Nutrition and Cancer
ISSN: 0163-5581 (Print) 1532-7914 (Online) Journal homepage: http://www.tandfonline.com/loi/hnuc20
The effect of various dietary fats on skin tumor initiation Mary Locniskar , Martha A. Belury , Adam G. Cumberland , Kelly E. Patrick & Susan M. Fischer To cite this article: Mary Locniskar , Martha A. Belury , Adam G. Cumberland , Kelly E. Patrick & Susan M. Fischer (1991) The effect of various dietary fats on skin tumor initiation, Nutrition and Cancer, 16:3-4, 189-196, DOI: 10.1080/01635589109514157 To link to this article: http://dx.doi.org/10.1080/01635589109514157
Published online: 04 Aug 2009.
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Date: 06 November 2015, At: 07:04
The Effect of Various Dietary Fats on Skin Tumor Initiation Downloaded by [York University Libraries] at 07:04 06 November 2015
Mary Locniskar, Martha A. Belury, Adam G. Cumberland, Kelly E. Patrick, and Susan M. Fischer
Abstract The type of dietary fat has been shown to modulate the initiation stage of mammary tumorigenesis, with saturated fat fed before and/or during carcinogen treatment resulting in increased tumor incidence. This study was designed to determine whether different types of dietary fat alter the initiation stage of skin carcinogenesis by use of the initiation-promotion mouse skin carcinogenesis model. Sencar mice were divided into three groups and maintained on one of the experimental diets. The AIN-76-based diets consisted of 10% total fat with various types of fat: 8.5% menhaden oil plus 1.5% corn oil, 8.5% coconut oil plus 1.5% corn oil, and 10% corn oil. After three weeks mice were initiated with 10 nmol dimethylbenz[a]anthracene (DMBA). Two weeks later, all mice were switched to a diet containing 5% corn oil. Promotion began four weeks after initiation with twice-weekly application of 1 μg 12-O-tetradecanoylphorbol-13-acetate and continuedfor 12 weeks. No statistically significant differences in kilocalories of food consumed or body weights were observed between diet groups during the study. The final papilloma incidence, yield, and size were not significantly different among the diet groups. In a parallel study, [3H]DMBA binding to epidermal DNA showed no dietary differences. Unlike the mammary carcinogenesis model, these data suggest that the type of fat fed during DMBA initiation had minimal effects on this stage of skin carcinogenesis. (Nutr Cancer 16, 189-196, 1991)
Introduction Studies on the effects of diet and cancer have demonstrated that dietary fat modulates tumorigenesis at several organ sites. Evidence from animal models suggests that the amount and type of dietary fat have effects primarily on the promotional stage of carcinogenesis. For example, corn oil, rich in the essential fatty acid linoleate, fed during promotion enhances M. Locniskar and M. A. Belury are affiliated with the Division of Nutrition, The University of Texas at Austin, Austin, TX 78712. A. G. Cumberland, K. E. Patrick, and S. M. Fischer are affiliated with the Research Division, The University of Texas, M. D. Anderson Cancer Center, Smithville, TX 78957.
Copyright © 1991, Lawrence Erlbaum Associates, Inc.
