcumin, Chlorogenic Acid, Caffeic Acid, and Fer
ulic Acid on TPA-induced Tumor Promotion in Mouse Epidermis. CD-I mice initiated with 200 nmol of DMBA and promoted with 5 nmol of TPA twice weekly for 20 weeks developedan average of 16.4 tumors/mouse. Topical application of 1, 3,or 10 ttmol of curcumin with 5 nmol of TPA twice a week for20 weeks inhibited the number of skin tumors per mouse by39,77, or 98%, respectively, and the percentage of animals withtumors was decreased by 21, 66, or 82%, respectively (Fig. 2).Additional groups of mice were initiated with DMBA and thentreated with acetone or 3 /¿molof curcumin twice weekly for 20weeks. None of these animals developed tumors, indicating thatcurcumin was not a tumor promoter. In a second experiment,mice that were initiated with 200 nmol of DMBA and promotedwith 5 nmol of TPA twice weekly for 19 weeks developed anaverage of 6.2 tumors/mouse. In this experiment, topical application of 10 Mmolof curcumin, chlorogenic acid, caffeic acid,or ferulic acid together with 5 nmol of TPA twice a week for19 weeks inhibited the number of tumors per mouse by 100,60,28, or 35%, respectively, and the percentage of animals withtumors was inhibited 100, 31, 28, or 7%, respectively (Table3). The topical application of 20 /¿molchlorogenic acid, 20/¿molcaffeic acid, or 50 jumol ferulic acid inhibited the averagenumber of tumors per mouse by 91, 82, or 97%, respectively,and the percentage of animals with tumors was inhibited 61,43, or 80%, respectively (Table 3).
Effect of Curcumin on TPA- and Arachidonic Acid-inducedEdema of Mouse Ears. The possibility that curcumin couldinhibit TPA- and arachidonic acid-dependent inflammation wasevaluated by studying the effects of curcumin on TPA- andarachidonic acid-induced edema of mouse ears. The applicationof 0.4 or 1 /¿molof curcumin to an ear of the mouse 30 minbefore 0.5 nmol of TPA inhibited TPA-induced edema by 79or 97%, respectively (Table 4, experiment 1). In additional
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INHIBITION OF TPA-INDUCED TUMOR PROMOTION
Table I Effects ofcurcumin, chlorogenic acid, caffeic add, andferulic acid on TPA-induced ornithine decarboxylase activity in mouse epidermis
TPA (S nmol) was applied topically alone or simultaneously with various doses of potential inhibitor in 200 »Iof acetone (experiment 1) or in 200 «Iofacetune:DMSO (90:10) (experiment 2). Mice were sacrificed 5 h later, the epidermis was isolated and ornithine decarboxylase activity was determined. Data areexpressed as the mean ±SE of duplicate assays from at least six mice.
ExperimentTreatment1
AcetoneTPA(5nmol)TPA(5 nmol) + curcumin (0.5/unni)TPA(5 nmol) + curcumin (1pmol)TPA(5 nmol) + curcumin (3jimol)TPA(5 nmol) + curcumin (10jimo!)TPA(5 nmol) + retinoic acid (1.3nmol)2
Acetone:DMSO(90:10)TPA(5nmol)TPA(5 nmol) + chlorogenic acid (10¿iinol)TPA(5 nmol) + chlorogenic acid (20 //nini)TPA(5 nmol) + caffeic acid (10/imol)TPA(5 nmol) + caffeic acid (20^mol)TPA(5 nmol) + ferulic acid (10nmol)TPA(5 nmol) + ferulic acid (20jimol)TPA(5 nmol) + ferulic acid (50nmol)TPA(5 nmol) + curcumin (1jjmol)TPA(5 nmol) + curcumin (10 mnol)Ornithine
decarboxylase(pmol "CO2 released/
mgprotein/hr)31±12°3831
±6822659±8942082±544647±186°123
±54°472±90°17±
11°2683
±2502022±5091215±
118°1553±439°1068
±159°1470±190°1032±171°901±244°1636±440°255±130°Percent
inhibition ofTPA-induced ornithinedecarboxylaseactivity3146849888255542614662673991°
Statistically different from TPA alone (P < 0.05).
Table 2 Effect ofcurcumin, chlorogenic acid, caffeic acid, andferulic acid on TPA-induced ['HJtHymidine incorporation into epidermal DNA
TPA (2 nmol or 5 nmol) was applied topically alone or simultaneously with potential inhibitor in 200 j.1 of acetone (experiments 1 and 2) or in 200 it\ ofacetone:DMSO (90:10) (experiment 3). Eighteen h after application of TPA alone or TPA with the various compounds, the mice were given injections of [3H]thymidine.
