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Effects of dietary n3ton6 polyunsaturated fattyacid ratio on mammary carcinogenesis in ratsTaku Sasaki a , Yasuhiko Kobayashi b , Jun Shimizu b , Masahiro Wada b , Satoshi In'nami b

, Yusuke Kanke a & Toshichika Takita ca Division of Bioregulation Studies , Tokyo University of Agriculture , Tokyo, 156, Japanb Department of Nutrition , Tokyo University of Agriculture , Tokyo, 156, Japanc Dept. of Nutrition , Tokyo University of Agriculture , 111 Sakuragaoka, Setagayaku,Tokyo, 1560054, Japan Phone: 81 03 5477 2443 Fax: 81 03 5477 2443Published online: 04 Aug 2009.

To cite this article: Taku Sasaki , Yasuhiko Kobayashi , Jun Shimizu , Masahiro Wada , Satoshi In'nami , Yusuke Kanke &Toshichika Takita (1998) Effects of dietary n3ton6 polyunsaturated fatty acid ratio on mammary carcinogenesis in rats,Nutrition and Cancer, 30:2, 137-143, DOI: 10.1080/01635589809514653

To link to this article: http://dx.doi.org/10.1080/01635589809514653

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NUTRITION AND CANCER, 30(2), 137-143Copyright 1998, Lawrence Eribaum Associates, Inc.

Effects of Dietary n-3-to-n-6 Polyunsaturated Fatty AcidRatio on Mammary Carcinogenesis in Rats

Taku Sasaki, Yasuhiko Kobayashi, Jun Shimizu, Masahiro Wada,Satoshi In'nami, Yusuke Kanke, and Toshichika Takita

Abstract: We investigated the effects of the dietary n-3-to-n-6 polyunsaturated fatty acid (PUFA) ratio (n-3/n-6 ratio)on mammary carcinogenesis induced by 7,12-dimethyl-benz[a]anthracene in rats by feeding them several types ofdietary fat with a fixed PUFA-to-saturated fatty acid ratio.Dietary fat was fed to the rats as 10% of the total feedweight, starting two weeks before the initiation. An increasein the n-3/n-6 ratio did not suppress the incidence or reducethe latency of mammary tumor development. The numberand weight of mammary tumors per tumor-bearing rattended to be large in the group with an n3/n-6 ratio of 7.84compared with those in the other groups. As the n3/n-6ratios were elevated, the total number and weight of tumorsincreased gradually. The prostaglandin E2 (PGE2) concen-tration in mammary tumor tissue was markedly low in thegroup with an n-3/n-6 ratio of 1.03 compared with thegroup with an n-3/n-6 ratio of 0.01. In addition, PGE2concentrations were almost constant when n-3/n-6 ratioswere >1.03. These results suggested that the increase in then3/n-6 ratio of dietary fat with the fixed PUFA-to-satu-rated fatty acid ratio cannot suppress the mammary carcino-genesis but can promote development of tumors, despitereduced PGE2 concentration in the tumor.

Introduction

The quantity and quality of fat ingested are representativedietary factors that greatly affect the development of mam-mary and colonie tumors (1-4). Epidemiologie studies in-volving Greenlanders, who consume large amounts of fat,revealed relationships between not only the amount of fatsbut also the type of fatty acids ingested and the developmentof tumors (1,2). The results of experiments using animalsalso suggested that development of mammary tumors, whichcommonly are observed in Europeans, was promoted by anincrease in the amount of fat ingested (3,4). Thus the rela-tionship between the development of tumors and quality offat ingested has become a subject of research (521).