Downloaded by [York University Libraries] at 07:04 06 November 2015
carcinogenesis of the rat mammary gland, with 4.4% linoleic acid being required for maximum tumor development (1). However, dietary fat has also been shown to alter initiation. By use of a similar mammary model, high levels of saturated fat including coconut oil and lard enhance tumor yield when fed before and/or during the initiation stage (2-4). We previously found that type of dietary fat alters skin tumor promotion. By use of the initiation-promotion mouse skin carcinogenesis model, Sencar mice fed corn oil-rich diets during treatment with the promoter 12-0-tetradecanoylphorbol-13-acetate (TPA) developed fewer tumors than mice fed diets containing saturated fats (5,6). Metabolites of arachidonic acid are thought to be required for the biochemical and histological changes that are associated with phorbol ester tumor promotion. This is based on evidence that TPA activates phospholipase A2 (7) and induces the production of prostaglandin E 2 (8). In addition, anti-inflammatory agents, which inhibit cyclooxygenase and lipoxygenase metabolism, also inhibit the TPA-elicited events and tumor promotion (9). The initiation stage of this skin carcinogenesis model can also be affected by arachidonic acid metabolism. For example, topical application of arachidonic acid before initiation with the carcinogen 7,12-dimethylbenz[cr]anthracene (DMBA) enhances tumor yield by 60% (10). Furthermore, using a dietary model, Birt et al. (11) recently demonstrated that in Sencar mice elevated levels of corn oil inhibit initiation and this inhibition parallels a reduction in the arachidonic acid content of epidermal phosphatidylcholme seen in another strain of mouse, the Swiss Webster, fed the same diet (12). One explanation for these findings is the effect of fatty acids on carcinogen metabolism. The n-6 fatty acid linoleate and the n-3 fatty acids found in fish oil, eicosapentaenoic acid (20:5, n-3) and docosahexaenoic acid (22:6, n-3), are of particular importance in determining the activity of carcinogen-metabolizing hydroxylases (reviewed in Refs. 13 and 14). Although dietary fat alters enzyme content and activity, this effect has been shown to be organ specific. For example, Kwei et al. (15) recently demonstrated that the level of corn oil alters the activity of arylhydrocarbon hydroxylase in the lung and kidney but not in liver. In addition to the effect of fatty acids on the activity of the metabolic enzymes, evidence suggests that polycyclic aromatic hydrocarbon carcinogens, such as DMBA, are metabolized through cooxygenation during the oxidative metabolism of arachidonic acid by prostaglandin endoperoxide synthase (16). It is therefore reasonable to hypothesize that incorporation of specific fatty acids into epidermal phospholipids would alter carcinogen metabolism and thereby alter the initiation process. The studies described in this report were undertaken to evaluate the effect of dietary fat on the initiation stage of tumorigenesis by use of the initiation-promotion mouse skin model in animals given a constant and reasonable amount of coconut oil, a saturated fat (SF); menhaden oil, a polyunsaturated fat containing n-3 fatty acids (PUF/n-3); or corn oil, a PUF containing n-6 fatty acid (PUF/n-6). In addition, the effect of dietary fat on the fatty acid profiles of epidermal phospholipids and DMBA binding to epidermal DNA was assessed. Materials and Methods Animals Weanling outbred female Sencar mice (Hsd:SENCAR BR; Harlan Sprague Dawley, Indianapolis, IN) were randomized and assigned their respective diet groups on arrival for each experiment described below. Mice were ear clipped for identification and housed 10 per cage in climate-controlled quarters (22 ± 1°C at 50% humidity) with 10:14-hour light-dark cycle. Animals were observed daily and individually weighed weekly or biweekly.