Forty min later the mice were sacrificed, the epidermis was removed and radioactivity in epidermal DNA was determined. Data are expressed as the mean ±SE of16 mice.
ExperimentTreatment1
AcetoneTPA (2 nmol)TPA (2 nmol) + curcumin (1 umuhTPA (2 nmol) -I-curcumin (3 nmol)TPA (2 nmol) + curcumin (10 nmol)TPA (2 nmol) + fluocinolone acetonide (2nmol)2
AcetoneTPA (5 nmol)TPA (5 nmol) + curcumin (1 nmol)TPA (5 nmol) + curcumin (3 nmol)TPA (5 nmol) + curcumin ( 10 ,,mol>3
Acetone:DMSO (90:10)TPA (5 nmol)TPA (5 nmol) + chlorogenic acid (20 nmol)TPA (5 nmol) + caffeic acid (20 nmol)TPA (5 nmol) + ferulic acid (20 nmol)[3H]Thymidine
incorporation
into epidermal DNA(dpm/ngDNA)34.8
±5.5"
147.9 ±25.1143.5 ±15.4132.0 ±17.593.0 ±6.1°50.5 ±10.5"33.4
±3.3°
136.7 ±9.5133.7± 15.6132.6 ±6.9106.7±4.0°27.7
±2.9°
82.3 ±3.578.3 ±4.664.5 ±6.0°74.2
±7.4Percent
inhibitionof TPA-inducedDNA synthesis4
14498634
297
3315'
Statistically different from TPA alone (P < 0.05).
studies, the application of 3 or 10 ¿¿molof curcumin to an earof the mouse 30 min before 1 /¿molof arachidonic acid inhibitedarachidonic acid-induced edema by 33 or 80%, respectively(Table 4, experiment 2). In another experiment, the topicalapplication of 1.5 or 5 ¿¿molofcurcumin 30 min before 1 ^rnolof arachidonic acid inhibited arachidonic acid-induced edemaby 32 or 58%, respectively (Table 4, experiment 3). Phenidoneand nordihydroguaiaretic acid are known inhibitors of arachidonic acid metabolism (22, 23), and both of these compoundswere effective inhibitors of arachidonic acid-induced edema
(Table 4, experiments 2 and 3).
DISCUSSION
The results of the present study demonstrate that topicalapplication of curcumin inhibits TPA-induced epidermal ornithine decarboxylase activity, epidermal DNA synthesis, andpromotion of skin tumors in mice. The related compoundschlorogenic acid, caffeic acid, and ferulic acid are also active as
inhibitors of TPA-induced tumor promotion and ornithinedecarboxylase activity in mouse skin, but these compounds areless effective than curcumin. Curcumin is widely used as ayellow coloring agent and spice in commonly ingested foods,and chlorogenic acid, caffeic acid, and ferulic acid are normallyoccurring constituents of many human foods and beverages.
Application of TPA to mouse skin results in the rapid accumulation of inflammatory cells such as neutrophils and macrophages (24) and an increase in the release of active oxygenspecies (25, 26). As shown in Table 4, curcumin markedlyinhibited TPA- and arachidonic acid-induced inflammation.The results could be explained by a possible effect of curcuminto inhibit the infiltration of inflammatory cells and/or to inhibitthe release and action of reactive oxygen species from thesecells.
It has been suggested that reactive oxygen species and otherfree radicals play an important role in tumor promotion (24-27). Free radical-generating compounds such as benzoyl peroxide, lauroyl peroxide, and chloroperbenzoic acid have tumor-
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INHIBITION OF TPA-INDUCED TUMOR PROMOTION
r A
TPA(5nmol>-f•^—'Curcumin(IO^mol)
5 10 15 20WEEKS OF TPA APPLICATION
Fig. 2. Effect of curcumin on TPA-induced tumor promotion in mouse skin.Thirty mice/group were initiated with 200 nmol of DMBA. One week later themice were treated topically with 5 nmol TPA alone or together with various dosesof curcumin in 200 ¡ilacetone twice a week for 20 weeks. A, tumors/mouse; B,percentage of mice with tumors.