Mammary carcinogenesis is promoted by 18:2, one of then-6 polyunsaturated fatty acids (PUFAs) (5,6). It was re-ported that the incidence of mammary tumors increased as aresult of ingesting fat containing a high concentration of 18:2,but not as a result of ingesting fat containing the sameconcentration of medium-chain fatty acids or 18:1 (7). How-ever, Ip and co-workers (8) indicated in their study, in whichthe 18:2 concentration of dietary fat was increased in astepwise manner, that the promotional effect of dietary fat on7,12-dimethylbenz[a]anthracene (DMBA)-induced mam-mary carcinogenesis was closely related to the level of 18:2.That is, the incidence and the number of mammary tumorsincreased linearly with increasing 18:2 concentration up to4.4%; however, the effect of 18:2 on mammary carcinogene-sis did not continue to change when the concentration in-creased beyond this point. In contrast, it has been verified bya number of studies (10-21) that when fish oil (FO)- or perillaoil-based diets, which contain large amounts of n-3 PUFA,were ingested, the incidence and the proliferation of mam-mary tumors were suppressed compared with cases in whichplant oil containing large amounts of n-6 PUFA [i.e., saf-flower oil (SO) or corn oil (CO)] was ingested.

However, the various kinds of fat used in the above ani-mal experiments differ not only in the kinds of PUFA theycontain but also in the amounts of total PUFA, monoun-saturated fatty acid (MUFA), and saturated fatty acid (SFA).Therefore, we cannot conclude that the suppression of theincidence of mammary tumors induced by the ingestion offish oil is due only to the effect of n-3 PUFA. The reasonfor this is that FO contains larger amounts of SFA andMUFA than does SO, so the incidence of mammary tumorscould be suppressed by the intake of SFA or MUFA (7,10).In the current study, we investigated the effects of dietaryfat with different n-3/n-6 ratios on the development of mam-mary carcinogenesis induced by the administration ofDMBA through administration of several types of dietaryfat with constant PUFA-to-SFA (P/S) ratio, for which then-3/n-6 ratio was increased in a stepwise manner, using FOand SO as sources of n-3 and n-6 PUFA, respectively.

T. Sasaki and Y. Kanke are affiliated with the Division of Bioregulation Studies and Y. Kobayashi, J. Shimizu, M. Wada, S. In'nami, and T. Takitawith the Department of Nutrition, Tokyo University of Agriculture, Tokyo 156, Japan.

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Methods Table 2. Fatty Acid Composition of Dietary Fats"*

Animals and Diets

Twenty-eight-day-old female Sprague-Dawley rats (To-kyo Laboratory Animal Science, Tokyo, Japan) were usedin all experiments. They were housed in an air-conditionedroom (22 1C and 50 10% humidity) under a 12:12-hourlight-dark cycle. For adaptation to the experimental regimen,the rats were fed MF-2 rat chow (Oriental Yeast, Tokyo,Japan) and divided into experimental dietary groups to givethe same values obtained in body weight in each of thegroups. They were housed in individual cages. Semipurifieddiets were prepared daily by adding the oils to the otherdietary components. The food was placed in the cages at1700 and removed at 0900 the following morning to avoidthe ingestion of lipid peroxides. The stock powder diet andtest oils were stored separately at -30C in dark bottlesunder nitrogen until use. The degree of lipid peroxidationof test oils was monitored by thiobarbituric acid-reactivesubstance (TBARS) assay (22), and no lipid peroxidationwas detected. The rats were allowed water ad libitum. Thesemipurified diet contained 20% casein, 0.3% DL-methio-nine, 60% sucrose, 3.5% mineral mix (AIN-76), 1% vitaminmix (AIN-76), 0.2% choline bitartrate, 5% cellulose, and10% (wt/wt) fat. The fats were prepared by mixing FO (sar-dine oil, Nihon Oil and Fats, Tokyo, Japan), SO (Nihon Oiland Fats), and coconut oil (CCO; Hayashi Chemicals, To-kyo, Japan). The fatty acid compositions of the dietary fatsused in each experiment were analyzed by gas chromatog-raphy (Tables 1 and 2). Briefly, the dietary fats were trans-Table 1. Fatty Acid Composition of Coconut, Safflower,and Fish Oil"*

