Dietary Modulation of Hepatocarcinoma in Mice

C57Bl/6 male mice were obtained from the Centro de Investigaciones Biológicas (CIB-CSIC) (Madrid, Spain). To induce hepatocarcinoma (Ethic code, Proex 220/17), the animals were randomly distributed into four different groups (n = 6 per group) under a standard (groups 1 and 2) or a high-fat diet (groups 3 and 4) (AIN93G mod. HF 43 kcal% fat, irradiated, Ssniff spezialdiäten gmbh): groups 1 and 3 received a saline solution as the ‘vehicle’ (control); groups 2 and 4 received SETIs (reconstituted lyophilisates).
HCC was induced by a combined treatment with diethynitrosamine (DEN)/thioacetamide (TAA) [3 (link),7 (link),8 (link)]: DEN (20 mg/kg, i.p.), given 3 times per week (for 2 weeks), and TAA (saturated solution—0.5 mL/kg i.g., dissolved in PBS—dose equivalent to 80 mg/kg) administered 3 times per week (for 8 weeks). The mice were sacrificed 7 weeks after the initial DEN injection (Figure 1).
Changes in body weight (BW) and food consumption were monitored every two days. After treatment, the mice were sacrificed by cervical luxation. Whole blood samples were preserved in EDTA-treated tubes (at room temperature) for analyses. Different sections (30–50 mg) of the liver were immersed in RNA later buffer (Qiagen, Hilden, Germany), Krebs’s buffer, and RIPA buffer and kept at −80 °C until analysis.