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Nutrition and Cancer 1991
Diets
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The three semipurified diets used for these studies were based on the AIN-76 formulation (17) but modified to contain 10% lipid with adequate levels of essential fatty acids and are described in detail elsewhere (6). The three fats used were coconut oil (Dyets, Bethlehem, PA), menhaden oil (Zapata-Haynie, Reedville, VA), and corn oil (Best Foods, Union, NJ). The fat composition of the experimental diets is shown in Table 1 and consisted of 8.5% coconut oil plus 1.5% corn oil (SF), 8.5% menhaden oil and 1.5% corn oil (PUF/n-3), and 10% corn oil (PUF/n-6). A diet containing 5% corn oil was prepared and used only during the promotion period of the tumor study. Diets were made on the premises biweekly with components purchased from Dyets, and butylated hydroxytoluene at 0.01% of the fat content was added to the diets to retard lipid peroxidation. Fresh diet was furnished every two days in clean powder feeders with stainless steel grids and lids to reduce spilling, and mice were allowed free access to food and water. Estimates of food consumption for each diet group were determined before DMBA and TPA treatment. Tumor Study Two hundred twenty-five weanling Sencar mice weighing 21.9 ± 3 (SD) g were randomized and assigned to one of the diet groups. They were then fed one of the experimental diets for three weeks. Three days before initiation the dorsal sides of all mice were shaved with electric clippers, and only those mice in the resting phase of the hair cycle were used for the study. Mice were treated with one application of the carcinogen DMBA (10 nmol; Sigma Chemical, St. Louis, MO). The mice were switched to the 5% corn oil diet two weeks later and were maintained on this diet for the remainder of the study. Four weeks after initiation, biweekly treatment with the promoter TPA (1 /tg in 200 y\ acetone per application) was begun (30 mice in each diet group) and continued for 16 weeks. A second group of initiated mice was treated twice weekly with 200 y\ acetone only (15 mice per diet). Mice were observed daily and papilloma incidence and multiplicity assessed weekly. Mice were not carried to carcinoma development because for this initiation-promotion mouse skin carcinogenesis model both end points (papilloma and carcinoma) usually parallel one another. Fatty Acid Analysis of Epidermal Phospholipids Mice were maintained on one of the experimental diets for four weeks. As described in detail previously (5), total lipids were extracted from epidermis, phospholipids were Table 1. Fat Composition of Experimental Diets" Diet 6
Fat Source
Corn oil Menhaden oil Coconut oil Saturated* Monosaturatedd Polyunsaturatedrf
SF C
1.5
8.5 88.7 3.4 7.5
PUF/n-3
PUF/n-6
1.5 8.5
10
28.4 24.5 34.8
12.2 26.0 61.4
a: Diets were prepared on the premises every 2 wks and stored 5 mm. A similar distribution was seen for the other diet groups (data not shown). Fatty Acid Analysis of Epidermal Phospholipids The effect of three types of dietary fat on the fatty acid profiles of phosphatidylcholine and phosphatidylethanolamine is shown in Table 2. As expected, eicosapentaenoic acid (20:5) and docosahexaenoic acid (22:6) in both phospholipid fractions were increased in the PUF/n-3 diet group. In addition, linoleate (18:2) was dramatically elevated in the phosphatidylethanolamine fraction of the PUF/n-6 diet group, and arachidonic acid tended to be reduced in the phosphatidylcholine fraction of the mice fed PUF/n-3. DMBA Binding to DNA As shown in Table 3, covalent binding of radiolabeled DMBA to DNA at either the 15- or 24-hour time point was not different among the diet groups. 100
Figure 2. Effect of diet on papilloma incidence in Sencar mice. Groups of 30 Sencar mice were fed 1 of the 3 experimental diets for 3 wks and initiated with 10 nmol DMBA. After 2 wks, mice were switched to the 5% corn oil diet and after an additional 2 wks were promoted twice weekly with 12-0-tetradecanoylphorbol13-acetate. Tumors were counted weekly, and incidence was calculated as percentage of animals bearing tumors in each diet group. Cumulative probability of observing a papilloma was not different among the groups (pB = 0.5359,pm.c = 0.5291). Symbols as in Figure 1.
Vol. 16, Nos. 3 & 4
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Figure 3. Effect of diet on papilloma yield in Sencar mice. Tumor yield was calculated as avg no. of papillomas per mouse for each diet group. Standard error is indicated for Week 21. Symbols as in Figure 1.
Table 2. Fatty Acid Profile of Epidermal Phosphatidylcholine and Phosphatidylethanolamine0 Diet SF 16:0 16:1 18:0 18:1 18:2 20:4 20:5 22:6 16:0 16:1 18:0 18:1 18:2 20:4 20:5 22:6
37 ± 7* 7 ± 4 10 ± 1 24 ± 3 10 ± 3 10 ± 5 NDC 3 ± 2 35 7 8 35 5 5
ISSN: 0163-5581 (Print) 1532-7914 (Online) Journal homepage: http://www.tandfonline.com/loi/hnuc20
The effect of various dietary fats on skin tumor initiation Mary Locniskar , Martha A. Belury , Adam G. Cumberland , Kelly E. Patrick & Susan M. Fischer To cite this article: Mary Locniskar , Martha A. Belury , Adam G. Cumberland , Kelly E. Patrick & Susan M. Fischer (1991) The effect of various dietary fats on skin tumor initiation, Nutrition and Cancer, 16:3-4, 189-196, DOI: 10.1080/01635589109514157 To link to this article: http://dx.doi.org/10.1080/01635589109514157
Published online: 04 Aug 2009.