Fatty Acids

8:010:012:014:016:018:018:ln-918:2n-620:5n-322:6n-3

SFAMUFAPUFAPUFA/SFAn-3 PUFAn-6 PUFAn-3/n-

CCO

4.25.4

47.718.610.63.28.51.7NDND

89.88.51.70.02ND1.70.00

Experiment

SO

NDNDND0.16.62.4

14.775.1NDND

9.215.075.8

8.250.4

75.10.01

1

FO

ND0.10.26.2

14.92.7

14.71.6

16.411.8

26.830.942.4

1.5836.73.79.87

CCO

3.85.4

49.119.210.12.78.21.7NDND

90.18.21.70.02ND1.70.00

Experiment 2

SO

NDNDND0.27.12.4

13.874.7NDND

9.714.076.47.910.9

74.70.01

FO

ND0.11.38.1

17.43.4

14.61.9

16.39.7

32.427.140.5

1.2533.24.08.40

Fatty Acids

8:010:012:014:016:018:018:ln-918:2n-620:5n-322:6n-3

SFAMUFAPUFAPUFA/SFAn-3 PUFAn-6PUFAn-3/n-6

GroupI

2.82.7

20.88.18.12.6

11.943.7NDND

43.712.044.3

1.010.3

43.70.01

Experiment 1

GroupII

0.61.2

12.18.0

11.72.6

13.117.38.76.3

39.021.739.3

1.0119.418.81.03

GroupIII

0.70.88.78.2

13.62.5

13.74.7

12.38.8

37.726.435.9

0.9627.2

6.93.96

GroupIV

0.50.77.68.1

13.92.5

13.61.6

13.910.1

36.426.936.7

1.0130.9

3.97.84

Experiment 2

GroupA

2.12.4

21.78.48.42.5

11.342.5NDND

45.611.443.1

0.950.3

42.50.01

GroupB

1.31.3

11.88.9

13.42.8

13.017.69.45.6

40.719.639.70.98

19.119.01.00

a: Values are percentages, except for ratios.b: Abbreviations are as follows: CCO, coconut oil; SO, safflower oil;

FO, fish oil; ND, not detected; SFA, saturated fatty acids; MUFA,monounsaturated fatty acids; PUFA, polyunsaturated fatty acids.

a: Values are percentages, except for ratios. See Table 1 footnote fordefinition of abbreviations.

b: Mixing proportions of the oils were as follows (wt%): in Experiment1, 42.3:57.7 CCO-SO (Group I), 22.0:18.4:59.6 CCO-SO-FO (GroupII), 14.6:4.1:81.3 CCO-SO-FO (Group III), and 12.6:0.3:87.1 CCO-SO-FO (Group IV); in Experiment 2, 45.3:54.7 CCO-SO (Group A)and 20.3:18.7:61.0 CCO-SO-FO (Group B).

methylated with boron trifluoride-methanol complex (23)and then analyzed on a gas Chromatograph (model GC-12A,Shimadzu, Kyoto, Japan) equipped with a flame ionizationdetector and a 0.25 mm x 40 m stainless steel capillarycolumn (Silar 5CP, Chromatotec, Tokyo, Japan). Analysiswas conducted at a column temperature of 200C and inlettemperature of 250C, with nitrogen gas (2.2 ml/min) as thecarrier. A single dose of 15 mg of DMBA (Sigma Chemical,St. Louis, MO) dissolved in 1 ml of SO was given to eachrat on Day 49 after birth. The rats were palpated daily tomonitor tumor development.

Analysis

Experiment 1: Ninety-six rats were divided into the fourdietary groups indicated in Table 2. The n-3/n-6 ratios ofindividual test lipids were adjusted to 0.01 (Diet I), 1.03 (DietII), 3.96 (Diet III), and 7.84 (Diet IV). Sixty-six days afterDMBA was administered, the rats were anesthetized withpentobarbital sodium (Somnopentyl, Kyoritsu, Tokyo, Ja-pan). Blood was collected from the abdominal aorta. Tumorswere removed rapidly, frozen in liquid N2, and stored untiluse in the prostaglandin E2 (PGEj) assay. PGEj in tumors wasextracted using a modification of the method of Fritshe andJohnston (24). Briefly, portions of the mammary tumors werequickly homogenized in phosphate-buffered saline using aTeflon-Potter homogenizer under liquid N2. The homogenatewas filtered through three layers of cheesecloth and thenincubated at 37C for one hour. Immediately after incubation,