promoting activity on mouse skin (27-29), and application of
TPA to mouse skin has been shown to stimulate the releaseand metabolism of arachidonic acid (30-33) and to increase theformation of reactive oxygen species (25, 26). Superoxide dis-
mutase is an important enzymatic defense for the removal ofSuperoxide anión,and the biomimetic copper(II) (3,4-diisopro-
pylsalicylic acid)2, which possesses Superoxide dismutase activity, has been demonstrated to inhibit TPA-induced ornithinedecarboxylase activity and tumor promotion in mouse skin (34,35). Several inhibitors of TPA-dependent tumor promotionpossess antioxidant activity. Butylated hydroxytoluene (10),butylated hydroxyanisole (10, 25), sodium selenite (36), quer-cetin (37), nordihydroguaiaretic acid (38), a-tocopherol (39),
ascorbic acid (12), and ascorbyl palmitate (12) are examples ofcompounds that possess antioxidant or reactive oxygen-scav
enging activity and that inhibit tumor promotion and/or certainbiochemical events associated with tumor promotion in mouseskin. Although the antioxidant activity of chlorogenic acid hasnot been compared with that of curcumin, the relative antioxidant activities of curcumin, caffeic acid, and ferulic acid roughly
parallel the effects of these compounds as inhibitors of tumorpromotion (see Ref. 13 and Table 3).
Several inhibitors of arachidonic acid metabolism inhibitTPA-dependent tumor promotion in mouse skin (37, 38, 40,41), and it is thought that products of arachidonic acid metabolism may play an important role in the tumor promotionprocess. Mouse epidermis has been reported to possess A5-,A8-, A12-,and A"-lipoxygenase activity (37, 42-44). HPETEs
and HETEs, leukotrienes, and prostaglandins are among thearachidonic acid metabolic products that are believed to play arole in TPA-induced inflammation and tumor promotion (45-47). It was found that TPA stimulates cell proliferation andincreases the production of HETEs and prostaglandins in epidermal cell culture (44). In addition, inhibition of arachidonicacid metabolism results in lowered levels of HETEs and prostaglandins in human platelets (48). These products of arachidonic acid metabolism may be important in cell proliferationsince high levels of arachidonic acid, HPETEs, and HETEs arefound in the hyperproliferative disease psoriasis (49). Fluoci-nolone acetonide and other glucocorticoids represent anotherclass of potent inhibitors of TPA-dependent tumor promotion(7), and these steroids are inhibitors of epidermal phospholipaseA; activity in vivo and inhibit the release of arachidonic acidfrom cellular membranes (50). Nordihydroguaiaretic acid andquercetin are examples of As-lipoxygenase inhibitors that alsoinhibit the TPA-dependent induction of ornithine decarboxylase activity and tumor promotion in mouse skin (23, 37, 41).As indicated above, arachidonic acid metabolism is believed togive rise to reactive oxygen species and other free radicals, andstudies on the TPA-induced generation of reactive oxygenspecies in mouse epidermal cells have indicated that inhibitorsof arachidonic acid metabolism can diminish the chemilumi-nescense response that is associated with the generation of freeradicals (51). It is possible that arachidonic acid metabolismmay provide a source of free radicals that play a role in tumorpromotion. The inhibitory effect of curcumin on arachidonicacid-dependent ear edema (Table 4) suggests that curcuminmay block arachidonic acid metabolism by inhibiting epidermalcyclooxygenase and/or lipoxygenase activity, or that curcuminmay serve as a scavenger of reactive free radicals which areproduced during the metabolism of arachidonic acid. It is alsopossible that curcumin can interfere with the activity of epidermal phospholipase A2 and/or inhibit TPA-induced release of
arachidonic acid from membranes, since curcumin appears tobe a more potent inhibitor of TPA-induced mouse ear edemathan arachidonic acid-induced ear edema (Table 4). Additionalstudies are needed to determine the possible inhibitory effectsof curcumin, chlorogenic acid, caffeic acid, and ferulic acid on
Table 3 Effect of curcumin, chlorogenic acid, caffeic acid, and ferulic acid on TPA-induced tumor promotion in mouse skin
Female mice were treated topically with 200 nmol of DMBA in 200 u\ of acetone. One week later the mice were treated topically with 5 nmol of TPA alone ortogether with curcumin, chlorogenic acid, caffeic acid, or ferulic acid in 200 pi of acetone:DMSO (90:10) twice a week for 19 weeks. The tumors per mouse areexpressed as the mean ±SE from 30 mice.