Submit your article to this journal
Article views: 3
View related articles
Citing articles: 6 View citing articles
Full Terms & Conditions of access and use can be found at http://www.tandfonline.com/action/journalInformation?journalCode=hnuc20 Download by: [York University Libraries]
Date: 06 November 2015, At: 07:04
The Effect of Various Dietary Fats on Skin Tumor Initiation Downloaded by [York University Libraries] at 07:04 06 November 2015
Mary Locniskar, Martha A. Belury, Adam G. Cumberland, Kelly E. Patrick, and Susan M. Fischer
Abstract The type of dietary fat has been shown to modulate the initiation stage of mammary tumorigenesis, with saturated fat fed before and/or during carcinogen treatment resulting in increased tumor incidence. This study was designed to determine whether different types of dietary fat alter the initiation stage of skin carcinogenesis by use of the initiation-promotion mouse skin carcinogenesis model. Sencar mice were divided into three groups and maintained on one of the experimental diets. The AIN-76-based diets consisted of 10% total fat with various types of fat: 8.5% menhaden oil plus 1.5% corn oil, 8.5% coconut oil plus 1.5% corn oil, and 10% corn oil. After three weeks mice were initiated with 10 nmol dimethylbenz[a]anthracene (DMBA). Two weeks later, all mice were switched to a diet containing 5% corn oil. Promotion began four weeks after initiation with twice-weekly application of 1 μg 12-O-tetradecanoylphorbol-13-acetate and continuedfor 12 weeks. No statistically significant differences in kilocalories of food consumed or body weights were observed between diet groups during the study. The final papilloma incidence, yield, and size were not significantly different among the diet groups. In a parallel study, [3H]DMBA binding to epidermal DNA showed no dietary differences. Unlike the mammary carcinogenesis model, these data suggest that the type of fat fed during DMBA initiation had minimal effects on this stage of skin carcinogenesis. (Nutr Cancer 16, 189-196, 1991)
Introduction Studies on the effects of diet and cancer have demonstrated that dietary fat modulates tumorigenesis at several organ sites. Evidence from animal models suggests that the amount and type of dietary fat have effects primarily on the promotional stage of carcinogenesis. For example, corn oil, rich in the essential fatty acid linoleate, fed during promotion enhances M. Locniskar and M. A. Belury are affiliated with the Division of Nutrition, The University of Texas at Austin, Austin, TX 78712. A. G. Cumberland, K. E. Patrick, and S. M. Fischer are affiliated with the Research Division, The University of Texas, M. D. Anderson Cancer Center, Smithville, TX 78957.
Copyright © 1991, Lawrence Erlbaum Associates, Inc.