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350

300 -

O)250 -

01'5

200"OOm

150 -

100

1 3 5 7 9 11

Weeks after experimental diet administeredFigure 1. Body weight changes in 7,12-dimethylbenz[a]nthracene(DMBA)-treated rats. Thin solid line, Group I (n-3/n-6 = 0.01); thin dashedline, Group II (n-3/n-6 = 1.03); thick dashed line, Group HI (n-3/n-6 =3.96); thick solid line, Group IV (n-3/n-6 = 7.84).

aliquots of the homogenate were frozen. These samples werethen stored at -80C until analysis. PGE2 concentration wasdetermined by enzyme-linked immunosorbent assay (Amer-sham, Tokyo, Japan).

Experiment 2: Twenty-four rats were divided into thetwo dietary groups indicated in Table 2. The n-3/n-6 ratiosof individual test lipids were adjusted to 0.01 (Diet A) and1.00 (Diet B). Blood was collected in the same manner as inExperiment 1. An aliquot of blood was heparinized rapidly.Tumors and livers were removed rapidly, frozen in liquid N2,and stored until use in the lipid peroxide assay.

Peripheral blood lymphocytes (PBLs) were isolated fromheparinized blood using Percoll (Sigma Chemical) gradientcentrifugation. Cell numbers and viability were determinedby trypan blue exclusion as detected microscopically. PBLswere stained with fluorescein isothiocyanate-conjugated mon-oclonal antibody to rat CD4 (W3/25, Serotec, Kidlington,Oxford, UK) and phycoerythrin-conjugated monoclonal an-tibody to rat CD8 (MRC OX-8, Serotec) for two-color stain-ing. Stained cells were analyzed using EPICs Elite (Coulter,Miami, FL) after the exclusion of dead cells by forward andside light scattering. Tumors and livers were homogenizedin 1.15% (wt/vol) KC1 buffer using an ice-cold Teflon-Potterhomogenizer. Lipid peroxide concentration was determinedby the method of Kikugawa and co-workers (22).

Table 4. Mammary Tumors in DMBA-Treated Ratsa

100

&

30 40 SO 60 70Time after Carcinogen Treatment

(days)Figure 2. Cumulative palpable tumor incidence in DMBA-treated rats.Lines as defined in Figure 1 legend.

150

Time after Carcinogen Treatment(days)

Figure 3. Cumulative palpable tumor number in DMBA-treated rats. Linesas defined in Figure 1 legend.

cated that the weights of mammary tumors per tumor-bear-ing rat were positively correlated with increasing n-3/n-6ratio (v = 1.278 xx- 6.35, r2 = 0.83).

The PGE2 concentration per gram of the mammary tu-mors was markedly lower in the rats in the Diet II groupthan in the rats in the Diet I group (Figure 4). However, the

1000 -

750 .

500 -

250 -

Groups I III IVFigure 4. Prostaglandin E2 (PGE2) concentration in tumors of DMBA-treated rats. PGE2 concentrations were determined by enzyme-linkedimmunosorbent assay. Groups sharing a letter (a, b) or without letters arenot significantly different (p > 0.05, analysis of variance combined withDuncan's new multiple comparison test), n-3/n- ratios of dietary fats areas follows: 0.01 (Group I), 1.03 (Group II), 3.96 (Group III), and 7.84(Group IV).

PGE2 concentration was almost constant when the n-3/n-6ratio was >1.03.

Experiment 2

The n-3/n-6 ratio was observed to have the same effectas in Experiment 1 on the incidence of mammary tumorsand on the body weight gain of the rats (data not shown).

Although the n-3/n-6 ratio in the dietary fat was differentin the two dietary groups, the number of PBLs was similar.The percentage of CD4+CD8", CD4-CD8+, and CD4"CD8-cells in the PBLs of the two groups was also similar (Table 5).

