Fig. 1

Transgenerational study design. a Pregnant C57BL/6NTac mice (F0) were fed either a high-fat (HF; n = 10) or control (CON; n = 10) diet. The HF diet was fed to dams from gestational day (GD) 10 to GD 20. All offspring were fed the CON diet after birth for the remainder of the study, including during pregnancies of F1 and F2 generation offspring. b All pups were cross-fostered at birth (postnatal day [PND] 1) to a CON mother to eliminate litter bias. Pups were weighed on PNDs 2 and 3 and weaned on PND 21. Tumorigenesis was initiated on PND 42 by priming female mice with medroxyprogesterone acetate (MPA; 15 mg/kg), followed by oral gavage of 7,12-dimethylbenz[a]anthracene (DMBA; 1 mg/dose/week) for 3 weeks. Tumorigenesis was monitored by palpation once per week starting 3 weeks after final DMBA administration up to 20 weeks post-DMBA. Mammary glands (MGs) and tumors were collected and processed from F1 and F3 offspring unexposed to DMBA on PND 50 for whole mounts and on PND 100 to perform RNA sequencing analysis