Percent of mice withtumorsTreatmentAcetone:DMSO(90:10)TPA
(5nmol)TPA(5 nmol) + chlorogenic acid(10nmol)TPA
(5 nmol) + chlorogenic acid(20(imol)TPA
(5 nmol) + caffeic acid (10^mol)TPA(5 nmol) + caffeic acid (20/imol)TPA(5 nmol) + ferulic acid (10»imol)TPA(5 nmol) + ferulic acid (50>imol)TPA(5 nmol) + curcumin (10 nmol)11
wk048.323.36.724.711.534.50015wk060.740.06.743.323.044.83.3019wk067.946.626.746.738.563.513.3011wk0°3.79
±1.370.91±0.44°0.07
±0.05"3.47
±0.090.31±0.21"2.21
±0.960°0°Tumors/mouse15
wk0«5.53
±1.842.50±0.750.53
±0.10°4.43
±1.531.12+0.49°4.00
±1.020.17±0.08°0°19
wko-6.18
±1.842.50±0.780.53
±0.05°4.43
±1.891.12±0.67°4.00
±1.190.17±0.03°0°
" Statistically different from TPA alone (P < 0.05).
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INHIBITION OF TPA-INDUCED TUMOR PROMOTION
Table 4 Effect ofcurcumin on TPA- and arachidonic acid-induced edema ofmouse ears
In experiment 1, female CD-I mice (10 animals/group) were treated on theright ear with 25 nI of acetone or curcumin in acetone 30 min before theapplication of 25 n' of acetone or TPA in acetone. The mice were sacrificed 5 hlater by cervical dislocation, and ear punches from two animals were weighedtogether (6 mm diameter punch/animal). In experiments 2 and 3, female CD-Imice (10 animals per group) were treated on the right ear with 25 jil of acetoneor the potential inhibitor in acetone 30 min before the application of 25 <ilofacetone or 1 nmol of arachidonic acid in acetone. The mice were sacrificed l hlater by cervical dislocation, and ear punches from two animals were weighedtogether (6-mm-diameter punch/animal). The data from each group representsthe mean ±SE from 5 pooled samples obtained from 10 mice.
Experiment123TreatmentAcetoneTPA
(0.5nmol)TPA(0.5 nmol) +curcuminTPA
(0.5 nmol) +curcumin(1nmol)TPA
(0.5 nmol) +curcumin(3nmol)TPA
(0.5 nmol) +curcumin(10nmol)AcetoneArachidonic
acid (1nmol)Arachidonicacid (1 nmol)tcurcumin(3nmol)Arachidonicacid (1 nmol)+curcumin(10nmol)Arachidonicacid (1 nmol)+phenidone(2mg)AcetoneArachidonic
acid (1nmol)Arachidonicacid (1 nmol)+curcumin
(1.5nmol)Arachidonicacid (1 nmol)+curcumin
(5nmol)Arachidonicacid (1 nmol)+nordihydroguaiaretic
acid(1.5nmol)Arachidonic
acid (1 nmol)+nordihydroguaiareticacidWt/punch
(mg)7.35±0.17°12.78
±0.538.51±0.30°7.49
±0.19°6.78
±0.09°7.08
±0.20°7.11
±0.24°11.
57±0.6910.11±0.357.99
±0.28°8.45
±0.36"7.30
±0.14°10.85
±0.639.70±0.728.80
±0.29°9.07
+0.31°8.98
±0.30°Percent
inhibition799710010033807032585053
"Statistically different from TPA or arachidonic acid alone (P < 0.05).
epidermal phospholipase A2, cyclooxygenase, and lipoxygenaseactivity. Since protein kinase C is believed to be involved in theregulation of cellular proliferation and is a receptor for TPA(52), additional studies are also needed to determine the effectsof curcumin, chlorogenic acid, caffeic acid, and ferulic acid onprotein kinase C activity in the presence and absence of TPA.
Studies on the absorption and metabolism of curcumin indicate that it is absorbed after p.o. administration to rodents andthat it is rapidly metabolized to glucuronide and sulfate conjugates that are excreted primarily in bile and to a lesser extentin urine (53-55). Low or undetectable blood levels of unchangedcurcumin were observed after p.o. administration (54, 55),possibly because of its rapid metabolism during absorption (53-
55).Toxicity studies with turmeric or curcumin in animals indi
cated no histopathological changes when these substances werefed to rats, dogs, guinea pigs, or monkeys (0.5 to 2 g/kg) for 8-60 weeks (56). In addition, studies with turmeric and curcuminin rats for three generations did not show any teratogenic orcarcinogenic effects (54). However feeding turmeric oleoresinto pigs at a dose of 296 to 1551 mg/kg for 16 weeks decreasedbody weight gain and concomitantly increased the weight of theliver and thyroid gland (56). Hyperplasia of the thyroid andepithelial changes in kidney, urinary bladder, and liver werealso observed in the pigs fed turmeric oleoresin (56). Furtherstudies are needed to determine whether these adverse effectswere caused by curcumin per se or by other components in the
turmeric oleoresin. Recent studies have shown that feedingcaffeic acid, chlorogenic acid, or ferulic acid (2% in the diet) torats for 4 weeks resulted in hyperplasia throughout the fore-stomach epithelium in the animals fed caffeic acid, but little orno effect was observed when the rats were fed chlorogenic acidor ferulic acid (57).