The concentration of lipid peroxides per liver weight in therats was higher in the Diet B group than in the Diet A group,but no difference was observed in the concentration of lipidperoxides per weight of mammary tumors (Figure 5).

Discussion

In cases where FO-based diets were administered, suppres-sion of mammary tumor incidence was indicated to result

Table 5.

Groups

AB

Number of PBLs and Flow Cytometry Analysis in

No. of PBLs, 106 cells/ml

1.3 0.21.8 0.2

CD4SP

43 445 3

DMBA-Treated Rats'"Flow Cytometry

CD8SP

22 427 3

Analysis, %

DN

35 727 5

CD4SP/CD8SP

2.3 0.31.8 0.1

a: Values are means SE.b: Abbreviations are as follows: PBLs, peripheral blood lymphocytes; SP, single positive; DN, double negative.c: n-3/n-6 ratios of dietary fats are as follows: 0.01 (Group A) and 1.00 (Group B).

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300

250

.200toXLm 150"5I loo

50

Groups B BFigure 5. Lipid peroxide concentration of liver (left) and tumor (right) in DMBA-treated rats. Lipid peroxides were analyzed by thiobarbituric acid-reactivesubstances assay. Values are means SE. n-3/n-6 ratios of dietary fats are as follows: 0.01 (Group A) and 1.00 (Group B).

from the intake of n-3 PUFA. Cave and Jurkowski (17)showed in their study, in which a mixture of FO and CO thatcomprised 20% of the total feed weight was administered, thatthe latency was longer in a group fed fat with an n-3/n-6 ratioof 1.4 than in a group fed CO. Bunce and colleagues (18)observed a suppression of the incidence of mammary tumorsin a group fed fat with an n-3/n-6 ratio of 1.18 compared withCO. Ip and associates (19) also observed suppression of theincidence of mammary tumors in a group fed fat with ann-3/n-6 ratio of 0.71 when dietary fat comprised 20% of thetotal feed weight similar to the above studies (17,18). Inaddition, in another study by Karmali and others (20), inwhich dietary fat comprised 23.5% of the total feed weight,suppression of mammary tumor incidence was observed in agroup fed fat with an n-3/n-6 ratio of 0.91.

In the current study, using FO, SO, and CCO, we preparedvarious fat mixtures with constant P/S ratio and variousn-3/n-6 ratios and fed the mixtures to rats as 10% of theirtotal feed weight. The results indicate that the incidence ofmammary tumors was not suppressed but that the total num-ber and weight of the tumors were increased as a result offeeding the rats dietary fat with a high n-3/n-6 ratio. Theseresults disagree with the results of previous studies (17-20).

Previously, Cohen and co-workers (21) reported that then-3/n-6 ratio had selective inhibitory effects that were ob-served only when equal parts of FO and CO were fed torats exhibiting Af-nitroso-TV-methylurea-induced mammarytumors. However, direct comparison of their results withours is difficult, because 1) the total level of dietary fat theyused (23.52%) is not equal to the level we used and maybe too high for rats, since the level recommended by theAmerican Institute of Nutrition is 5% of the total feedweight; 2) the P/S ratios of the dietary fat in the study ofCohen and co-workers varied from 2.0 to 4.7; and 3) wedid not study the n-3/n-6 ratio of 0.5 with which they founda selective effect.

Many studies (18,25-28) regarding the relationship be-tween eicosanoids and the incidence of mammary tumors

have been carried out; however, the results are inconsistent.Some investigators reported a close relationship between theproliferation of tumor cells and the concentration of PGE2,which is responsible for the suppressive effects of n-3 PUFAon mammary cancer. However, the other researchers deniedany role for PGE2 in the development of mammary cancer.

The analysis of PGE2 concentration indicates the absenceof a correlation between the PGE2 concentration and theincidence of mammary tumors. This result supports theresults of Carter and co-workers (29) and Fritsche andJohnston (24). Unfortunately, the leukotriene concentrationwas not determined in the current study, and therefore acomplete evaluation of eicosanoid metabolism was impossi-ble. However, in a previous study in which dietary fat with ann-3/n-6 ratio of 1.18 was used, which is similar to that usedin our study, although the P/S ratios were different, a decreasein the leukotriene concentration was observed (18). Accord-ingly, the relationship between eicosanoid metabolism andthe incidence of mammary tumors is assumed to be weak.