Downloaded by [York University Libraries] at 07:04 06 November 2015
carcinogenesis of the rat mammary gland, with 4.4% linoleic acid being required for maximum tumor development (1). However, dietary fat has also been shown to alter initiation. By use of a similar mammary model, high levels of saturated fat including coconut oil and lard enhance tumor yield when fed before and/or during the initiation stage (2-4). We previously found that type of dietary fat alters skin tumor promotion. By use of the initiation-promotion mouse skin carcinogenesis model, Sencar mice fed corn oil-rich diets during treatment with the promoter 12-0-tetradecanoylphorbol-13-acetate (TPA) developed fewer tumors than mice fed diets containing saturated fats (5,6). Metabolites of arachidonic acid are thought to be required for the biochemical and histological changes that are associated with phorbol ester tumor promotion. This is based on evidence that TPA activates phospholipase A2 (7) and induces the production of prostaglandin E 2 (8). In addition, anti-inflammatory agents, which inhibit cyclooxygenase and lipoxygenase metabolism, also inhibit the TPA-elicited events and tumor promotion (9). The initiation stage of this skin carcinogenesis model can also be affected by arachidonic acid metabolism. For example, topical application of arachidonic acid before initiation with the carcinogen 7,12-dimethylbenz[cr]anthracene (DMBA) enhances tumor yield by 60% (10). Furthermore, using a dietary model, Birt et al. (11) recently demonstrated that in Sencar mice elevated levels of corn oil inhibit initiation and this inhibition parallels a reduction in the arachidonic acid content of epidermal phosphatidylcholme seen in another strain of mouse, the Swiss Webster, fed the same diet (12). One explanation for these findings is the effect of fatty acids on carcinogen metabolism. The n-6 fatty acid linoleate and the n-3 fatty acids found in fish oil, eicosapentaenoic acid (20:5, n-3) and docosahexaenoic acid (22:6, n-3), are of particular importance in determining the activity of carcinogen-metabolizing hydroxylases (reviewed in Refs. 13 and 14). Although dietary fat alters enzyme content and activity, this effect has been shown to be organ specific. For example, Kwei et al. (15) recently demonstrated that the level of corn oil alters the activity of arylhydrocarbon hydroxylase in the lung and kidney but not in liver. In addition to the effect of fatty acids on the activity of the metabolic enzymes, evidence suggests that polycyclic aromatic hydrocarbon carcinogens, such as DMBA, are metabolized through cooxygenation during the oxidative metabolism of arachidonic acid by prostaglandin endoperoxide synthase (16). It is therefore reasonable to hypothesize that incorporation of specific fatty acids into epidermal phospholipids would alter carcinogen metabolism and thereby alter the initiation process. The studies described in this report were undertaken to evaluate the effect of dietary fat on the initiation stage of tumorigenesis by use of the initiation-promotion mouse skin model in animals given a constant and reasonable amount of coconut oil, a saturated fat (SF); menhaden oil, a polyunsaturated fat containing n-3 fatty acids (PUF/n-3); or corn oil, a PUF containing n-6 fatty acid (PUF/n-6). In addition, the effect of dietary fat on the fatty acid profiles of epidermal phospholipids and DMBA binding to epidermal DNA was assessed. Materials and Methods Animals Weanling outbred female Sencar mice (Hsd:SENCAR BR; Harlan Sprague Dawley, Indianapolis, IN) were randomized and assigned their respective diet groups on arrival for each experiment described below. Mice were ear clipped for identification and housed 10 per cage in climate-controlled quarters (22 ± 1°C at 50% humidity) with 10:14-hour light-dark cycle. Animals were observed daily and individually weighed weekly or biweekly.
