Antitumor Effects of an Anthocyanin-Rich Grain Diet in a Mouse Model of Lewis Lung Carcinoma

Abstract

Functional foods enriched with plant polyphenol anthocyanins attract particular attention due to their health-promoting properties, including antitumor activity. We evaluated the effects of a grain diet rich in anthocyanins in a mouse model of Lewis lung carcinoma. Mice of the C57BL/6 strain were fed with wheat of near-isogenic lines differing in the anthocyanin content for four months prior to tumor transplantation. Although a significant decrease in the size of the tumor and the number of metastases in the lungs was revealed in the groups with both types of grain diet, the highest percentage of animals without metastases and with attenuated cell proliferation in the primary tumor were observed in the mice with the anthocyanin-rich diet. Both grain diets reduced the body weight gain and spleen weight index. The antitumor effects of the grain diets were associated with the activation of different mechanisms: immune response of the allergic type with augmented interleukin(IL)-9 and eotaxin serum levels in mice fed with control grain vs. inhibition of the IL-6/LIF system accompanied by a decrease in the tumor-associated M2 macrophage marker arginase 1 gene mRNA levels and enhanced autophagy in the tumor evaluated by the mRNA levels of Beclin 1 gene. Thus, anthocyanin-rich wheat is suggested as a promising source of functional nutrition with confirmed in vivo antitumor activity.

Keywords: autophagy; bioflavonoids; blood serum; cancer; cytokines; functional food; metastases; mice; tumor-associated macrophages; wheat grain.

Figure 1

Effects of the type of diet on the body weight gain (a) and spleen index (b) in mice. Data are presented as the mean ± S.E.M. of the values obtained in an independent group of animals (n = 8–10 per group). Statistically significant differences: ^a p < 0.05, ^aa p < 0.01, ^aaa p < 0.001 vs. values of a respective group in a previous month; ^cc p < 0.01, ^ccc p < 0.001 vs. values of a respective group in the month before a previous month; ^b p < 0.05, ^bbb p < 0.001 vs. groups given a standard diet; ^^^ p < 0.001 vs. Intact group; *** p < 0.001 vs. “LLC_St. diet” group.

Figure 2

Effects of the type of diet on the tumor weight (a), number of metastases in the lungs (b), and percentage of mice with metastases (c) in an LLC model. Data are presented as the mean ± S.E.M. (a,b) or share of animals with metastases (c) of the values obtained in an independent group of animals (n = 21–24 per group). Statistically significant differences: * p < 0.05, ** p < 0.01, *** p < 0.001 vs. “LLC_St. diet” group; # p < 0.05 vs. “LLC_CGr” group.

Figure 3

Effects of the type of diet on the expression of Ki67 in the tumor tissue of mice in an LLC model. (a) Quantitative results. Data are presented as the mean ± S.E.M. of the values obtained in an independent group of animals (n = 4–5 per group). Statistically significant differences: ** p < 0.01 vs. “LLC_St. diet” group; # p < 0.05 vs. “LLC_CGr” group. (b) Ki67 immunofluorescence in the tumor tissue. Alexa Flur 488 secondary antibodies were used for staining the samples. The fluorescence images were finally obtained by an Axioplan 2 microscope. Magnification: ×200; scale bar: 100 μm.

Figure 4

Effects of the type of diet on the serum levels of IL-6 (a) and the percentage of mice with undetectable serum levels of LIF (b) in an LLC model. Data are presented as the median (Q1; Q3) (a) or share of animals with metastases (b) of the values obtained in an independent group of animals (n = 6–9 per group). Statistically significant differences: ^^ p < 0.01, ^^^ p < 0.001 vs. Intact group; * p < 0.05 vs. “LLC_St. diet” group; # p < 0.05 vs. “LLC_CGr” group.

Figure 5

Effects of the type of diet on the mRNA levels of Arg1 (a), Nos2 (b), and Becn1 (c) in the tumor tissue of mice in an LLC model. Data are presented as the median (Q1; Q3) of the values obtained in an independent group of animals (n = 9–10 per group). Statistically significant differences: * p < 0.05, ** p < 0.01 vs. “LLC_St. diet” group.