However, in Experiment 1 of the current study, althoughthe 18:2 level in the diet fed to the rats in the Diet IV groupwas 1.8%, the incidence of mammary tumors was not sup-pressed but was promoted. According to the above-men-tioned reports (9,17-20), this 18:2 level should besufficiently low to suppress the incidence of mammary tu-mors. Considering the suppression of the incidence of mam-mary tumors equal to FO-fed groups observed in theCCO-fed groups by Craig-Schmidt and colleagues (10) andthe current results, the suppression of the incidence of mam-mary tumors may depend not only on reducing the 18:2level in dietary fat but also on the SFA content in the dietaryfat; since in the current experiment the P/S ratio of fat fedto rats in all the groups was set at 1.0, the effect of sup-pression of mammary carcinogenesis induced by n-6 PUFAmight possibly be masked by the effect of SFA. The factthat we did not observe any relationship between PGEj con-centrations in tumor tissues and the incidence of mammarytumors may support this possibility.

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Previously, Gonzalez and associates (15) reported thatproliferation of tumor cells was suppressed by increasedlevels of lipid peroxide reactions (TBARS) in the tumorsinduced by FO administration. In Experiment 2 of the cur-rent study, although the concentration of lipid peroxides inthe livers of rats was higher in the Diet A group than in theDiet B group, increases in the amounts of lipid peroxidesin mammary tumor tissues were not observed. This fact maysupport a relationship between an increasing n-3/n-6 ratioand the increasing weight of mammary tumors observed inthe rats in Experiment 1, which was contradictory to theresults of previous reports. In contrast, Bertoli and others(30) reported recently that four weeks of low-dose admin-istration of n-3 PUFAs that have the same ratio as ourcondition (n-3/n-6 ratio = 1.20) at lower quantity (as 5.5%of total feed weight) does not induce harmful modificationsof oxidative cell metabolism. This apparent conflict betweenour results and their results may be explained by differencesin quantity of dietary fats and/or the period of administration.However, these current results cannot be explained in termsof the P/S ratio or the n-3/n-6 ratio alone. In addition, theTBAR assay, which was used in Experiment 2 and the pre-vious study (15), is a nonspecific method. Therefore, it isdifficult to explain the effect of high n-3/n-6 ratio on tumorincidence by the results shown in Figure 5 alone.

In addition, in Experiment 2, to investigate whether anunbalanced immunosurveillance system is involved in thedevelopment of tumors observed in rats fed an n-3 PUFA-rich diet, we analyzed the effect of dietary fat on PBLs. Theresults indicate no effect of the n-3/n- ratio on the per-centage of T lymphocyte subsets in PBLs. Without admin-istration of DMBA, however, the activity of natural killercells in PBLs in the group fed Diet A was significantly (p< 0.05) reduced compared with the group fed Diet B (un-published data; the mean specific cytotoxicities were 15.3%and 9.0%, respectively). Despite the results obtained whenDMBA was not administered, the decrease in activity ofnatural killer cells may explain the increase in the incidenceof tumor development.

Nevertheless, further investigation is required not onlyof the response of organisms to lower n-3/n-6 ratios, be-cause those used in the current study were higher than thosein actual human diets, but also of the cause of the promo-tional effect of a diet with a high n-3/n-6 ratio on tumordevelopment.

Acknowledgments and Notes

The authors thank Dr. Satoru Moriguchi (Dept. of Nutrition, School ofMedicine, University of Tokushima) for advice on the preparation of PBLs,Dr. Akio Ni'ibe for valuable advice in statistical analysis, and Keita Kudohfor excellent assistance. Address reprint requests to Toshichika Takita,Dept. of Nutrition, Tokyo University of Agriculture, 1-1-1 Sakuragaoka,Setagaya-ku, Tokyo 156-0054, Japan. Phone: 81 03 5477 2443. FAX: 8103 5477 2626.