Extracts of turmeric were reported to cause chromosomalaberrations in several mammalian cell lines (58) and in plantcells (59). In vivo studies with both turmeric and curcumin didnot show any genotoxic effects in the micronuclei test in mice,the bone marrow chromosome analysis test in mice and rats,or the dominant lethal test in mice (60, 61). Turmeric andcurcumin were inactive and evaluated for mutagenicity in Sal-monella typhimurium test systems in the presence of hepaticenzymes (62, 63). Curcumin was reported to possess cytotoxic(64) and antibacterial activity (65) and to inhibit the microsome-mediated mutagenicity of benzo[a]pyrene, 7,12-dimethylbenz-[a]anthracene, capsaicin, chili extract, and cigarette smoke condensate (62,66). Curcumin was also shown to inhibit the growthof tumor cells in vitro and to increase the survival of animalswith lymphomas (64).
The present study, which demonstrates an inhibitory effectof curcumin, chlorogenic acid, caffeic acid, and ferulic acid ontumor promotion by TPA in the two-stage initiation-promotionmodel in mouse skin, suggests a need for additional studieswith these compounds in several other experimental carcino-genesis models. These studies are particularly important because of the widespread dietary ingestion of curcumin, chlorogenic acid, caffeic acid, ferulic acid, and other plant phenolicsin varying proportions and amounts by the human population.
REFERENCES
1. Chopra, R. V, Chopra, I. ( '., Handa, K. 1... and Kapur. I.. D. In: Indigenous
Drugs of India, Ed. 2, pp. 325-327. Calcutta: Dhur, 1958.2. Nadkarn, K. M. Curcuma longa. ¡n:K. M. Nadkami (ed.), India Materia
Medica, pp. 414-416. Bombay: Popular Prakashan Publishing Co., 1976.3. Wealth of India. In: Raw Materials, p. 402. New Delhi: CSIR, 1950.4. Bacon, E. Health protection branch regulation of food additives. Can. Pharm.
J., 112: 340-344, 1979.5. Marmion, D. M. Colorants exempt from certification. In: Handbook of U.
S. Colorants for Food, Drugs and Cosmetics, pp. 86-87. New York: JohnWiley & Sons, 1979.
6. Govindarajan, V. S. Turmeric—chemistry, technology and quality. CRCCrit. Rev. Food Sci. Nutr., 12:199-301, 1980.
7. Schwarz, J. A., Viaje, A., and Slaga, T. J. Flucinolone acetonide: a potentinhibitor of mouse skin tumor promotion and epidermal DNA synthesis.Chem. Biol. Interact., 17:331-347, 1977.
8. Slaga, T. J., and Scribner, J. D. Brief communication: inhibition of tumorinitiation and promotion by anti-inflammatory agents. J. Nati. Cancer Inst.,51:1723-1725,1973.
9. Viaje, A., Slaga, T. J., Wigler, M., and Weinstein, I. B. Effects of antiinflam-matory agents on mouse skin tumor promotion, epidermal DNA synthesis,phorbol ester-induced cellular proliferation, and production of plasminogenactivator. Cancer Res., 37: 1530-1536, 1977.
10. Kozombo, W. J., Seed, J. L., and Kensler, T. W. Inhibition by 2(3)-fert-butyl-4-hydroxyanisole and other antioxidants of epidermal omithine decarboxyl-ase activity induced by 12-O-tetradecanoylphorbol-13-acetate. Cancer Res.,43: 2555-2559, 1983.
11. Nakadate, T., Yamamoto, S., Aizu, E., and Kato, R. Effects of flavonoidsand antioxidants on 12-O-tetradecanoylphorbol-13-acetate caused epidermalomithine decarboxylase induction and tumor promotion in relation to lipoxygenase inhibition by these compounds. Gann, 75: 214-222, 1984.
12. Smart, R. C, Huang, M-T., Han, Z. T., Kaplan, M. C, Focella, A., andConney, A. H. Inhibition of 12-O-tetradecanoylphorbol-l 3-acetate inductionof omithine decarboxylase activity, DNA synthesis, and tumor promotion inmouse skin by ascorbic acid and ascorbyl palmitate. Cancer Res., 47: 6633-6638, 1987.
