Consumption of high ω-3 fatty acid diet suppressed prostate tumorigenesis in C3(1) Tag mice

Abstract

Prostate cancer incidence and mortality are high in the Western world and high ω-6/ω-3 PUFA in the Western diet may be a contributing factor. We investigated whether changing from a diet that approximates ω-6 fat content of the Western diet to a high ω-3 fat diet at adulthood might reduce prostate cancer risk. Female SV 129 mice that had consumed a high ω-6 diet containing corn oil for 2 weeks were bred with homozygous C3(1)Tag transgenic male mice. All male offspring were weaned to the corn oil diet (CO) until postpuberty when half of the male offspring were transferred to a high ω-3 diet containing canola oil and fish oil concentrate (FS). High ω-3 diet increased ω-3 and decreased ω-6 fat content of mice tissues. Average weights of prostate and genitourinary bloc were significantly lower in mice consuming high ω-3 diet at adulthood (CO-FS) than mice fed a lifetime high ω-6 diet (CO-CO). There was slower progression of tumorigenesis in dorsalateral prostate of CO-FS than in CO-CO mice. CO-FS mice had slightly lower plasma testosterone level at 24 and 40 weeks, significantly lower estradiol level at 40 weeks and significantly less expressed androgen receptor (AR) in the dorsalateral prostate at 40 weeks than CO-CO mice. Consumption of high ω-3 diet lowered the expression of genes expected to increase proliferation and decrease apoptosis in dorsalateral prostate. Our results suggest that consumption of high ω-3 diet slows down prostate tumorigenesis by lowering estradiol, testosterone and AR levels, promoting apoptosis and suppressing cell proliferation in C3(1)Tag mice.

Fig. 1.

Effect of diets on average body weight gain, prostate weight and GU weight between CO–CO and CO-FS mice. (A) Average body weight gain. Data are from measurements of body weight gain for individual mouse from the time of weaning to 40 weeks of age. Analysis by t-test showed no significant difference in body weight gain between the two groups at P < 0.05. n = 9/group. Bar is mean of body weight gain per week. (B) Prostate weight at 24 and 40 weeks of age. Mouse DL, anterior prostate and ventral prostate lobes were dissected and weighed, and the sums were expressed as milligrams per 25 grams body weight. The average weight was significantly different between the two groups at 24 weeks (n = 15–19/group) and 40 weeks (n = 12/group) by t-test and at P < 0.05. Average prostate weight significantly increased in both groups with time by t-test at P < 0.05. C and D) GU weight at 24 and 40 weeks of age. Mouse GU were dissected at euthanasia and weighed. The weights were expressed as milligrams per 25 grams body weight. The average GU weight was not significantly different between the two groups at 24 weeks but significantly lower at 40 weeks in the CO-FS group than in the CO–CO group by t-test and at P < 0.05 (n = 19/group). Average GU weight increased significantly in CO–CO group with time. *Significantly different between groups, ^†significantly different across time at P < 0.05. Arrow bars represent SEM.

Fig. 2.

Effect of diets on the incidence of PIN and ADC lesions in the DL prostate. (A–D) Representative hematoxylin- and eosin-stained sections from 24 to 40 weeks old mice. A and C illustrate the presence of LGPIN and HGPIN and ADC lesions as demonstrated by epithelial cells proliferation, hyperchromasia, nuclear enlargement and membrane irregularity. (B and D) Consumption of high ω-3 diet resulted in marked reduction of epithelial cells proliferation, hyperchromasia, nuclear enlargement and membrane irregularity. (E) Incidence of PIN lesions and ADC in CO–CO and CO-FS groups at 24 and 40 weeks of age. Prostate specimens were histologically examined for lesions by two independent evaluators and scores were averaged (n = 11–12/group). Pictures were taken at ×40 magnification (Star indicates LGPIN; arrow indicates HGPIN; arrow head indicates ADC. *^{, †, ‡} represent significant difference in LGPIN, HGPIN and ADC, respectively, between CO–CO and CO-FS group at P < 0.05).

Fig. 3.

Effect of diets on plasma testosterone and estradiol levels. Mice from both diet groups were killed at 24 and 40 weeks of age. Plasma was obtained for individual assay from blood collected from mice. (A) The level of testosterone was slightly less in mice from CO-FS group than mice from CO–CO group at 24 and 40 weeks (n = 12–19/group) and increased slightly as mice progressed in age in the CO–CO group (Mann–Whitney test at P < 0.05). (B) Expression of AR was lower significantly in the CO-FS group than the CO–CO group at 40 weeks by t-test, n = 5/group. A total of 1000 epithelial cells within viable areas of lesions per slide were counted for AR expression assay. (C) Estradiol levels were significantly different between the two groups at 24 and 40 weeks by Mann–Whitney test at P < 0.05 (n= 15–20/group). Estradiol level increased slightly as mice progressed in age in CO–CO group. Estradiol level was significantly less in CO-FS group than CO–CO group at 40 weeks and decreased significantly as mice progressed in age in CO-FS group. *Significantly different at P < 0.05, ^†significantly different across time at P < 0.05. Arrow bars represent SEM.

Fig. 4.

Effect of diets on proteins in NF-κB and apoptosis pathways in DL prostate. (A) Protein expression of NF-κBp65, IKα, IKKβ and NOS2 in the DL prostate and their corresponding densitometric analyses in 24 weeks old mice. Expression of IKα protein was slightly higher, whereas the expressions of NF-κBp65, IKKβ and NOS2 proteins were significantly lower in DL prostate of mice that consumed high ω-3 fat diet than mice that consumed high ω-6 fat diet. (B) Protein expression of BCL2 in the DL prostate and the densitometric analysis in 24 weeks old mice. Expression of BCL2 protein was lower significantly in the DL prostate of mice that consumed high ω-3 fat diet compared with mice that consumed high ω-6 fat diet. Equal loading of protein in the lanes was confirmed by β-actin. Values are relative density corrected for β-actin. *Significantly different at P < 0.05, n = 3–5/group. Arrow bars represent SD or SEM where applicable.

Fig. 5.

Effect of diets on the extent of proliferation and apoptosis in DL prostate. DL prostate sections from the two groups were assessed for proliferation by immunohistochemistry analysis of PCNA at 24 and 40 weeks (A–D). Proliferation was significantly less in mice that consumed high ω-3 diet compared with mice that consumed high ω-6 diet by t-test at P < 0.05 (E). Apoptosis was assessed by morphological identification of apoptotic cells in hematoxylin- and eosin-stained sections (F–I) and was significantly greater in mice that consumed high ω-3 diet compared with mice that consumed high ω-6 diet by t-test at P < 0.05 (J). A total of 1000 epithelial cells within viable areas of lesions per slide were counted for proliferation and apoptosis assays. n = 5/group for proliferation assay, n = 7/group for apoptosis assay. Pictures were taken at ×40 magnification. Arrow indicates apoptotic cells. Arrow bars represent SEM. *Significantly different at P < 0.05.