Submitted 11 July 1997; accepted in final form 4 December 1997.

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References

1. Sincair, HM: The relative importance of essential fatty acids of thelinoleic and linolenic families: studies with the Eskimo diet. ProgLipid Res 20, 897-899, 1981.

2. Kaizer, L, Boyd, MF, Kriukov, V, and Tritchler, D: Fish consumptionand breast cancer risk: an ecological study. Nutr Cancer 12, 61-68, 1989.

3. Carter, CA, Milholland, RJ, Shea, W, and Ip, MM: Effect of theprostaglandin synthetase inhibitor indomethacin on 7,12-dimethyl-benz[a]anthracene-induced mammary tumorigenesis in rats fed differ-ent levels of fat. Cancer Res 43, 3559-3562, 1983.

4. Lane, HW, Teer, P, Keith, RE, White, MT, and Strahan, S: Reducedenergy intake and moderate exercise reduce mammary tumor incidencein virgin female BALB/c mice treated with 7,12-dimethylbenz[a]an-thracene. J Nutr 121, 1883-1888, 1991.

5. Hopkins, GJ, Kennedy, TG, and Carroll, KK: Polyunsaturated fatty acidsas promoters of mammary carcinogenesis induced in Sprague-Dawleyrats by 7,12-dimethylbenz[a]anthracene. JNCI 66, 517-522, 1981.

6. Carroll, KK, and Hopkins, GJ: Dietary polyunsaturated fat versus satu-rated fat in relation to mammary carcinogenesis. Lipids 14, 155-158,1979.

7. Cohen, LA: Differing effects of high-fat diets rich in polyunsaturated,monounsaturated or medium-chain saturated fatty acids on rat mam-mary tumor promotion. In Polyunsaturated Fatty Acids and Ei-cosanoids, WEM Lands (ed). Champaign, IL: Am Oil Chem Soc,1987, pp 241-247.

8. Ip, C, Carter, CA, and Ip, MM: Requirement of essential fatty acid formammary tumorigenesis in the rat. Cancer Res 45, 1997-2001, 1985.

9. Tinsley, IJ, Schmitz, JA, and Pierce, DA: Influence of dietary fattyacids on the incidence of mammary tumors in the C3H mouse. CancerRes 41, 1460-1465, 1981.

10. Craig-Schmidt, M, White, MT, Teer, P, Johnson, J, and Lane, HW:Menhaden, coconut and corn oils and mammary tumor incidence inBALB/c virgin female mice treated with DMBA. Nutr Cancer 20,99-106, 1993.

11. Jurkowski, JJ, and Cave, WT: Dietary effects of menhaden oil on thegrowth and membrane lipid composition of rat mammary tumors. JNCI74, 1145-1150, 1985.

12. Braden, LM, and Carroll, KK: Dietary polyunsaturated fat in relationto mammary carcinogenesis in rats. Lipids 21, 285-288, 1986.

13. Hirose, M, Masuda, A, Ito, N, Kamano, K, and Okuyama, H: Effectsof dietary perilla oil, soybean oil and safflower oil on 7, 12-dimethyl-benz[a]anthracene (DMBA)- and 1,2-dimethylhydrazine (DMH)-in-duced mammary gland and colon carcinogenesis in female SD rats.Carcinogenesis 11, 731-735, 1990.

14. Gabor, H, and Abraham, S: Effects of dietary menhaden oil on tumorcell loss and the accumulation of mass of a transplantable mammaryadenocarcinoma in BALB/c mice. JNCI 76, 1223-1229, 1986.

15. Gonzalez, MJ, Scemmel, RA, Gray, JI, Dugan, L, Sheffield, LG, etal.: Effects of dietary fat on growth of MCF-7 and MDA-MB231human breast carcinomas in athymic nude mice: relationship betweencarcinoma growth and lipid peroxidation product levels. Carcinogene-sis 12, 1231-1235, 1991.