13. Sharma, O. P. Antioxidant activity of curcumin and related compounds.Biochem. Pharmcol., 25: 1811-1812, 1976.
14. Toda, S., Miyase, T., Arichi, H., Tanizawa, H., and Takino, Y. Naturalantioxidant. III. Antioxidative components isolated from rhizome of Curcuma longa L. Chemical and Pharmaceutical Bulletin (Tokyo), 33: 1725-1728, 1985.
15. Srimal, R. C., and Dhawan, B. N. Pharmacology of diferuloyl methane
5945
on May 28, 2018. © 1988 American Association for Cancer Research. cancerres.aacrjournals.org Downloaded from
INHIBITION OF TPA-INDUCED TUMOR PROMOTION
(curcumin), a non-steroidal anti-inflammatory agent. J. Pharm. Pharmacol.,25:447-452, 1973.
16. Rao, T. S., Basu, N., and Siddiqui, H. H. Anti-inflammatory activity ofcurcumin analogues. Indian J. Med. Res., 75: 574-578, 1982.
17. Mukhopadhyay, A., Basu, N., Ghatak, N., and Gujral, P. K. Anti-inflammatory and irritant activities of curcumin analogues in rats. Agents andActions, 72:508-515, 1982.
18. Smart, R. C., Huang, M-T., and Conney, A. H. sn-l,2-Diacylglycerols mimicthe effects of 12-O-tetradecanoylphorbol-13-acetate in vivo by inducing biochemical changes associated with tumor promotion in mouse epidermis.Carcinogenesis (Lond.), 7:1865-1870, 1986.
19. Slaga, T. J., Das, S. B., Rice, J. M., and Thompson, S. Fractionation ofmouse epidermal chromatin components. J. Invest. Dermatol., 63:343-349,1974.
20. Lowry, O. H., Rosebrough, N. J., Farr, A. L., and Randall, R. J. Proteindetermination with Folin phenol reagent. J. Biol. Chem., 193: 265-275,1951.
21. Verma, A. K., and Boutwell, R. K. Vitamin A acid (retinoic acid), a potentinhibitor of 12-O-tetradecanoylphorbol-13-acetate-induced ornithine decar-boxylase activity in mouse epidermis. Cancer Res., 37: 2196-2201, 1977.
22. Fischer, S. M. Arachidonic acid metabolism and tumor promotion. In: S. M.Fischer and T. J. Slaga (eds.), Arachidonic Acid Metabolism and TumorPromotion, pp. 21-27. The Hague: Martinus Nijhoff Publishers, 1985.
23. Higgs, G. A., and Vane, J. R. Inhibition of cyclo-oxygenase and lipoxygenase.Br. Med. Bull., 39: 265-270, 1983.
24. Lewis, J. G., and Adams, D. O. Early inflammatory changes in the skin ofSENCAR and C57BL/6 mice following exposure to 12-O-tetradecanoylphorbol-13-acetate. Carcinogenesis (Lond.), 8: 889-898, 1987.
25. Copeland, E. S. (éd.).A National Institutes of Health Workshop Report.Free radicals in promotion—a chemical pathology study section workshop.Cancer Res., 43: 5631-5637, 1983.
26. Cerutti, P. A. Prooxidant states and tumor promotion. Science (Wash. DC),227: 375-381, 1985.
27. Slaga, T. J., Solanki, V., and Logan!, M. Studies on the mechanism of actionof antitumor promoting agents: suggestive evidence for the involvement offree radicals in promotion. In: O. F. Nygaard and M. G. Simic (eds.),Radioprotectors and Anticarcinogens, pp. 471-485. New York: AcademicPress, 1983.
28. Slaga, T. J., Klein-Szanto, A. J. P., Triple«, L. L., and Yotti, L. P. Skintumor-promoting activity of benzoyl peroxide, a widely used free radical-generating compound. Science (Wash. DC), 213:1023-1025, 1981.
29. Klein-Szanto, A. J. P., and Slaga, T. J. Effects of peroxides on rodent skin:epidermal hyperplasia and tumor promotion. J. Invest. Dermatol., 79: 30-34, 1982.
30. Bresnick, E., Bailey, G., Bonney, R. J., and Wightman, P. Phospholipaseactivity in skin after application of phorbol esters and 3-methylcholanthrene.Carcinogenesis (Lond.), 2: \ 119-1122, 1981.
31. Bresnick, E., Meunier. P., and Lamden, M. Epidermal prostaglandins aftertopical application of a tumor promoter. Cancer Lett., 7:121-125, 1979.