190
Nutrition and Cancer 1991
Diets
Downloaded by [York University Libraries] at 07:04 06 November 2015
The three semipurified diets used for these studies were based on the AIN-76 formulation (17) but modified to contain 10% lipid with adequate levels of essential fatty acids and are described in detail elsewhere (6). The three fats used were coconut oil (Dyets, Bethlehem, PA), menhaden oil (Zapata-Haynie, Reedville, VA), and corn oil (Best Foods, Union, NJ). The fat composition of the experimental diets is shown in Table 1 and consisted of 8.5% coconut oil plus 1.5% corn oil (SF), 8.5% menhaden oil and 1.5% corn oil (PUF/n-3), and 10% corn oil (PUF/n-6). A diet containing 5% corn oil was prepared and used only during the promotion period of the tumor study. Diets were made on the premises biweekly with components purchased from Dyets, and butylated hydroxytoluene at 0.01% of the fat content was added to the diets to retard lipid peroxidation. Fresh diet was furnished every two days in clean powder feeders with stainless steel grids and lids to reduce spilling, and mice were allowed free access to food and water. Estimates of food consumption for each diet group were determined before DMBA and TPA treatment. Tumor Study Two hundred twenty-five weanling Sencar mice weighing 21.9 ± 3 (SD) g were randomized and assigned to one of the diet groups. They were then fed one of the experimental diets for three weeks. Three days before initiation the dorsal sides of all mice were shaved with electric clippers, and only those mice in the resting phase of the hair cycle were used for the study. Mice were treated with one application of the carcinogen DMBA (10 nmol; Sigma Chemical, St. Louis, MO). The mice were switched to the 5% corn oil diet two weeks later and were maintained on this diet for the remainder of the study. Four weeks after initiation, biweekly treatment with the promoter TPA (1 /tg in 200 y\ acetone per application) was begun (30 mice in each diet group) and continued for 16 weeks. A second group of initiated mice was treated twice weekly with 200 y\ acetone only (15 mice per diet). Mice were observed daily and papilloma incidence and multiplicity assessed weekly. Mice were not carried to carcinoma development because for this initiation-promotion mouse skin carcinogenesis model both end points (papilloma and carcinoma) usually parallel one another. Fatty Acid Analysis of Epidermal Phospholipids Mice were maintained on one of the experimental diets for four weeks. As described in detail previously (5), total lipids were extracted from epidermis, phospholipids were Table 1. Fat Composition of Experimental Diets" Diet 6
Fat Source
Corn oil Menhaden oil Coconut oil Saturated* Monosaturatedd Polyunsaturatedrf
SF C
1.5
8.5 88.7 3.4 7.5
PUF/n-3
PUF/n-6
1.5 8.5
10
28.4 24.5 34.8
12.2 26.0 61.4
a: Diets were prepared on the premises every 2 wks and stored 5 mm. A similar distribution was seen for the other diet groups (data not shown). Fatty Acid Analysis of Epidermal Phospholipids The effect of three types of dietary fat on the fatty acid profiles of phosphatidylcholine and phosphatidylethanolamine is shown in Table 2. As expected, eicosapentaenoic acid (20:5) and docosahexaenoic acid (22:6) in both phospholipid fractions were increased in the PUF/n-3 diet group. In addition, linoleate (18:2) was dramatically elevated in the phosphatidylethanolamine fraction of the PUF/n-6 diet group, and arachidonic acid tended to be reduced in the phosphatidylcholine fraction of the mice fed PUF/n-3. DMBA Binding to DNA As shown in Table 3, covalent binding of radiolabeled DMBA to DNA at either the 15- or 24-hour time point was not different among the diet groups. 100
Figure 2. Effect of diet on papilloma incidence in Sencar mice. Groups of 30 Sencar mice were fed 1 of the 3 experimental diets for 3 wks and initiated with 10 nmol DMBA. After 2 wks, mice were switched to the 5% corn oil diet and after an additional 2 wks were promoted twice weekly with 12-0-tetradecanoylphorbol13-acetate. Tumors were counted weekly, and incidence was calculated as percentage of animals bearing tumors in each diet group. Cumulative probability of observing a papilloma was not different among the groups (pB = 0.5359,pm.c = 0.5291). Symbols as in Figure 1.
Vol. 16, Nos. 3 & 4
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Figure 3. Effect of diet on papilloma yield in Sencar mice. Tumor yield was calculated as avg no. of papillomas per mouse for each diet group. Standard error is indicated for Week 21. Symbols as in Figure 1.
Table 2. Fatty Acid Profile of Epidermal Phosphatidylcholine and Phosphatidylethanolamine0 Diet SF 16:0 16:1 18:0 18:1 18:2 20:4 20:5 22:6 16:0 16:1 18:0 18:1 18:2 20:4 20:5 22:6
37 ± 7* 7 ± 4 10 ± 1 24 ± 3 10 ± 3 10 ± 5 NDC 3 ± 2 35 7 8 35 5 5
