Long-term HF–HC–HSD induces hepatic tumor development. Liver mRNA was determined for CD133 (a), Nanog (b) and vimentin (c) by qPCR. Gross images of tumors that were detected in two of the mice and tumor histopathology (d). Detectable liver p53 protein expression was observed by western blotting after 27 and 49 weeks of HF–HC–HSD (e). Low but detectable AFP was observed in liver tumor samples after 49 weeks of HF–HC–HSD on immunohistochemistry (f). *p < 0.01, #p = 0.03, %p = 0.05 versus chow fed controls, n = 6–12/group.