32. Ashendel, C. L., and Boutwell, R. K. Prostaglandin E and F levels in mouseepidermis are increased by tumor-promoting phorbol esters. Biochem. Biophys. Res. Commun., 90: 623-627, 1979.
33. /¡boh,V. A. Arachidonic acid metabolism in the skin. In: S. M. Fischer andT. J. Slaga (eds.), Arachidonic Acid Metabolism and Tumor Promotion, pp.6-20. The Hague: Martinus Nijhoff Publishers, 1985.
34. Egner, P. A., and Kensler, T. W. Effects of a biomimetic Superoxide dismutaseon complete and multistage Carcinogenesis in mouse skin. Carcinogenesis(Lond.),«: 1167-1172, 1985.
35. Kensler, T. W., Bush, D. M., and Kozumbo, W. J. Inhibition of tumorpromotion by a biomimetic Superoxide dismutase. Science (Wash. DC), 227:75-77, 1983.
36. Shamberger, R. J. Increase of peroxidation in Carcinogenesis. J. Nati. CancerInst., «.-1491-1497, 1972.
37. Kalo, R., Nakadate, T., Yamamoto, S., and Sugimura, T. Inhibition of 12-O-tetradecanoylphorbol-13-acetate-induced tumor promotion and ornithinedecarboxylase activity by quercetin: possible involvement of lipoxygenaseinhibition. Carcinogenesis (Lond.), 4:1301-1305, 1983.
38. Nakadate, T., Yamamoto, S., Ishii, M., and Kato, R. Inhibition of 12-O-tetradecanoylphorbol- 13-acetate-induced epidermal ornithine decarboxylaseactivity by lipoxygenase inhibitors: possible role of product(s) of lipoxygenasepathway. Carcinogenesis (Lond.), 3: 1411-1414, 1982.
39. Perchellet. J-P., Owen, M. D., Posey, T. D., Orten, D. K., and Schneider, B.A. Inhibitory effects of glutathione level-raising agents and D-a-tocopherolon ornithine decarboxylase induction and mouse skin tumor promotion by12-O-tetradecanoylphorbol-13-acetate. Carcinogenesis (Lond.), 6: 567-573,1985.
40. Fischer, S. M., Mills, G. D., and Slaga, T. J. Inhibition of mouse skin tumorpromotion by several inhibitors of arachidonic acid metabolism. Carcinogenesis (Lond.), 3:1243-1245, 1982.
41. Nakadate, T., Yamamoto, S., Ishii, M., and Kalo, R. Inhibition of 12-0-tetradecanoylphorbol-13-acetate-induced epidermal ornithine decarboxylaseactivity by phospholipase A., inhibitors and lipoxygenase inhibitors. CancerRes., 42: 2841-2845, 1982.
42. Nakadate, T., Aizu, E., Yamamoto, S., and Kato, R. Some properties oflipoxygenase activities in cytosol and microsomal fractions of mouse epidermal homogenate. Prostaglandins Leukotrienes Med., _'/: 305-319, 1986.
43. Gschwendt, M., Furstenberger, G., Kittstein, W., Besemfelder, E., Hull, W.E., Hagedorn, H., Opferkuch, H. J., and Marks, F. Generation of thearachidonic acid metabolite 8-HETE by extracts of mouse skin treated withphorbol ester in vivo: identification by H-NMR and GC-MS spectroscopy.Carcinogenesis (Lond.), 7:449-455, 1986.
44. Fischer, S. M., Baldwin, J. K., Jasheway, D. W., Patrick, K. E., and Cameron,G. S. Phorbol ester induction of 8-lipoxygenase in inbred SENCAR (SSIN)but not C57BL/6J mice correlated with hyperplasia, edema, and oxidantgeneration but not ornithine decarboxylase induction. Cancer Res., 48:658-664, 1988.
45. Higgs, G. A., Salmon, J. A., and Spayne, J. A. The inflammatory effects ofhydroperoxy and hydroxy acid products of arachidonate lipoxygenase inrabbit skin. British Journal of Pharmacology, 74: 429-433, 1981.
46. Camp, R. O. R. Prostaglandins, hydroxyfatty acids, leukotrienes, and inflammation of the skin. Clin. Exp. Dermatol., 7:435-444, 1982.
47. Chang, J., Carlson, R. P., O'Neill-Davis, L., Lamb, B., Sharma, R. N., and
Lewis, A. J. Correlation between mouse skin inflammation induced byarachidonic acid and eicosanoid synthesis. Inflammation, 10:205-214,1986.
