4.1. Mice and Diets

Wild-type C57BL/6J female mice were generated at the mouse facility of the European Institute of Oncology, Milan, and maintained at the University of Milan in pathogen-free, controlled temperature, and relative humidity and light conditions (21 ± 1 °C, 50 ± 10%, 12 h light/12 h dark cycle). Food and tap water were supplied ad libitum, except for the CR cohort, which received a daily amount of a standard diet formula (4RF21, Mucedola srl, Settimo Milanese, Italy) sufficient to provide 70% of normal caloric intake.

Special diets were produced by replacing the maize content (29%) in the standard diet (4RF21, Mucedola srl, Settimo Milanese, Italy) with powder from seeds of the anthocyanin-rich B1 Pl1 hybrid (Red diet, RD) or the near-isogenic anthocyanin-free b1 pl1 yellow hybrid (Yellow diet, YD). The RD contained about 0.21 ± 0.01 mg/g anthocyanins. The SD, YD, and RD were equal in energy content (3.8 kcal/g), with 53.50% carbohydrate, 18.50% protein, 3% fat, and 6% fiber (Mucedola srl). The HF diet contained 20% protein, 20% carbohydrate, and 60% fat, with an energy content of 5.24 kcal/g (D12492, Research Diets). The body weight of mice and food consumption were controlled weekly.