16. Karmali, RA, Marsh, J, and Fuchs, C: Effect of -3 fatty acids ongrowth of a rat mammary tumor. JNCI 73, 457-461, 1984.

17. Cave, WT, and Jurkowski, JJ: Comparative effects of 3 and -6dietary lipids on rat mammary tumor development. In Proceedings ofthe American Oil Chemists' Society Short Course on PolyunsaturatedFatty Acids and Eicosanoids, WEM Lands (ed). Champaign, IL: AmOil Chem Soc, 1987, pp 261-266.

18. Bunce, OR, and Abou-El-Ela, SH: Eicosanoid synthesis and ornithinedecarboxylase activity in mammary tumors of rats fed varying levelsand types of n-3 and/or n-6 fatty acids. Prostaglandin Leukot EssentFatty Acids 41, 105-113, 1990.

19. Ip, C, Ip, MM, and Sylvester, P: Relevance of trans fatty acids andfish oil in animal tumorigenesis studies. In Progress in Clinical andBiological Research, C Ip, DF Bin, AE Rogers, and C Mettlin (eds).New York: Liss, 1986, vol 222, pp 283-294.

Nutrition and Cancer 1998

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20. Karmali, RA, Doshi, RU, Adams, L, and Choi, K: Effect of n-3 fattyacids on mammary tumorigenesis. In Advances in Prostaglandin,Thromboxane, and Leukotriene Research, B Samuelsson, R Paoletti,and PW Ramwell (eds). New York: Raven, 1987, vol 17, pp 886-889.

21. Cohen, LA, Chen-Backlund, JY, Sepkovic, DW, and Sugie, S: Effectof varying proportions of dietary menhaden and corn oil on experi-mental rat mammary tumor promotion. Upids 28, 449-456, 1993.

22. Kikugawa, K, Kojima, T, Yamaki, S, and Kosugi, H: Interpretationof the thiobarbituric acid reactivity of rat liver and brain homogenatesin the presence of ferric ion and ethylenediaminetetraacetic acid. AnalBiochem 202, 249-255, 1992.

23. Metcalee, LD, and Schmitz, AA: The rapid preparation of fatty acidesters for gas chromatography. Anal Chem 33, 363-364, 1961.

24. Fritsche, KL, and Johnston, PV: Effect of dietary -linolenic acid ongrowth, metastasis, fatty acid profile and prostaglandin production oftwo murine mammary adenocarcinomas. J Nutr 120, 1601-1609,1990.

25. Rose, DP, and Connolly, JM: Effects of fatty acids and inhibitors ofeicosanoid synthesis on the growth of a human breast cancer cell linein culture. Cancer Res 50, 7139-7144, 1990.

26. Abou-El-Ela, SH, Prasse, KW, Carroll, R, Wade, AE, Dharwadkar,S, et al.: Eicosanoid synthesis in 7,12-dimethylbenz[a]anthracene-in-duced mammary carcinomas in Sprague-Dawley rats fed primrose oil,menhaden oil or corn oil diet. Lipids 23, 948-954, 1988.

27. Karmali, RA: Fatty acids: inhibition. Am J Clin Nutr 45, 225-229,1987.

28. Noguchi, M, Earashi, M, Minami, M, Kinoshita, K, and Miyazaki, I:Effects of eicosapentaenoic and docosahexaenoic acid on cell growthand prostaglandin E and leukotriene B production by a human breastcancer cell line (MDA-MB-231). Oncology 52, 458-464, 1995.

29. Carter, CA, Ip, MM, and Ip, C: A comparison of the effects of theprostaglandin synthesis inhibitors indomethacin and caprofen on 7, 12-dimethylbenz[a]anthracene induced mammary tumorigenesis in ratsfed different amounts of essential fatty acid. Carcinogenesis 10, 1369-1374, 1989.

30. Calviello, G, Palozza, P, Franceschelli, P, and Bartoli, GM: Low-doseeicosapentaenoic or docosahexaenoic acid administration modifiesfatty acid composition and does not affect susceptibility to oxidativestress in rat erythrocytes and tissues. Lipids 32, 1075-1083, 1997.

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