48. Hamberg, M., and Samuelsson, B. Prostaglandin endoperoxides. Novel transformations of arachidonic acid in human platelets. Proc. Nati. Acad. Sci.USA, 71: 3400-3404, 1974.
49. Hammarstrom, S., Hamberg, M., Samuelsson, B., Duell, E. A., Stawiski,M., and Voorhees, J. J. Increased concentrations of nonesterified arachidonicacid, 12£-hydroxy-5,8,10,14-eicosatetraenoic acid, prostaglandin I-"...,and
Prostaglandin F^ in epidermis of psoriasis. Proc. Nati. Acad. Sci. USA, 72:5130-5134, 1975.
50. Gryglewski, R. J. Effects of antiinflammatory steroids on arachidonatecascade. In: G. Weissman, B. Samuelsson, and R. Paoletti (eds.), Advancesin Inflammation Research, Vol. 1, pp. 505-513. New York: Raven Press,1979.
51. Fischer, S. M., and Adams, L. M. Suppression of tumor promoter-inducedchemiluminescence in mouse epidermal cells by several inhibitors of arachidonic acid metabolism. Cancer Res., 45: 3130-3136, 1985.
52. Niedel, J. E., Kühn,L. J., and Vandenbank, G. R. Phorbol diester receptorcopurifies with protein kinase C. Proc. Nati. Acad. Sci. USA, 80: 36-40,1983.
53. Ravindranath, V., and Chandrasekhara, N. Absorption and tissue distributionof curcumin in rats. Toxicology, 16: 259-265, 1980.
54. Wahlstrom, B., and Blennow, G. A study on the fate of curcumin in the rat.Acta Pharmacol. Toxicol., 43:86-92, 1978.
55. Ravindranath, V., and Chandrasekhara, N. Metabolism of curcumin—studieswith [3H]curcumin. Toxicology, 22:337-344, 1982.
56. Bille, N., I.arsen. J. C., Hansen, E. V., and Wurtzen, G. Subchronic oraltoxicity of turmeric oleoresin in pigs. Food Chem. Toxicol., 23: 967-973,1985.
57. Hirose, M., Masuda, A., Imaida, K., Kagawa, M., Tsuda, H., and Ito, N.Induction of forestomach lesions in rats by oral administrations of naturallyoccurring antioxidants for four weeks. Jpn. J. Cancer Res. (Gann), 78: 317-321, 1987.
58. Goodpasture, C. E., and Arrighi, F. E. Effects of food seasonings on the cellcycle and chromosome morphology of mammalian cells in vitro with specialreference to turmeric. Food Cosmet. Toxicol., 14: 9-14, 1976.
59. Abraham, S., Abraham, S. K., and Radhamony, G. Mutagenic potential ofthe condiments, ginger and turmeric, Cytologia (Tokyo), 41: 591-595,1976.
60. Vijayalaxmi. Genetic effects of turmeric and curcumin in mice and rats.Mutât.Res., 79:125-132, 1980.
61. Abraham, S. K., and Kesavan, P. C. Genotoxicity of garlic, turmeric andasafoetida in mice. Mutât.Res., 136:85-88, 1984.
62. Nagabhushan, M., and Bhide, S. V. Nonmutagenicity of curcumin and itsantimutagenic action versus chili and capsaicin. Nutr. Cancer, 8: 201-210,1986.
63. Jensen, N. J. Lack of mutagenic effect of turmeric oleoresin and curcuminin the Sa/monf/to/mammalian microsome test. Mutat. Res., 105: 393-396,1982.
64. Kuttan, R., Bhanumathy, P., Ninnala, K., and George, M. C. Potentialanticancer activity of turmeric (Curcuma longa). Cancer Lett., 29: 197-202,1985.
65. Ramaprasad, C., and Sirsi, M. Indian medicinal plants. Curcuma longa invitro antibacterial activity of curcumin and the essential oil. J. Sei. Ind. Res.(India), 156: 239-241, 1956.
66. Nagabhushan, M., Amonkar, A. J., and Bhide, S. V. In vitro antimutagenicityof curcumin against environmental mutagens. Food Chem. Toxicol., 25:545-547, 1987.
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1988;48:5941-5946. Cancer Res Mou-Tuan Huang, Robert C. Smart, Ching-Quo Wong, et al. -Tetradecanoylphorbol-13-acetate
Oand Ferulic Acid on Tumor Promotion in Mouse Skin by 12-Inhibitory Effect of Curcumin, Chlorogenic Acid, Caffeic Acid,
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